[Federal Register Volume 59, Number 122 (Monday, June 27, 1994)]
[Unknown Section]
[Page 0]
From the Federal Register Online via the Government Publishing Office [www.gpo.gov]
[FR Doc No: 94-13784]


[[Page Unknown]]

[Federal Register: June 27, 1994]


_______________________________________________________________________

Part II





Environmental Protection Agency





_______________________________________________________________________



40 CFR Part 79




Fuels and Fuel Additives Registration Regulations; Final Rule
ENVIRONMENTAL PROTECTION AGENCY

40 CFR Part 79

[FRL-4892-7]
RIN 2060-AC10

 
Fuels and Fuel Additives Registration Regulations

AGENCY: Environmental Protection Agency (EPA).

ACTION: Final rule.

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SUMMARY: This final rule establishes new requirements for the 
registration of designated fuels and fuel additives (F/FAs) as 
authorized by sections 211(b)(2) and 211(e) of the Clean Air Act (CAA). 
The registration requirements are organized within a three-tier 
structure. Tier 1 requires F/FA manufacturers to perform a literature 
search on the health and welfare effects of F/FA emissions, 
characterize the emissions, and provide qualitative exposure 
information. Tier 2 requires biological testing for the examination of 
subchronic systemic and organ toxicity, as well as the assessment of 
specific health effects endpoints. When necessary, Tier 3, which 
includes follow-up studies or other additional tests, may be required. 
The rule permits adequate existing test data to be submitted in lieu of 
conducting new duplicative tests. It also includes special provisions 
for small businesses and certain types of products, and a grouping 
system which permits manufacturers of similar F/FA products to share 
the costs of compliance.

DATES: This regulation is effective May 27, 1994. The incorporation by 
reference of certain publications listed in the regulations is approved 
by the Director of the Federal Register as of June 27, 1994.
    The information collection requirements contained in 40 CFR 79.51, 
79.52, and 79.57 through 79.68 have not been approved by the Office of 
Management and Budget (OMB) and are not effective until OMB has 
approved them. EPA will publish a document in the Federal Register 
announcing OMB approval of the information collection requirements.

ADDRESSES: The record for this rulemaking is contained in Docket No. A-
90-07. The docket is located at the Air Docket, Room M-1500, 401 M 
Street SW., Washington, DC 20460; phone (202) 260-7548 or 7549; fax 
(202) 260-4000. The docket is open for public inspection from 8 a.m. 
until 4 p.m., Monday through Friday. As provided in 40 CFR part 2, a 
reasonable fee may be charged by EPA for photocopying services. 
Electronic copies of major F/FA rulemaking documents can be obtained 
through the Office of Air Quality Planning and Standards (OAQPS) 
Technology Transfer Network Bulletin Board System (TTNBBS). Details on 
how to access TTNBBS are included in Section XIV of this preamble.

FOR FURTHER INFORMATION CONTACT: Regarding F/FA registration 
procedures: James Caldwell (phone 202-233-9303) or Joseph Fernandes 
(phone 202-233-9016). USEPA, OMS Field Operations and Support Division, 
Mail Code 6406J, 401 M Street SW., Washington, DC 20460. Regarding 
technical requirements: Kent Helmer (phone 313-741-7825). USEPA, OMS 
Regulation Development and Support Division, Mail Code RDSD-12, 2565 
Plymouth Road, Ann Arbor, MI 48105.

SUPPLEMENTARY INFORMATION:

Table of Contents

I. Introduction
II. Background
    A. Legal Authority and Statutory History
    B. Public Participation
    C. Additional Information on the Effective Date
III. Overview of Program Requirements
    A. Overall Scope and Approach
    B. Health Evaluation Requirements
    C. Welfare Evaluation Requirements
    D. Requirements for Emission Control System Testing
IV. Grouping System
    A. Objectives and Rationale
    B. Grouping Approach and Criteria
    C. Implementation of Grouping System and Cost-Sharing Provisions
V. Base Fuel Specifications and Formulation Requirements
    A. Gasoline
    B. Diesel
    C. Alternative Fuels
VI. Emission Generation
    A. General Approach
    B. Combustion Emission Generation
    C. Evaporative Emission Generation
    D. Vehicle Selection
    E. Mileage Accumulation
    F. Special Requirements for Additives
VII. Tier 1 Requirements
    A. Literature Search
    B. Characterization of Emissions
    C. Exposure Analysis
VIII. Tier 2 Requirements
    A. General Methodology
    B. Subchronic Inhalation Study and Endpoint Tests
    C. Adequate Endpoint Information in Lieu of Tier 2 Tests
    D. Alternative Tier 2 Provision
IX. Tier 3 Requirements
    A. Scope
    B. Criteria for Referral to Tier 3
    C. Potential Tier 3 Tests
X. Special Provisions
    A. Experimental F/FAs
    B. Relabeled Products
    C. Aerosols
    D. Small Business Provisions
XI. Timing and Compliance Requirements
XII. Reporting Requirements
    A. Basic Registration Data
    B. Summary Report
    C. Appendices
    D. Tier 3 Report
    E. Confidential Business Information
XIII. Administrative Requirements
    A. Administrative Designation and Regulatory Analysis
    B. Regulatory Flexibility Act
    C. Recordkeeping Requirements
XIV. Electronic Availability of Rulemaking Documents

I. Introduction

    Over 2,300 fuels and 4,800 fuel additives were registered by EPA as 
of March 1994 and, to some degree, each of them produces emissions 
which may contribute to potentially harmful air pollution. The primary 
purpose of today's rule is to establish registration requirements which 
will provide information for identifying and evaluating the potential 
adverse effects of designated F/FA emissions and for guiding the 
direction of related regulatory actions in the future as specified in 
section 211 of the CAA.
    Previous actions have implemented CAA sections 211(a) and 
211(b)(1), which govern the general registration of F/FAs, as well as 
CAA section 211(f). Today's rule amends 40 CFR part 79 by adding 
regulatory provisions requiring the testing of F/FAs as a requirement 
for registration, as stipulated in section 211(b)(2) and section 211(e) 
of the CAA.
    In addressing these additional statutory provisions, this rule 
focuses on the identification and evaluation of potential adverse 
health effects associated with F/FA evaporative and combustion 
emissions. The required health effects evaluation is organized in a 
tiered structure, and includes emission characterization, literature 
search, and biological testing requirements. Although this testing 
framework focuses on the evaluation of health effects, F/FA 
manufacturers are also required to perform data searches to obtain 
information on the potential welfare effects of F/FA emissions. In 
addition, EPA will continue to use existing procedures under CAA 
section 211(f) for the evaluation of potential effects of F/FAs on ECS 
performance.
    The ultimate use of the registration information to be submitted in 
compliance with this rule is to guide EPA in potential future 
regulatory actions under CAA section 211(c). Section 211(c) provides 
authority for the possible control or prohibition of any fuel or fuel 
additive whose emission products cause or contribute to air pollution 
which may reasonably be anticipated to endanger the public health or 
welfare. Evidence of adverse effects of F/FA emissions on ECS 
performance, obtained under CAA section 211(f) or from other sources, 
could also be used by EPA to support such regulatory decisions.

II. Background

A. Legal Authority and Statutory History

    The legal authority for the F/FA registration program is provided 
by section 211 of the CAA. Section 211(a), 42 U.S.C. section 7545, 
authorizes EPA to designate any fuel or fuel additive and prohibits 
manufacturers of designated fuels or additives from selling such 
products unless they have been registered by EPA in accordance with CAA 
section 211(b). In 1975, EPA issued regulations (40 CFR part 79) 
implementing basic registration requirements, as stipulated by CAA 
section 211(b)(1), that included: commercial identifying information, 
range of concentration, purpose-in-use, and chemical composition.
    Section 211(b)(2) of the CAA also gives EPA discretionary authority 
to establish additional registration requirements. According to this 
section, EPA ``may also require the manufacturer of any fuel or fuel 
additive to conduct tests to determine potential public health effects 
of such fuel or fuel additive (including, but not limited to, 
carcinogenic, teratogenic, or mutagenic effects),'' and to furnish 
other ``reasonable and necessary'' information to identify F/FA 
emissions and determine their effects on vehicular emission control 
performance and on the public health and welfare. The statute further 
stipulates that testing for health effects is to be conducted according 
to procedures and protocols established by the Administrator, and that 
test results will not be considered confidential. Once the manufacturer 
has completed registration requirements and has given assurances that 
the Agency will be notified of future changes in that information, CAA 
section 211(b)(3) directs the Administrator to register the fuel or 
fuel additive.
    EPA did not exercise its discretionary authority to require testing 
of F/FAs under CAA section 211(b)(2) as part of the general 
registration regulations issued in 1975. However, in the CAA Amendments 
of 1977 (PL 95-95, August 7, 1977), Congress added section 211(e), 
which made implementation of section 211(b)(2) mandatory and contained 
additional provisions requiring the implementation of the regulations 
within one year of enactment of the CAA Amendments. In an effort to 
fulfill this requirement, EPA published an Advanced Notice of Proposed 
Rulemaking (ANPRM) in 1978 (see 43 FR 38607, August 29, 1978; Docket 
ORD-78-01). However, the rulemaking process did not go forward during 
the next ten years and the rule was not finalized. Nevertheless, this 
action remained on EPA's regulatory agenda and a development plan for 
the rulemaking was created in 1988.
    In 1989, a citizens group brought a lawsuit [Thomas v. Browner, 
C.A. No. 89-6269 (D. Oreg. 1989)] challenging EPA's failure to 
promulgate F/FA testing regulations within the one-year deadline 
stipulated in CAA section 211(e). EPA entered into a Consent Decree in 
settlement of this lawsuit which, together with subsequent 
modifications, established the rulemaking schedule. Accordingly, a new 
ANPRM was published on August 7, 1990 (55 FR 32218) and a Notice of 
Proposed Rulemaking (NPRM) was published on April 15, 1992 (57 FR 
13168). Public hearings as well as periods for written commentary 
followed both of these publications. On February 24, 1994, EPA 
published a Notice of Reopening of Comment Period (59 FR 8886) 
requesting public comment on several compliance-related and technical 
issues that needed clarification and/or reconsideration. Today's action 
culminates the rulemaking process by promulgating F/FA registration 
requirements under CAA sections 211(b)(2) and 211(e).

B. Public Participation

    In the months following the publication of the ANPRM (55 FR 32218) 
and the associated public hearing (on September 26, 1990), EPA explored 
the feasibility and appropriateness of applying regulatory negotiation 
procedures (under the provisions of section 583 of the Negotiated 
Rulemaking Act of 1990) to the development of this rule. Interviews and 
meetings were held with representatives of a variety of affected 
industry groups and environmental organizations, to assess their 
interest and willingness to participate in potential negotiations. This 
process indicated that there was insufficient support for regulatory 
negotiation among a number of key parties. A traditional rulemaking 
procedure was then followed to develop this rule.
    Following publication of the proposed rule, EPA held a public 
hearing on May 28, 1992, and accepted comments until June 30, 1992. 
Public response on the NPRM included five oral presentations at the 
hearing and the subsequent submission of 42 written comments. EPA also 
received 13 written comments on the issues discussed in the Notice of 
Reopening of Comment Period (referred to as the ``Reopening Notice'' in 
later portions of this document). A transcript of both public hearings 
and copies of all written comments are available in public Docket No. 
A-90-07.
    A discussion of comments received since the NPRM and EPA's 
responses are included in the ``Summary and Analysis of Comments for 
the Fuels and Fuel Additives Registration Regulations,'' which is 
available in the public docket referenced above. All public commentary 
was carefully considered in developing this final rule. Major areas of 
comment are described in the relevant sections of this preamble.

C. Additional Information on the Effective Date

    The effective date of this rule is May 27, 1994. EPA notes that the 
general requirement (under 5 U.S.C. 553(d), the Administrative 
Procedure Act (APA)), that publication or service of a substantive rule 
be made not less than 30 days before its effective date, does not apply 
here. Under 5 U.S.C. 559, the APA states that a subsequent statute does 
not supersede or modify the APA except to the extent that it does so 
expressly. CAA section 307(d)(1)(E) specifically applies to the 
promulgation or revision of any regulation pertaining to any fuel or 
fuel additive under CAA section 211. CAA section 307(d)(1) further 
provides that ``[t]he provisions of sections 533 through 557 and 
section 706 of title VI shall not, except as expressly provided in this 
subsection, apply to actions to which this subsection applies.'' 
Nowhere does subsection 307(d) expressly provide that section 553(d) of 
title 5 applies.
    Further, CAA section 211(e)(2) expressly provides that the time 
period for providing the ``requisite information'' under section 
211(e)(2) is based on the ``date of promulgation'' of the rule. 
Therefore, the requirements under CAA section 211(e)(2) are effective 
on May 27, 1994. Additionally, even if section 553(d) were to apply to 
the portion of the rule promulgated under the authority of section 
211(b) of the CAA, there is good cause under section 553(d)(3) of the 
APA to provide less than 30 days notice following publication in order 
to simplify implementation of the rule by establishing one effective 
date for the rule's requirements. As discussed in the following 
sections, this final rule provides a six year time period for 
completing Tier 2 testing, commencing on May 27, 1994. EPA believes 
this to be sufficient for the regulated industry to comply with the 
rule and that, given this compliance schedule, a reasonable amount of 
notice is provided for this type of information-gathering regulation. 
Finally, EPA has taken steps to provide notice of this final action to 
the regulated industry upon signature of the rule. For these reasons, 
EPA believes that establishing the effective date as May 27, 1994 is 
reasonable.

III. Overview of Program Requirements

A. Overall Scope and Approach

    The requirements of this rule apply to all types of F/FAs which 
have been designated to be registered by EPA (see Section III.A.3). 
Based on the provisions of CAA section 211(e), the requirements must be 
satisfied both by manufacturers of F/FA products registered at the time 
of promulgation as well as manufacturers of F/FA products seeking 
registration after promulgation. Considering the large number of F/FA 
products to be evaluated and the potential burden of the program on the 
regulated industry, this final rule maintains the grouping system and 
the tiered approach proposed in the NPRM.
    The grouping system allows manufacturers of similar products to 
share the costs of testing. Rather than mandating comprehensive testing 
as a routine registration requirement for every registered fuel and 
fuel additive, the grouping system permits the testing of one product 
as a representative of all relatively similar products (see Section IV 
of this preamble). In addition, the testing program is designed to 
address testing needs on a tiered basis, with allowance for more 
rigorous, resource-intensive requirements contained in each successive 
tier. A detailed description of the scope and requirements of each 
individual tier is provided in Sections VII, VIII, and IX of this 
preamble, and procedures for generating the emissions to be tested are 
explained in Section VI. Additional special provisions to reduce the 
burden on the regulated industry are discussed in Section X.
    EPA believes that the program required by this final rule is 
consistent with the CAA and reflects a reasonable and cost-conscious 
approach to a very complex regulatory area. The following sections 
present a general overview of the main provisions of the rule.
1. Tiered Approach
    As depicted in Figure 1, the registration program's requirements 
are organized within a three-tier structure. In part, each tier is 
intended to function as a screen for determining the need for more 
rigorous requirements in subsequent tiers. Consistent with CAA section 
211(e)(3), which authorizes EPA to avoid duplication of effort, the 
tiered approach permits F/FA manufacturers to use test results and 
other information which may already be available about their products.

    BILLING CODE 6560-50-P

TR27JN94.000


    BILLING CODE 6560-50-C
    a. Content of Tiers. Except as may be modified by any applicable 
special provisions, the requirements of Tiers 1 and 2 are mandatory for 
all fuels and fuel additives. These requirements may be satisfied by 
manufacturers either on an individual basis or by way of a group 
submission consistent with the provisions of the grouping system.
    Under Tier 1, F/FA manufacturers are required to perform a 
literature search on the health and welfare effects of F/FA emissions, 
characterize the emissions, and provide a qualitative exposure analysis 
based on total annual production volume and market distribution data 
(see Section VII for details on Tier 1 requirements). The modeling 
analyses proposed in the NPRM have been eliminated from Tier 1 as 
explained in Section III.C of this preamble. Tier 2 (see Section VIII) 
includes biological testing for specific health effect endpoints, as 
well as general systemic and organ toxicity. The Tier 2 biological 
testing requires the exposure of laboratory animals to the whole 
emissions of fuels or additive/base fuel mixtures.\1\ To the extent 
that previously conducted studies are available which are at least 
comparable to the specified guidelines for the chemical and/or 
biological tests required in Tiers 1 and 2, such existing data may be 
submitted in lieu of performing and reporting on new duplicative tests 
(see Section VII.A.2).
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    \1\An additive must be mixed with the base fuel of its 
associated fuel family prior to generating emissions for testing 
(see Sections IV.A and IV.B.1).
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    The results of Tiers 1 and 2 are to be reported to EPA according to 
the report formats described in Section XII. EPA will evaluate these 
results to determine if additional testing or analysis may be indicated 
under the provisions of Tier 3. For the purpose of peer review during 
this evaluation process, EPA may furnish the submitted data to, and 
consult with, other organizations, such as the Health Effects 
Institute. Tier 3 tests will be determined on a case-by-case basis at 
EPA's discretion, as discussed in Section IX. The specific objectives 
and scope of Tier 3 tests will vary depending on the concerns 
identified in the earlier tiers or any other information available to 
EPA.
    b. Timing of Requirements: Registered F/FAs. EPA proposed to 
require that Tier 1 and Tier 2 data be submitted within three years for 
registered F/FAs. In their comments on the proposal the regulated 
industry suggested that it would not be possible to complete Tier 2 
testing within three years. As detailed in the Summary and Analysis of 
Comments, these commenters stated that the number of F/FAs to be 
tested, the time needed for development of detailed test protocols, and 
the lack of available test facilities were inconsistent with the three-
year time frame for completion of Tier 1 and Tier 2 for all F/FAs (or 
groups). None of the commenters, however, suggested an alternative time 
frame within which the testing of all F/FAs (or groups) could be 
completed.
    EPA has thoroughly considered these comments and, accordingly, has 
taken a number of measures to streamline the program. As discussed 
later in this preamble, such measures include the deletion from the 
final rule of some requirements proposed in the NPRM (e.g., 
quantitative modeling requirements), modification of the Tier 2 testing 
scenario for greater efficiency, simplification of some of the grouping 
rules, and the addition of special provisions which will result in a 
smaller number of F/FA products requiring testing.
    Nevertheless, EPA recognizes that the number of laboratory 
facilities currently available to conduct the required emission-based 
toxicological tests is very limited. EPA expects that the promulgation 
of this rule will create a demand for testing laboratories which will 
encourage the reactivation, modification, and/or expansion of existing 
laboratories, as well as the development of new facilities, to 
accommodate the requirements of the F/FA registration program. However, 
there is likely to be a lag between the demand for and the availability 
of laboratory capacity. Thus, while EPA believes that some groups could 
complete the testing required by the rule in three years, it is likely 
that not all of the F/FAs to be tested could complete the requirements 
in the three-year time frame.
    Considering these factors, the final rule allows a six-year period 
for the conduct of Tier 2 testing for registered F/FAs. This longer 
period will provide the necessary start-up time for laboratories with 
previous experience in conducting studies reasonably similar to those 
required in Tier 2 to adapt or build the necessary facilities, organize 
the key technical personnel, and conduct verification procedures. In 
this regard, it is not necessary for each laboratory providing services 
for this program to employ experts in each of the toxicology specialty 
fields covered by Tier 2. EPA recognizes that some of the required 
expertise may be in short supply, and envisions that laboratories may 
subcontract with subspecialists as needed for evaluation of test 
results. Furthermore, biological laboratories which currently offer 
inhalation toxicology testing services, but are not equipped to 
generate and deliver engine emissions for such testing, should be able 
to obtain the additional equipment and engineering expertise they will 
need in a relatively short period of time. Addition of emission 
generation capabilities by such laboratories will be facilitated by the 
fact that the final rule allows the use of relatively inexpensive and 
possibly portable engine dynamometers for generating the required 
emissions for toxicological testing related both to light-duty and 
heavy-duty engine applications.
    EPA estimates that the necessary toxicology laboratory capacity 
will begin to come on line within 8-15 months of the effective date of 
this rule, with expansion of capacity continuing for an additional 12-
24 months thereafter. The initial period should coincide with the 
start-up time needed by the regulated industry prior to beginning the 
required Tier 2 testing. Manufacturers' start-up activities will 
include review and understanding of the requirements, formation and 
functional organization of groups, acquisition of required test fuel 
supplies, and contracting for data gathering and testing services. EPA 
estimates that these activities can generally be accomplished in 6-12 
months.
    With six years provided for full Tier 2 completion, sufficient time 
should then be available for completion of all Tier 2 testing. However, 
this assumes that the regulated industry will not purposely delay the 
onset of testing. If all F/FA groups wait until the fourth and fifth 
years to begin the Tier 2 testing, it will again become likely that 
some will not be able to complete the requirements on time. In such 
instances, the responsible manufacturers will have failed to comply 
with the requirements of this rule and will be subject to enforcement 
action and/or loss of registration.
    Thus, in the case of registered F/FA products, this final rule 
requires the submission within three years from the effective date of 
all applicable Tier 1 requirements plus either: (1) Submittal of all 
Tier 2 requirements or (2) evidence of a contractual obligation with a 
qualified laboratory to conduct the required Tier 2 tests.\2\ If, 
within the first three years, a contract for Tier 2 is submitted rather 
than the Tier 2 data itself, then the final Tier 2 report is due to EPA 
no later than six years of the effective date of this rule. Both Tiers 
1 and 2 are mandatory. Failure to submit Tier 2 data for a registered 
F/FA within six years of the effective date of this rule will subject 
the manufacturer to enforcement action and/or revocation of the 
registration. In the case of F/FAs for which Tier 2 testing is not 
required because of special provisions, all applicable requirements are 
due to EPA within the initial three-year period after promulgation of 
this rule.
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    \2\To be qualified, a laboratory must be able to perform 
inhalation toxicology tests in compliance with the Good Laboratory 
Practice requirement in this rule, including monitoring by an onsite 
Quality Assurance Unit. It must also be able to properly and safely 
store, transport, and use F/FAs. The study director must be a 
professional scientist with a doctoral degree in toxicology or 
equivalent. Other individuals engaged in the conduct of the studies 
shall have the education, training, and/or experience to enable 
proper performance of the assigned functions. The laboratory's 
animal handling facility must be registered and in good standing 
with the U.S. Department of Agriculture. Accreditation with a 
recognized independent organization which sets laboratory animal 
handling standards [e.g., the American Association for Accreditation 
of Laboratory Animal Care (AAALAC)] is required.
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    Existing F/FA registrations are also conditional on satisfaction of 
any Tier 3 requirements which might be prescribed by the Agency 
pursuant to CAA section 211(b). When Tier 3 testing is prescribed for a 
registered F/FA product, the existing registration will be extended for 
that time which EPA specifies as necessary for completion of the 
additional requirements. Maintenance of registration will depend upon 
satisfactory compliance with these requirements.
    EPA is promulgating Tier 2 testing requirements under the authority 
of both sections 211(b) and 211(e) of the CAA. The requirements for 
Tier 2 testing are all within EPA's discretion under section 211(b). 
Section 211(b) gives the Administrator broad authority ``for the 
purpose of registration of fuels and fuel additives'' to require 
manufacturers ``to conduct tests to determine potential public health 
effects of such fuel or fuel additive.'' EPA interprets ``for the 
purpose of registration'' to encompass both gaining and maintaining 
registration for F/FAs. This interpretation is supported by section 
211(e), which requires implementation of section 211(b) authority with 
respect to both registered and new F/FAs and mandates that EPA require 
testing of F/FAs. The legislative history supports such a view. The 
1977 House Report,\3\ upon which section 211(e) was based, states:

    \3\H. Rept. No. 294, 95th Cong., 1st Sess. 308, reprinted in 
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1977, U.S. CODE CONG. & ADMIN. NEWS 1077, 1387.

    Section 220 of the Committee bill is intended to express the 
Committee's disapproval of EPA's past handling of its authority in 
this area and of its proposed future plans * * * Instead, an 
aggressive, preventative approach to the gathering of necessary 
information is mandated * * *
    The bill mandates the Administrator to promulgate regulations 
within one year after enactment. * * * These regulations must 
require testing by the manufacturer of the fuel or fuel additive, 
except insofar as paragraph (3) otherwise permits.
    All of these requirements are mandatory.

    Tier 2 involves testing ``to determine potential public health 
effects'' of F/FAs and, therefore, is within the type of testing that 
section 211(b) allows EPA to require. The timing requirements of Tier 2 
are also within EPA's discretion under section 211(b), for section 
211(b) leaves submission deadlines to EPA's discretion.
    The timing for Tier 2 submissions is also governed, at least in 
part, by section 211(e)(2). That section requires that, for F/FAs 
registered when the rule is promulgated, ``requisite information'' be 
submitted within three years of the promulgation date. ``Requisite 
information'' is not defined in the statute. EPA proposed to interpret 
``requisite information'' as the data required by Tiers 1 and 2. In 
part this was based on EPA's understanding that Congress intended that 
the testing rule promulgated under section 211(e) would require 
manufacturers to conduct testing--not merely conduct a literature 
search and compile studies that had already been conducted. As a 
practical matter, however, EPA now believes that the Tier 2 tests 
cannot be completed for all F/FAs within three years. EPA believes this 
makes its proposed definition of ``requisite information'' unreasonable 
and requires a different interpretation of ``requisite information.''
    EPA considered redefining ``requisite information'' to mean studies 
that could be completed for all F/FAs within three years. Given the 
time frame, laboratory availability, and the number of groups to be 
tested, EPA was not sure that any meaningful health effects testing 
could be accomplished for all groups within three years. Certainly, 
such testing could not include testing of combustion and evaporation 
products for all groups. As discussed elsewhere, EPA believes that the 
testing of combustion and evaporative emissions included in Tier 2 is 
part of the basic testing necessary to evaluate potential health 
effects, because people are exposed to both combustion and evaporative 
emissions. Therefore, EPA would require this testing under section 
211(b) regardless of whether the testing is required by section 
211(e)(2). If EPA had chosen to interpret ``requisite information'' to 
be testing that could be done in three years, EPA would have, in any 
case, required such testing in addition to the Tier 2 testing required 
by this rule. This would likely have delayed submission of the 
information that EPA believes to be necessary (i.e., the combustion 
emissions testing and evaporative emissions testing).
    In today's rule, EPA interprets ``requisite information'' as either 
data required by Tiers 1 and 2 or data required by Tier 1 and a 
commitment to conduct Tier 2 testing. EPA believes that this meets the 
congressional mandate to require emissions speciation testing and a 
demonstration that manufacturers are making progress in their testing 
by requiring submission of evidence of a contract with a qualified 
laboratory to conduct the Tier 2 studies. In addition, this 
interpretation imposes testing costs on manufacturers only for types of 
studies that EPA believes are necessary and useful, and it requires 
manufacturers to finish testing in a time frame that EPA believes is 
possible to meet.
    Even if ``requisite information'' were interpreted to mean only 
data required by Tier 1, EPA would still impose Tier 2 testing and 
timing requirements as contained in today's rule. In that case, section 
211(e) would require Tier 1 data to be submitted within three years of 
this rule's promulgation, and section 211(b) would provide authority 
for EPA to impose the Tier 2 testing requirements in the time frame set 
forth in this rule.
    c. Timing of Requirements: Registrable and New F/FAs. Consistent 
with section 211(e), for F/FAs not yet registered, all test 
requirements must be satisfied prior to registration, including any 
Tier 3 requirements which EPA judges to be necessary. However, as 
discussed in the Reopening Notice, this final rule makes a distinction 
between ``registrable'' and truly ``new'' F/FA products. Registrable F/
FAs are products\4\ that are not registered as of the effective date of 
this rule but that meet the program's criteria for grouping with a 
currently registered fuel or bulk additive\5\ in the same fuel 
family.\6\ Conversely, a F/FA product not registered as of the 
effective date of this rule is designated as ``new'' if it does not 
meet the program's criteria for grouping with a currently registered 
fuel or bulk additive in the same fuel family. In the above 
definitions, the term ``currently'' refers to the date on which EPA 
receives the basic registration data for the F/FA product in 
question.\7\
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    \4\Registration is product-specific. Thus, if a particular fuel 
or additive product has not been registered by its manufacturer, 
then that manufacturer does not have the right to introduce, market, 
and/or sell this product, even if a compositionally similar or 
identical product has been registered by another manufacturer.
    \5\A ``bulk additive,'' sometimes called a ``general use'' 
additive, is defined as a product added to fuel at the refinery as 
part of the original blending stream or after the fuel is 
transported from the refinery, but before the fuel is purchased for 
introduction into the fuel tank of a motor vehicle. In contrast, an 
``aftermarket additive,'' sometimes called a ``consumer additive,'' 
is an additive product which is added by the end-user directly to 
fuel in a motor vehicle or engine to modify the performance or other 
characteristics of the fuel, the engine, or its emissions.
    \6\``Fuel family'' refers to the primary categorization of F/FAs 
in the grouping system of this registration program, as described in 
Section IV. A fuel family is defined as a set of F/FAs which share 
basic chemical and physical formulation characteristics and can be 
used in the same engine or vehicle. In the definition of 
``registrable'', the restriction ``in the same fuel family'' means 
that the similarity of an applicant F/FA product to a bulk additive 
currently registered for use in another fuel family will not suffice 
to make the applicant F/FA product registrable. This restriction is 
consistent with the general principles of the grouping system, which 
permits grouping of F/FAs only within the defined fuel families.
    \7\Revision of an existing registration (e.g., addition or 
deletion of a currently-registered bulk additive to an existing fuel 
registration) does not constitute a new registration (assuming any 
added bulk additive is registered for use in fuels of the same 
type). However, test requirements may change if the revision causes 
the fuel product to change from one group to another.
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    For registrable products, similar testing and compliance 
requirements apply as those pertaining to currently registered F/FAs in 
the same group. Upon the manufacturer's submittal of the basic 
registration data and other pre-Tier 1 application requirements for a 
registrable product, registration will be granted by EPA. Once 
registered, these products will be legally able to enter the market. 
The manufacturer will have the same period of time after the effective 
date of this rule for the completion of Tiers 1 and 2 as the applicable 
group of existing F/FAs, and can satisfy these requirements either by 
joining the existing group or by testing individually. On the other 
hand, manufacturers of new F/FA products (i.e., F/FAs not registered as 
of the effective date of this rule and not fitting the registrable 
criteria) are required to submit all testing requirements prior to 
registration, including Tier 3 when prescribed by the Agency. Thus, if 
EPA identifies a need for additional testing at the Tier 3 level for a 
new F/FA product, registration will not occur until satisfactory 
completion of all such requirements.
    As discussed in the NPRM and in the Reopening Notice, EPA 
interprets CAA section 211(b) in conjunction with CAA section 211(c), 
which gives EPA authority to control or prohibit the manufacture, 
introduction into commerce, offering for sale, or sale of any fuel or 
fuel additive if the Administrator finds that the emission products of 
such fuel or fuel additive ``cause[s], or contribute[s], to air 
pollution which may reasonably be anticipated to endanger the public 
health or welfare.'' In light of this responsibility, EPA believes that 
it should exercise particular caution in registering new F/FA products 
and that it should have the necessary information to evaluate fully the 
potential public health consequences of such new F/FAs prior to 
allowing their introduction into the market. Thus, before granting 
registration to manufacturers of new F/FAs, under the authority of CAA 
section 211(b), this final rule requires that they comply with all 
testing requirements.
    Figure 2 summarizes the decision process for determining whether an 
unregistered F/FA product (i.e., a F/FA product not registered as of 
the effective date of this rule) is ``registrable'' and thus handled 
much like a currently registered F/FA product, or whether an 
unregistered F/FA product is ``new'' and must complete all testing 
requirements before registration is granted.

    BILLING CODE 6560-50-P

TR27JN94.001


    BILLING CODE 6560-50-C
    An unregistered F/FA product which meets the criteria for grouping 
only with a currently registered aftermarket additive (and not with a 
currently registered fuel and/or bulk additive) is not designated as 
registrable. This does not necessarily preclude an unregistered 
aftermarket additive from being registrable (since aftermarket 
additives can group with fuels and bulk additives), nor does it affect 
the registration status of currently registered aftermarket additives.
    For example, an unregistered detergent additive (either bulk or 
aftermarket) intended for use in gasoline and meeting the criteria for 
grouping with baseline gasoline fuels and bulk additives will be 
registrable.\8\ On the other hand, an unregistered chromium-containing 
additive (either bulk or aftermarket) intended for use in gasoline will 
be considered ``new'' rather than ``registrable,'' because there are no 
currently registered chromium-containing fuels or bulk additives in the 
gasoline fuel family with which the applicant additive could be 
grouped. Even if a chromium-containing product had previously been 
registered as an aftermarket additive for gasoline [e.g., as a 
``grandfathered'' product registered prior to the ban of such 
aftermarket additives under CAA section 211(f)(1)(B)]\9\ or as a bulk 
additive for use in another fuel family (e.g., diesel fuel), the 
applicant additive would still be considered ``new''.
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    \8\The grouping criteria for the baseline gasoline category are 
described in Section IV.B.2.a.
    \9\Until the 1990 CAA Amendments went into effect, the statutory 
language of section 211(f) was interpreted as applying only to 
unleaded gasoline fuels and related bulk additives. Thus, prior to 
November 15, 1990 (the effective date of the CAA Amendments), 
aftermarket additives intended for use in unleaded gasoline and 
containing elements in addition to carbon, hydrogen, oxygen, 
nitrogen, and/or sulfur were allowed to be registered. Under the 
1990 CAA Amendments, all types of motor vehicle F/FAs were placed 
under section 211(f) jurisdiction. [However, section 211(f) 
provisions do not apply until certification requirements are 
promulgated for the specific motor vehicle fuel or fuel additive.] 
All aftermarket additives that were not ``substantially similar'' 
and were introduced on or after November 15, 1990 were banned. 
However, this ban did not apply retroactively. Thus, ``non- 
substantially similar'' gasoline aftermarket additives which had 
been registered prior to November 15, 1990 have been allowed to 
retain their registrations. These are so-called ``grandfathered'' 
aftermarket additives.
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    As discussed in the Reopening Notice, EPA believes that the 
distinctions between registrable and new F/FAs, both in terms of their 
definitions and their respective compliance requirements, reflect 
reasonable regard for the public health and welfare without undue 
interference in the F/FA marketplace. Because registrable F/FAs are 
defined such that they must be reasonably similar in composition and 
usage to current F/FAs, their entry into the market will generally not 
be expected to increase the health or welfare risks potentially related 
to current F/FA emission exposures, assuming the overall rate of usage 
does not increase substantially.
    Today's rule implements EPA's policy that F/FAs that may pose new 
or different health risks to the public should not be allowed on the 
market until EPA has determined that adequate health testing has been 
conducted. Because it would cause significant hardships to pull all 
currently used products off the market until they were tested, products 
that are already registered may continue to be sold. If these 
principles were strictly applied (i.e., EPA refused to register any 
specific product that does not currently have a registration), there 
could be significant stagnation in the marketplace--a new company that 
wanted to sell the same unleaded gasoline that everyone else is selling 
would be prevented from getting a registration until it had tested its 
gasoline product. Thus, today's rule allows new registrants to sell 
products that are similar to registered products in terms of: (1) 
Expected health effects; and (2) usage (and, therefore, type or extent 
of exposure) currently allowed by law.
    If an unregistered product can group with a registered product, EPA 
has determined that the products should have similar health effects. To 
ensure that usage (and, therefore, type or extent of exposure) is 
similar to a registered product, under today's rule, a manufacturer 
cannot rely on registration of an aftermarket fuel additive or on a F/
FA in a different fuel family. Considering a fuel or bulk additive to 
be registrable based on an aftermarket additive registration could 
significantly increase the public exposure to that F/FA. Generally, 
aftermarket additives are relatively limited in distribution and usage 
and, therefore, in exposure. Thus, in seeking a registration for a new 
product, a manufacturer cannot rely on the registration of aftermarket 
additives or any F/FA product in a fuel family other than the one for 
which the registration is sought.
    This is consistent with Congress' intent in CAA section 
211(f)(1)(B) to preclude introduction into commerce of new aftermarket 
additives which do not fit the ``substantially similar'' 
criteria.\10\, \11\ Looking only to F/FAs in the same fuel family 
to determine registrability is also intended to prevent potential 
increases in exposure to untested products. Expanding the use of an 
additive from one fuel family to another (e.g., from diesel fuel to 
gasoline) would significantly increase the overall size of the 
potential market for the product and thus the potential exposure to its 
emissions.
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    \10\See memorandum from James W. Caldwell to Mary T. Smith 
regarding ``Review of Notifications Submitted Pursuant to 40 CFR 79 
for Compliance with the `Substantially Similar' Rule for Unleaded 
Gasoline,'' available in Docket A-90-07, Item No. IV-B-07.
    \11\The grouping criteria in this final rule (see Section IV) 
allow aftermarket additives to join the baseline group if they 
contain no elements in addition to carbon, hydrogen, oxygen, 
nitrogen, and/or sulfur, even if they may sometimes be used by 
consumers in an amount greater than the gasoline ``substantially 
similar'' restriction of 2500 ppm.
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    EPA interprets section 211(e) to support the distinction between 
registered or registrable F/FAs and new F/FAs. EPA believes that the 
reference in CAA section 211(e)(2) (A) and (B) to a ``fuel or fuel 
additive which is registered'' or ``which is not registered'' is 
ambiguous as to whether it refers to the F/FA product generally or to a 
particular product-specific registration. Given this ambiguity, EPA 
believes that it is reasonable to interpret the phrase ``fuel or fuel 
additive which is registered'' to refer to the F/FAs generally. A 
contrary interpretation would result in EPA allowing numerous types of 
unleaded gasoline to be sold under existing registrations, while at the 
same time denying a registration (until completion of necessary 
testing) to a new company that wanted to sell the same type of unleaded 
gasoline that many others now sell. As discussed above, for 
``registrable'' products, such an interference in the market would not 
likely result in any public health benefit. Therefore, EPA believes it 
is reasonable to interpret the phrase ``registered'' to include both F/
FAs that are either ``registered'' or ``registrable.'' ``Registrable'' 
F/FAs are sufficiently similar in composition and use to existing F/FAs 
that one would not expect them to have dissimilar health effects; and 
therefore, it is reasonable to interpret the phrase ``registered fuel 
or fuel additive'' to include not only those F/FAs that are identical, 
but also those that can group with existing F/FAs.
    Alternatively, EPA believes that the above approach is consistent 
with section 211 because EPA interprets section 211(e)(3) (A) and (B) 
in conjunction with section 211(e)(3)(C), which gives EPA authority to 
exempt any F/FA product from duplicative testing. Thus, even if one 
interprets the phrase ``fuel or fuel additive which [is/is not] 
registered'' to mean either that an identical F/FA product must already 
have a registration, or to refer to a product-specific registration, 
EPA believes it is reasonable to interpret section 211(e)(3)(C) to 
allow F/FAs that are similar in composition and usage to those already 
on the market to group with those similar F/FAs and complete the 
testing with the other F/FAs in their group. At the same time, EPA 
believes that for F/FAs that differ significantly in composition or 
usage from currently registered F/FAs, such testing would not be 
duplicative of testing of registered F/FAs; and therefore, EPA is 
authorized under section 211(e) to require this information prior to 
registration. Under either theory, EPA's authority to obtain 
information prior to registration is not limited to Tier 1 and Tier 2 
data, because section 211(b)(2)(B) gives EPA authority to require for 
registration any information necessary to assess the effects of 
emissions on public health or welfare. Therefore, EPA interprets 
section 211 (e) and (b) to give it the authority to require any 
necessary health or welfare effects information for F/FAs that are 
significantly different in composition or usage from currently 
registered products.
    d. Changes to 40 CFR Part 79. This final rule includes revisions to 
the current 40 CFR part 79 registration regulations which are necessary 
to properly implement the new testing requirements in Subpart F. These 
consist of various conforming changes in registration procedures, 
requirements, and terminology.
    The submission by which a manufacturer requests registration of a 
fuel or fuel additive product is now called an ``application'' rather 
than a ``notification,'' in order to better reflect the additional 
submissions and requirements upon which registration is now contingent, 
and to avoid confusion with the various notifications concerning 
testing requirements which will be transmitted to applicants and 
registrants by EPA. Conforming changes have been made in procedures for 
notifying an applicant that a submission does not comply with 
registration requirements and for granting registration.
    As discussed in Section IV of this preamble, in order to determine 
in which of the F/FA group(s) established under the criteria set forth 
in this rule (see Sec. 79.56) a particular fuel additive should be 
enrolled, and to administer applicable testing requirements separately 
for each such F/FA group, it is essential that fuel additives be deemed 
to be registered only for those specific types of fuel for which they 
will be sold and used. The Agency interprets and is already 
administering the existing part 79 in this manner. However, in the 
conforming changes included in this rule, part 79 has been clarified to 
confirm that registration of fuel additives is fuel specific, and that 
EPA considers sale or distribution of a fuel additive product for use 
in a fuel for which it is not registered to be unlawful.
    In addition, a manufacturer seeking to register a fuel product or a 
fuel additive product for use in vehicles manufactured after 1974 must 
demonstrate that it is ``substantially similar'' to fuels or additives 
utilized in the certification of vehicles for 1975 or subsequent model 
years, or that the manufacturer has obtained a waiver under CAA section 
211(f)(4). This expressly codifies the manner in which EPA has 
administered the registration program since it adopted criteria 
implementing the statutory prohibition in CAA section 211(f)(1) on 
introduction into commerce of fuels and fuel additives which are not 
``substantially similar.''
2. Program Focus On Emissions
    CAA section 211 gives the Agency discretion to determine the focus 
of the F/FA testing program under CAA section 211(b). EPA is exercising 
its discretion by focusing this rule on the testing of emissions, 
because the main purpose of the testing program is to provide EPA with 
information that can be used in regulatory decision-making under CAA 
section 211(c).
    Section 211(c) of the CAA gives EPA the authority to regulate F/FAs 
based on the impact of their emissions on public health or welfare. 
Specifically, it allows the Administrator to control or prohibit the 
manufacture, introduction into commerce, or sale of any fuel or fuel 
additive whose emission product(s) cause or contribute to harmful air 
pollution. The legislative history of the provision also supports a 
focus on emissions, since House and Senate Reports on the CAA 
Amendments of 1970 link the information to be obtained under CAA 
section 211(b) to EPA's authority to regulate emissions under CAA 
section 211(c).\12\ Thus in the NPRM, EPA proposed to focus this rule's 
requirements on the potential emissions-based effects of F/FAs rather 
than on the effects of the raw (i.e., uncombusted) F/FA product. Public 
comment received after publication of the NPRM generally supported the 
proposed emissions-based focus of the rule. Accordingly, EPA has 
retained this focus in today's action. The health effects testing 
requirements of this final rule specifically address the effects of 
inhalation exposure to F/FA combustion and evaporative emissions. The 
required testing focuses on the evaluation of health effects of the 
whole emissions of the fuel or additive/base fuel mixture of interest 
and not on the toxicity of the individual emission products.
---------------------------------------------------------------------------

    \12\H. Rep. No. 1146, 91st Cong. 2nd Sess. (1980) at 13, 
reprinted in Environment and Natural Resources Division of the 
Library of Congress, 93rd Cong., 2nd Sess.; A Legislative History of 
the Clean Air Act Amendments of 1970 (Comm. Print 1974) (``Leg. 
Hist.'' at 433-434).
---------------------------------------------------------------------------

    For the purposes of this rule, combustion emissions are the primary 
exhaust products of the combustion of a fuel or additive/base fuel 
mixture in a motor vehicle engine and do not include secondary 
atmospheric transformation products. EPA recognizes that secondary air 
pollutants are a factor in the characterization of overall risks 
associated with F/FA emissions. However, it is not feasible to include 
this type of laboratory testing as a standard requirement at this time. 
When required in specific instances, transformation products will be 
addressed under Tier 3, as described in Section III.C.
    As proposed, evaporative emission testing is to be performed for F/
FAs meeting specific volatility criteria. While some commenters asked 
EPA to eliminate the evaporative emission testing from the program, 
today's rule maintains this requirement. EPA's decision is supported by 
the legislative history, which expresses the concerns of Congress about 
the public health impacts of emissions from both combustion and 
evaporative sources. Public exposure to evaporative emissions is still 
significant and, for many F/FAs, the toxicity of evaporative emissions 
as a whole mixture has not been characterized. Thus, this rule includes 
requirements for the characterization and biological testing of 
evaporative emissions in certain circumstances.
    While combustion emissions are inevitable products of the engine 
combustion process, the significance of evaporative emissions depends 
on the type of F/FA product. As proposed in the NPRM, this final rule 
specifies criteria for determining the need for evaporative emission 
testing. For fuels that are supplied to motor vehicle engines by way of 
sealed containment and delivery systems, evaporative emissions testing 
is less important, since human and environmental exposure should be 
extremely low or nonexistent. Thus, evaporative emissions testing under 
this final rule would not apply to methane (compressed natural gas or 
liquified natural gas) or propane (liquified petroleum gas) 
formulations.
    For liquid F/FAs, the significance of vaporization varies widely, 
depending largely on the volatility of the fuel or additive/base fuel 
mixture. Thus, this final rule uses the Reid Vapor Pressure (RVP) of a 
fuel or additive/base fuel mixture to determine its applicability for 
evaporative emissions testing. An RVP of 2.0 pounds per square inch 
(psi) is designated as the threshold for determining the need for 
evaporative emission testing for fuels. That is, fuels with RVP of 2.0 
psi or greater are subject to the evaporative emissions testing 
requirements, while those with RVP less than 2.0 psi are excused from 
the evaporative emission testing requirements under Tier 1 and Tier 2.
    With respect to additives, the NPRM proposed to require evaporative 
emission testing if the RVP of the additive/base fuel mixture was 
increased by 0.1 psi or more in comparison with the RVP of the base 
fuel alone. However, methods for measurement of vapor pressure have a 
reproducibility of about 0.3 psi.\13\ To account for this limitation of 
measurement accuracy, especially when dealing with low pressure 
measurements, today's rule uses a 0.4 psi criterion (i.e., 0.1 
0.3) for additives. Accordingly, this final rule requires 
the evaporative emission testing of additives when the RVP of the 
associated fuel in the additive/base fuel mixture is increased by 0.4 
psi and the resulting RVP of the additive/base fuel mixture is 2.0 psi 
or more.\14\ For example, an additive that causes an increase of 0.6 
psi when mixed with a fuel with a vapor pressure of 1.0 psi (i.e., the 
resulting RVP of the additive/base fuel mixture is 1.6), need not be 
tested for evaporative emissions. On the other hand, an additive that 
causes an increase of 1.1 psi when mixed with a fuel with RVP of 1.0 
psi is required to undergo evaporative emission testing because the 
resulting RVP of the additive/base fuel mixture is 2.1 psi.
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    \13\See ``Standard Test Method for Vapor Pressure of Petroleum 
Products (Mini Method),'' ASTM D 5191-91.
    \14\The requirement to test the evaporative emissions of a 
qualifying additive product do not apply if the manufacturer intends 
to satisfy the test requirements of the additive as part of a group, 
of which another member product or a base fuel serves as the group 
representative, and the manufacturer does not specifically test the 
additive apart from the group. See Section IV for a discussion on 
grouping provisions.
---------------------------------------------------------------------------

    The above defined thresholds are used by EPA in determining the 
applicability of evaporative emission testing for the purposes of Tier 
1 and Tier 2. However, EPA retains the authority to require evaporative 
emission testing under Tier 3 for fuels or additive/base fuel mixtures 
with low vapor pressure, e.g., RVP less than 2.0, if there is a health 
or welfare concern associated with the evaporative emissions of the 
fuel or additive/base fuel mixture in question. For example, if a 
highly toxic substance is present in a fuel or additive/base fuel 
mixture, EPA could require evaporative emission testing under Tier 3, 
even if the RVP of the F/FA product in question is below 2.0 psi. These 
special cases will be handled on a case-by-case basis under Tier 3.
3. Program Applicability
    The requirements of this rule apply to manufacturers (including 
importers) of designated fuels or fuel additives and to any F/FA 
manufacturer [see Sec. 79.2(d) and (f) as amended in this rule] seeking 
registration under CAA section 211(a) and 211(b). A fuel is defined to 
be any material which is capable of releasing energy or power by 
combustion or other chemical or physical reaction [see 40 CFR 
Sec. 79.2(c)]. A fuel additive is defined as any substance that is 
intentionally added to a fuel (including any added to a motor vehicle's 
fuel system) and that is not intentionally removed prior to sale or use 
(see 40 CFR Sec. 79.2(e), as amended in this rule), including both bulk 
and aftermarket additives.
    At the present time, the designation of F/FAs encompasses both 
leaded and unleaded gasoline F/FAs and diesel
F/FAs produced and commercially distributed for use in motor vehicles.
 F/FAs intended only for off-road use (e.g., farm and construction 
equipment, aircraft, boats, railroad engines) are not currently 
designated to be registered, and thus are not subject to the 
requirements of today's rule. However, if off-road F/FAs become 
designated in the future (according to provisions under CAA section 
213), this rule will be modified, as needed, to cover them as well.
    While the designated F/FAs include leaded gasoline formulations, 
CAA section 211(n) provides that ``after December 31, 1995 it shall be 
unlawful for any person to sell, offer for sale, supply, offer for 
supply, dispense, transport, or introduce into commerce, for use as 
fuel in any motor vehicle (as defined in section 7554(a) of this title) 
any gasoline which contains lead or lead additives.'' Because of the 
upcoming ban of leaded F/FAs, compliance with the requirements in 
today's rule will be superfluous for manufacturers of leaded products. 
Thus, the leaded fuel family has been deleted from this final rule.
    While alternative fuels and their additives are currently on the 
market, they are not yet designated and thus not yet required to be 
registered. However, because they are currently used and EPA 
contemplates their future designation, this final rule includes 
provisions for their registration and testing. The alternative fuels 
for which provisions are included are: methanol, ethanol, compressed 
natural gas (CNG), liquified natural gas (LNG), and liquified petroleum 
gas (LPG). EPA is currently developing the proposal for the designation 
of these alternative fuels and their additives. In that proposal, EPA 
currently intends to propose to set an effective date for the final 
designation rule far enough in the future so that these F/FAs could 
complete whatever pre-registration testing would be required prior to 
the time they would be required to be registered. EPA currently 
believes the delay of the effective date of the designation rule would 
be justified by the need to minimize disruptions in an existing market 
for alternative F/FAs.
    This rule contains a number of special provisions which reduce or 
modify the program's requirements for certain manufacturers or certain 
classes of products. Such special provisions apply to small businesses 
(as defined in this rule) and manufacturers of experimental F/FAs, 
relabeled products, and aerosols. These provisions are described in 
Section X of this preamble.

B. Health Evaluation Requirements

    The testing program established in this rule focuses on the 
identification and evaluation of potential adverse health effects 
associated with inhalation exposure to F/FA emissions. The Tier 2 
testing program of this rule addresses, in addition to the areas of 
inquiry mandated by the statute (carcinogenicity, teratogenicity, and 
mutagenicity), specific assessments designed to detect potential 
pulmonary, neurotoxic, and general reproductive effects of F/FA 
emissions.
    In the NPRM, short-term (42-day) tests were proposed under Tier 2 
to address each of the health effect endpoints described above. 
However, in response to public commentary and EPA's own analysis, the 
Tier 2 testing requirements have been modified in this final rule to 
enhance the efficiency and feasibility of the program. Today's rule 
uses a comprehensive 90-day subchronic inhalation protocol and 
ancillary tests to examine general systemic and organ toxicity 
(including pulmonary toxicity), as well as the specific areas of 
concern described above.
    These evaluations require the exposure of laboratory animals to the 
whole emissions of F/FAs. Tier 2 tests are to be conducted for both 
combustion and (when applicable) evaporative emissions. The subchronic 
inhalation protocol allows the examination of specific endpoints within 
the 90-day testing framework. For example, pulmonary and neurotoxic 
effects are examined in conjunction with the subchronic study standard 
histopathological requirements. The neurotoxicity assessment also 
includes a biochemical assay to measure the level of glial fibrillary 
acidic protein (GFAP). Coordinated with the 90-day study is a battery 
of three assays used in the evaluation of carcinogenicity and 
mutagenicity: the in vitro Salmonella assay, the in vivo micronucleus 
assay, and the in vivo sister chromatid exchange assay. A fertility 
assessment that looks at both reproductive and teratogenic effects is 
also coordinated with the general toxicity study. The assessment for 
reproductive effects involves the mating of exposed animals, the 
measurement of reproductive cycles, and the histopathology of male and 
female reproductive organs. The teratogenic assessment requires the 
exposure of pregnant females to F/FA emissions and the subsequent 
examination of the uterus and its contents just prior to the normal 
time of parturition.
    In addition to the evaluation of the health effects described 
above, EPA retains the authority under Tier 3 to require additional 
testing on a case-by-case basis on those endpoints evaluated under Tier 
2 and/or on other endpoints of concern. Further discussion about the 
specific requirements of Tiers 1, 2, and 3 is provided in Sections VII-
IX.

C. Welfare Evaluation Requirements

    CAA section 211(b)(2)(B) states that the Administrator may require 
manufacturers to furnish ``reasonable and necessary'' information for 
determining ``the extent to which F/FA emissions affect the public 
health or welfare''. The term ``welfare effects'' encompasses a variety 
of complex and interrelated factors. In terms of motor vehicle F/FA 
emissions, welfare effects could include the impact of air pollution on 
the public health and the environment, including a broad range of 
effects on aquatic and terrestrial ecosystems, cultivated crops and 
other vegetation, natural and man-made materials, wildlife, and 
stratospheric ozone. Air pollution effects on the public welfare also 
include important environmental concerns such as noxious odors or 
visibility impairment, which may detract from human well-being.
    Except for stipulating that welfare effects should be addressed, 
the statute gives EPA broad discretion about how to address welfare 
effects. EPA recognizes that, at the present time, scientific 
experience and laboratory screening methods for the evaluation of 
welfare effects are more limited than in the area of health effects. 
Thus, today's rule limits the routinely required welfare evaluation to 
requirements that are coincident or concurrent with the evaluation of 
health effects. These include the literature search, emission 
characterization, and exposure analysis requirements of Tier 1. While 
at this time EPA is not requiring biological testing for welfare 
effects, the Administrator retains the authority to require additional 
evaluation and/or testing of welfare effects at the Tier 3 level, when 
the outcome of lower tiers demonstrates both significant environmental 
toxicity and exposure potential. EPA will determine the need for Tier 3 
welfare effects testing on a case-by-case basis.
    In the NPRM, EPA proposed to require modeling analyses for 
atmospheric reactivity, environmental fate/partitioning, and exposure 
as part of the welfare evaluation. EPA requested comments on the 
feasibility of requiring such modeling analyses as a routine 
requirement for registration. Commenters urged EPA to limit the 
modeling requirements due to the lack of standardized methods in this 
area of study. Recognizing the limitations of modeling methods and the 
availability of existing data for some of the areas of study of 
interest in this rule, this final rule does not require modeling 
analyses as part of Tier 1.
    EPA recognizes that other EPA programs are actively researching and 
controlling mobile and stationary source contributions to major air 
pollution problems such as tropospheric/stratospheric ozone, global 
warming, and acid rain. Furthermore, modeled ozone reactivity data are 
already available for most conventional and alternative fuels. EPA 
believes that if additional modeling is deemed necessary, this could be 
performed by manufacturers under Tier 3. Also, EPA may conduct simple 
modeling, using the emission data submitted by registrants under Tier 
1, if needed for regulatory decisions.
    Regarding environmental fate and exposure modeling, EPA recognizes 
that these types of analyses will be extremely difficult due to the 
complex nature of F/FA emissions. Because both environmental 
partitioning models and exposure models address single compounds rather 
than mixtures, it would be unduly burdensome and unreasonable to 
require all registrants to perform these analyses on each individual 
emission constituent. Requiring the modeling of each individual 
emission product would also result in duplication of information and, 
therefore, would be inconsistent with the original intent of the 
statute.
    In addition, the environmental models are applicable only to a 
limited number of emission products for which appropriate physical/
chemical data are available in order to perform the analysis. In terms 
of available exposure models, these usually rely on carbon monoxide 
monitoring data and related emission rates to estimate potential 
exposures. This means that the applicability of available exposure 
models is somewhat limited to the analysis of compounds whose chemical/
physical behavior is similar to carbon monoxide.
    Based on the above factors, EPA believes that quantitative 
evaluations of potential exposures and environmental fate/partitioning 
of F/FA emissions will be better addressed at the Tier 3 level on a 
case-by-case basis, where they can be focused on specific compounds of 
potential environmental concern.

D. Requirements for Emission Control System Testing

    CAA section 211(b)(2) requires F/FA manufacturers to provide 
information to determine their products' effects on ECS performance. 
The NPRM stated that EPA intended to continue addressing ECS 
performance through the existing waiver application program under CAA 
section 211(f). The waiver program prevents the introduction into 
commerce of F/FAs which would significantly degrade the performance of 
emission control equipment. Under CAA section 211(f), F/FA formulations 
which do not meet specific chemical and physical criteria considered to 
be ``substantially similar'' to EPA certification fuel (see 
interpretive rule in 56 FR 5352), cannot be introduced into commerce 
unless a waiver is issued by EPA. The waiver process then requires the 
applicant to demonstrate, through testing if necessary, that ``such 
fuel or fuel additive or a specified concentration thereof, and the 
emission products of such fuel or fuel additive or a specified 
concentration thereof, will not cause or contribute to a failure of any 
emission control device or system (over the useful life of any vehicle 
in which such device or system is used) to achieve compliance by the 
vehicle with the emission standards to which it has been certified.'' 
For products already registered that do not meet ``substantially 
similar'' criteria, i.e., grandfathered products that were registered 
prior to the implementation of the waiver application program, EPA 
proposed to establish a mechanism that would permit the public to 
submit petitions to EPA requesting ECS testing for a particular fuel or 
fuel additive of concern.
    Today's rule reflects EPA's judgment that the mechanisms already 
established under CAA section 211(f) are adequate for the ECS testing 
of F/FAs. EPA's previous experience with the waiver application process 
has demonstrated the practical value of the ``substantially similar'' 
concept for determining whether a F/FA product needs to be tested for 
its effects on emission control equipment. EPA is not aware of 
instances in which products meeting ``substantially similar'' criteria 
were later discovered to have adverse effects on vehicular emission 
control performance. The implementation of another ECS testing program 
under section 211(b) would be duplicative and, therefore, inconsistent 
with Congress' intent. Thus, as proposed, today's rule refers to the 
waiver application process under CAA section 211(f)\15\ for the ECS 
testing of ``substantially similar'' F/FAs and
F/FAs required to obtain a waiver under CAA section 211(f)(4). Products 
which conform to applicable ``substantially similar'' criteria are not 
required to undergo ECS testing before they can be registered. On the 
other hand, new
F/FAs which do not meet ``substantially similar'' criteria are subject 
to the standard 211(f) application process prior to registration.
---------------------------------------------------------------------------

    \15\An example of a waiver decision can be found in 53 FR 33846.
---------------------------------------------------------------------------

    EPA recognizes that there are grandfathered F/FA products (see 
Section III.A above) which fall outside the regulatory domain of CAA 
section 211(f). These grandfathered products include gasoline 
aftermarket additives introduced prior to the 1990 CAA Amendments. 
Therefore, statutory authority for the ECS evaluation and regulatory 
control of grandfathered products exists under CAA sections 211 (b) and 
(c) rather than section 211(f). EPA judges that requiring ECS 
evaluation of all grandfathered products, without evidence of ECS 
problems, would be unreasonable and unnecessarily burdensome on the 
industry. Instead, today's rule provides a petition mechanism for the 
ECS evaluation of grandfathered products. Under this mechanism, EPA 
could require ECS testing of grandfathered products, similar to the 
testing which a waiver applicant would generally conduct, if so 
petitioned by outside parties or if other information available to the 
Agency indicates that such evaluation is appropriate. Such information 
might be obtained as a result of the emission characterization 
requirements included in this final rule. In addition, vehicle 
manufacturers or other outside parties are allowed to submit petitions 
to EPA requesting the testing of grandfathered products based on 
evidence of potential harm to vehicular ECS. If EPA judges that ECS 
testing is warranted after reviewing the petition arguments, emission 
characterization results and/or other available information, the 
authority provided by CAA sections 211(b) and 211(c) to require 
specific grandfathered products to test for ECS effects.

IV. Grouping System

A. Objectives and Rationale

    CAA section 211(e) provides a number of mechanisms by which EPA may 
reduce the costs and burdens of compliance with the registration 
requirements set forth in CAA section 211(b). In particular, CAA 
section 211(e)(3)(B) permits the Administrator to ``provide for cost-
sharing with respect to the testing of any fuel or fuel additive which 
is manufactured or processed by two or more persons, or otherwise 
provide for shared responsibility'' so that the program requirements 
can be met without duplication of effort. In accordance with this 
provision, today's rule maintains the grouping system proposed in the 
NPRM, which permits manufacturers of similar F/FAs, on a voluntary 
basis, to pool their resources and efforts to satisfy the registration 
requirements. The groups defined by the specifications in this final 
rule are the only groups permitted for satisfying the requirements of 
the registration program.
    As proposed in the NPRM, the grouping system allows similar fuels 
and additives to be grouped together, rather than creating separate 
fuel groups and additive groups. This convention recognizes that, to 
meet the requirements of this final rule, an additive must be mixed 
with its associated base fuel\16\ prior to generating the emissions for 
testing. To the extent that the resulting additive/base fuel mixture is 
similar to existing fuel formulations, the tests conducted on the 
emissions of the additive/base fuel mixture will be duplicative of 
tests conducted on the related fuels. To avoid potential duplication, 
this final rule maintains the proposed approach, in which closely-
related fuels and additives are grouped together. Accordingly, the 
manufacturers of fuels and the related additives can fulfill their 
individual registration responsibilities through jointly-supported 
testing rather than through duplicative independent efforts. By 
grouping similar fuels and additives together, the grouping scheme also 
avoids the need to define each generic product or product component as 
either a ``fuel'' or an ``additive.'' This would otherwise present a 
problem when a given substance (or mixture) can serve as either a fuel 
or an additive (e.g., ethanol).
---------------------------------------------------------------------------

    \16\Base fuel specifications for each fuel family are described 
in Section V.
---------------------------------------------------------------------------

    In the NPRM, EPA developed criteria for sorting individual F/FAs 
into groups of related formulations based on similarities in the 
chemical/physical properties of the ``raw'' fuel or additive/base fuel 
mixture. EPA has maintained this approach in the final rule. EPA 
expects F/FAs within each group to have similar emission 
characteristics and thus essentially the same general effects on the 
public health and welfare. Therefore, chemical or toxicologic 
information associated with individual members of a given group can 
reasonably be generalized to all F/FAs in the group. EPA will consider 
tests performed on a selected representative of a group to apply to all 
members of the group for purposes of compliance with registration 
requirements, for deciding whether to require additional testing under 
Tier 3, or for taking regulatory action under CAA section 211(c).
    While each manufacturer of a fuel or fuel additive will still be 
held individually accountable for compliance with the registration 
program, the grouping system provides an opportunity for meeting the 
program requirements in a more cost-effective manner. Participation in 
the F/FA grouping system is strictly voluntary, and any manufacturer 
may choose to fulfill the requirements on an individual basis. Those 
who choose to take advantage of the grouping opportunity will be able 
to share their planning efforts, research capabilities, and financial 
resources to satisfy the information-gathering and testing requirements 
of the F/FA registration program. To satisfy the chemical and 
biological testing requirements, the required tests will be done on the 
selected representative for the respective group, rather than being 
repeated for each of the F/FAs in the group. The results of the tests 
on the group representative will then be submitted jointly for all 
members of the group, with applicable costs to be shared by the 
respective manufacturers (based on their cost-sharing agreements, as 
discussed in Section IV.C). Manufacturers who question whether the 
results obtained for their group's representative are valid for their 
own products may conduct confirmatory tests on their products on an 
independent basis and at their own cost. However, until such 
independent test results are made available to EPA, the original 
results submitted on behalf of the group will be considered valid for 
all member products, and could be applied by EPA to support regulatory 
decisions under CAA section 211(c) or requirements for further testing 
under CAA section 211(b).
    The F/FA grouping system is expected to provide a number of 
benefits to the F/FA manufacturers who are responsible for registration 
while increasing the efficiency and functionality of the registration 
program itself. First, the grouping system will reduce the overall 
costs of the registration program by avoiding the generation and 
submission of essentially redundant information by individual 
manufacturers with similar products. In addition, by reducing the 
number of individual formulations that will be subject to testing, the 
grouping system is expected to ease the pressure and demands on limited 
laboratory capacity.

B. Grouping Approach and Criteria

    The basic conceptual framework for the grouping system is 
illustrated in Figure 3. First, each fuel or additive is sorted into 
one of six broad ``fuel families.'' F/FAs in each fuel family are then 
subdivided into three ``F/FA categories.'' The categories are further 
subdivided into ``F/FA groups''--the ``working'' units of the grouping 
system. It is among the members of the F/FA groups that cooperative 
evaluation and testing efforts can be pursued using designated group 
representatives. This grouping system is very similar to the approach 
that was proposed in the NPRM, with the exception that the original 
proposal has been simplified in today's rule by eliminating the 
separate concept of ``formulation class.'' EPA judged that the 
``formulation class'' concept could be confusing, and was not necessary 
for the structure or implementation of the grouping system. The key 
parameters and relationships within this grouping framework are further 
explained in the following sections. A summary of the grouping system 
is provided in Table F94-7 (see Sec. 79.56) of the accompanying 
regulatory text for this rule.

    BILLING CODE 6560-50-P

TR27JN94.003


    BILLING CODE 6560-50-C
    Generic rules for categorization and grouping are used to determine 
specific F/FA groups based on the raw composition of the particular 
products under consideration. The first step entails the selection of 
the applicable fuel family and category for the product according to 
the criteria discussed below. Rules specific to the categories then 
define the proper F/FA group. After the group is formed and 
arrangements are made for cooperative testing efforts, applicable 
criteria will be applied to select a representative of the group to be 
used in group-sponsored testing. In determining the category and group 
to which a fuel or fuel additive belongs, impurities present in trace 
amounts can be ignored unless otherwise restricted in the definition of 
the particular fuel family. Impurities in fuels or fuel additives are 
substances which are present through contamination, or remain 
naturally, after processing is completed.
1. Fuel Families
    This final rule defines six fuel families, as follows: (1) Gasoline 
(containing more than 50 percent gasoline by volume), (2) diesel 
(containing more than 50 percent diesel by volume; includes both diesel 
#1 and diesel #2 formulations), (3) methanol (containing at least 50 
percent methanol by volume), (4) ethanol (containing at least 50 
percent ethanol by volume), (5) methane (includes compressed natural 
gas and liquified natural gas containing at least 50 mole percent 
methane), and (6) propane (i.e., liquid petroleum gas containing at 
least 50 percent propane by volume). A manufacturer seeking to register 
a formulation which does not fit the criteria for inclusion in any of 
the above fuel families shall contact EPA for guidance in classifying 
and testing such formulation.
    In the NPRM, EPA proposed to establish two gasoline fuel families: 
unleaded and leaded. As explained previously, EPA is not including a 
leaded fuel family in this final rule due to the upcoming ban of leaded 
F/FAs under CAA section 211(n). Thus, the unleaded fuel family has been 
renamed to become simply the ``gasoline'' fuel family in this final 
rule.
    Fuel families consist of a constellation of F/FA products sharing 
basic characteristics in regard to their chemical/physical properties 
and engine/vehicle applicability. For ease of reference, the gasoline 
and diesel families are regarded as the ``conventional'' fuel families, 
while the remaining four are referred to as the ``alternative'' fuel 
families. If a manufacturer develops a F/FA product that does not meet 
the definition of any of the fuel families included in this rule, EPA 
will define additional fuel families to include such a product.
    Each fuel family includes not only the fuels referenced in the name 
of the family, but also bulk and aftermarket additives which are 
intended for use in such fuels. Additives which are registered for use 
in more than one type of fuel are assigned to each applicable fuel 
family. For example, an additive product that is registered as both a 
gasoline additive and a diesel additive belongs to both the gasoline 
fuel family and the diesel fuel family. Furthermore, the manufacturer 
of such additive product will be required to test the additive in each 
applicable fuel family. The multiple testing responsibility can be 
satisfied by the manufacturer individually or by participating in the 
applicable groups for each fuel family. For instance, if an additive 
product meets the baseline criteria for both gasoline and diesel, then 
the product will be assigned to two groups, i.e., the gasoline baseline 
group and the diesel baseline group. The manufacturer of such additive 
product will then be able to share the costs of testing with other 
manufacturers of baseline gasoline F/FAs and baseline diesel F/FAs.
    Today's rule specifies the chemical and physical characteristics of 
``base fuel'' formulations for each defined fuel family. These are 
generic formulations (rather than any particular commercial fuel) with 
average or normative characteristics for a given fuel family. Once an 
additive has been assigned to the applicable fuel family or families, 
determination of the proper category and group (for each applicable 
fuel family) for the additive is based on the properties of the mixture 
that results when the additive is mixed in the base fuel of the 
applicable family at the maximum concentration recommended for use by 
the additive manufacturer. Moreover, it is this mixture that is used 
for the generation and testing of additive emissions.\17\ Tests 
conducted on the emissions of the base fuel then serve as one control 
(the other being a clean-air exposure group) against which tests on the 
emissions of the additive/base fuel mixture are to be compared. Further 
discussion on the definition and use of base fuels is presented in 
Section V.
---------------------------------------------------------------------------

    \17\Special provsions related to the testing of additives are 
discussed in Section VI.F.
---------------------------------------------------------------------------

2. F/FA Categories
    Fuel families are subdivided into three F/FA categories: 
``baseline,'' ``non-baseline,'' and ``atypical.'' The baseline category 
consists of fuels and associated fuel additives which resemble the 
respective base fuel for a particular fuel family in terms of elemental 
composition and which conform with certain quantitative limits for 
particular constituents. It is important to understand that a baseline 
category is not limited to base fuels; the baseline category and group 
criteria defined below for each fuel family are considerably less 
restrictive than the respective base fuel definitions (specified in 
Section V). Non- baseline F/FAs contain no chemical elements other than 
those allowed in the baseline category, but they exceed the allowable 
baseline limit for certain constituents for the respective fuel family. 
The atypical category consists, in general, of F/FAs that contain 
chemical elements in addition to those allowed in the baseline 
category. (In a few circumstances, the atypical category also includes 
F/FAs that exceed specified baseline limits for certain constituents, 
as discussed below.) As mentioned above, the category determination for 
fuel additives is based on the properties of the mixture which results 
when the additive is mixed in the appropriate base fuel at the maximum 
concentration recommended for use by the additive manufacturer. If the 
fuel or additive/base fuel mixture contains both non-baseline and 
atypical constituents, the formulation is characterized as atypical. 
Thus, atypical constituents take precedence over non-baseline 
constituents.
    In establishing the F/FA categories (and the groups within them), 
EPA has sought to avoid overly narrow definitions which would result in 
unnecessary and duplicative testing by manufacturers, as well as overly 
broad definitions which would cause potentially important toxicologic 
differences between F/FAs to be obscured. A basic guideline EPA has 
used to find the proper balance between these two unsatisfactory 
scenarios is to ask whether the emissions of a single group 
representative (i.e., test substance) can reasonably be expected to 
reflect the chemical and toxicologic properties of the emissions of the 
F/FAs proposed to be classified together. In applying this guideline, 
EPA has kept in mind both the homogenizing effect of combustion 
processes, as well as the need in some cases to examine the effects of 
evaporative emissions, which generally retain the characteristics of 
the raw F/FA to a greater degree.
    These considerations have led EPA to depend on the chemical 
elements in F/FA formulations as a primary criterion for categorization 
and grouping. Another key criterion is the presence of significant 
amounts (by volume, weight, or potential potency) of F/FA constituents 
that are likely to change the characteristics of the combustion or 
evaporative emissions in substantive ways.
    Accordingly, the baseline category in each fuel family is generally 
comprised of F/FAs containing no elements in addition to carbon, 
hydrogen, oxygen, nitrogen, and/or sulfur.\18\ These elements are the 
fundamental chemical building blocks of all of the fuel families 
defined in this rule. Omitting any of these elements from the list of 
permissible baseline elements would eliminate all or most F/FAs from 
the baseline categories. On the other hand, allowing additional 
elements in the baseline definition would introduce substances not 
characteristic of most F/FA products in the fuel family. If a group 
representative (test substance) did not contain the additional element, 
it could not reasonably be assumed to reflect the presence and activity 
of F/FAs that do contain the element. Conversely, if the group 
representative did contain the additional element, then the results of 
the testing would be influenced by the presence and activity of this 
element, and would therefore not be valid for the large majority of the 
baseline F/FAs. Thus, allowing F/FAs with additional elements to be 
included in the baseline categories (and groups) would violate the 
principles of the grouping system and the objectives of the testing 
program.
---------------------------------------------------------------------------

    \18\The exceptions to this generalization (discussed in later 
sections) are small amounts of chlorine permitted in baseline 
methanol F/FAs, and small amounts of chlorine and copper permitted 
in baseline ethanol F/FAs. Also, trace contamination with elements 
other than carbon, hydrogen, oxygen, nitrogen, and sulfur do not 
cause F/FAs to be eliminated from baseline categories or groups.
---------------------------------------------------------------------------

    F/FA formulations containing elements in addition to those allowed 
in the baseline category of a fuel family are classified in the 
atypical category for that family. As described further below, such F/
FAs are then subdivided into groups based primarily on which atypical 
element(s) they contain. Moreover, the rules for choosing 
representatives of these atypical groups emphasize the atypical 
constituent(s). This approach assures separate testing of F/FA products 
with unique compositional characteristics that can reasonably be 
expected to appear in the emissions and may thus have distinct 
emissions-based toxicologic effects. EPA believes that this approach 
best effectuates CAA section 211(e) by avoiding duplicative testing of 
similar F/FA products while requiring ``each'' F/FA to be tested.
    Between the baseline category and the atypical category in each 
fuel family is the non-baseline category. Broad generalizations about 
the non-baseline F/FA categories are somewhat more difficult to make, 
since they are distinguished from the respective baseline categories by 
various attributes other than elemental composition. In the case of 
gasoline and diesel F/FAs, the distinction is based primarily on the 
presence of significant concentrations of oxygenating compounds. As 
discussed further below, the presence of such compounds may have a 
large impact on F/FA emission profiles. Classification of the 
oxygenated F/FAs into separate categories from the baseline F/FAs (and 
further subdivision into separate groups) is necessary to assure 
testing of representatives that can reasonably reflect the differences 
in these emission mixtures and, possibly, their health effects. Similar 
principles apply to the non-baseline categories in the alternative fuel 
families. In the case of alcohol fuels, non-baseline F/FAs are those 
which have a substantial non-alcohol and non-gasoline component in the 
formulation. Non-baseline propane and methane formulations are those 
containing significant amounts of substances other than propane and 
methane, respectively. In all of these cases, the non-baseline 
definitions serve to ensure that F/FAs with properties that are likely 
to result in significantly different emission profiles, with possibly 
different toxicologic effects, are not subsumed in the baseline 
category.
    The following sections describe the criteria which determine F/FA 
categories for each fuel family. It should be noted that the criteria 
that define baseline F/FA products for each fuel family are not meant 
to be fuel specifications. The baseline criteria used for grouping 
purposes in this final rule consider the potential health implications 
of the composition of the fuel or additive/base fuel mixture and might 
differ from previously established commercial fuel specifications, such 
as those established by the American Society for Testing and Materials 
(ASTM), the California Air Resources Board (CARB), or federal 
``substantially similar'' criteria.
    a. Gasoline. EPA discussed in the NPRM two options (Option A and 
Option B) for distinguishing the baseline and non-baseline categories 
for the gasoline fuel family (see 57 FR 13187-13188). Today's action 
uses Option A for the classification of gasoline products. A discussion 
supporting this decision is included in the ``Summary and Analysis of 
Comments for the Fuels and Fuel Additives Registration Regulations'' 
(available in public docket A-90-07). A synopsis of EPA's analysis 
follows.
    The major differences between the two alternative grouping options 
considered by EPA in the NPRM were: (1) The cutoff point for oxygen 
content to distinguish between baseline and non-baseline products, and 
(2) the approach for handling F/FAs that have received a waiver under 
CAA section 211(f). Under Option A, the baseline category was to be 
limited to F/FAs having less than 1.5 weight percent oxygen by weight. 
Because the cut-off point of 1.5 percent is consistent with the minimum 
oxygen requirement for reformulated gasolines, reformulated gasolines 
would not be considered baseline under Option A. Instead, gasolines 
with 1.5 percent or more oxygen were to be sorted into different non-
baseline groups depending on the oxygenated compounds they contained. 
In contrast, Option B used an oxygen cutoff point of 2.7 weight 
percent, based on current ``substantially similar'' criteria (see 56 FR 
5352). F/FAs which exceeded this limit but had been granted a waiver 
for the excess oxygen were also to be designated as baseline.
    After careful evaluation, EPA has selected Option A for the 
grouping of gasolines in this final rule. In arriving at this decision, 
EPA considered the testing and potential health effects implications of 
both grouping options. The main purpose of the grouping system is to 
sort F/FAs based on the similarities of their emission components. 
After analyzing existing emission characterization data, EPA concluded 
that Option B did not provide an adequate approach for the testing of 
gasoline F/FAs because it does not distinguish between formulations 
that may have significantly different emission characteristics. EPA's 
evaluation showed that the emissions from oxygenated gasolines are not 
the same as for non-oxygenated gasolines and that, furthermore, the 
emission profiles differ according to the particular oxygenated 
compound present in a fuel formulation. Differences in emission species 
will affect the toxicological characteristics of the fuel or additive/
base fuel mixture. Option B was found inappropriate because it would 
have allowed the grouping of gasolines containing significant amounts 
of different oxygenated compounds into one single group. These 
different compounds may have distinguishable toxic effects. Therefore, 
EPA selected Option A for the grouping of gasolines in this final rule 
in order to adequately examine the potential health effects of the 
different oxygenated compounds. EPA believes this best effectuates the 
goal of CAA Section(e) to require testing of each fuel or fuel 
additive.
    In this final rule, gasoline formulations are defined as those 
containing more than 50 percent gasoline by volume. Based on current 
``substantially similar'' criteria (see interpretative rule at 56 FR 
5352), the sulfur content for all gasoline formulations in the gasoline 
fuel family is limited to 0.1 percent by weight.
    F/FAs in the baseline gasoline category must contain no elements in 
addition to carbon, hydrogen, nitrogen, oxygen, and/or sulfur. Gasoline 
baseline formulations must possess, at the time of manufacture, all the 
physical and chemical properties of an unleaded gasoline and applicable 
volatility class standards as specified in the latest version of ASTM 
standard for Automotive Spark-Ignition Engine Fuel, D 4814. As 
discussed above, the oxygen content of baseline gasolines must be less 
than 1.5 percent by weight. The baseline gasoline category includes all 
gasoline fuels and additives (evaluated as additive/base fuel mixtures) 
meeting the above criteria.
    The non-baseline gasoline category is comprised of F/FAs which 
conform to the baseline specifications in terms of elemental 
composition, but exceed the specified baseline oxygen limit. Thus, this 
category includes gasoline formulations with no elements in addition to 
carbon, hydrogen, oxygen, nitrogen, and/or sulfur, which have been 
blended with oxygenates (i.e., alcohol, ether, ester, furan, and any 
other compound used to increase the oxygen content of the gasoline 
formulation), such that the total oxygen content of the gasoline-
oxygenate blend is at least 1.5 weight percent. Included in the non-
baseline gasoline category are reformulated gasolines and oxygenated 
gasolines with at least 1.5 percent oxygen (by weight), including a 
number of formulations which have previously been granted CAA section 
211(f) waivers on oxygen content.
    The atypical category in the gasoline fuel family includes F/FAs 
which contain elements in addition to carbon, hydrogen, oxygen, 
nitrogen, and/or sulfur. (Trace contamination by other elements does 
not cause a F/FA to be classified as atypical, however.)
    The baseline gasoline group is restricted to formulations that are 
derived from conventional petroleum sources. Thus, gasolines derived 
from synthetic crude oils are excluded from the baseline category. 
Synthetic crude oils can be prepared from coal, shale and tar sands, 
heavy oil deposits, and other non-conventional petroleum sources. 
Compared to petroleum, these synthetic crude oils must be extensively 
upgraded before they can be refined into useful products. Because of 
the nature of their sources, these synthetic products are likely to 
contain a variety of unknown contaminants with unknown health effects. 
With little specific data currently available on their composition, EPA 
believes that separate non-baseline classifications are most 
appropriate for grouping these products. Gasoline formulations derived 
from a particular synthetic crude oil source (e.g., coal) will be 
permitted to group together. The representative of each such group will 
be the first such product to seek registration.
    b. Diesel. Diesel formulations are defined as those containing more 
than 50 percent diesel by volume. The sulfur content for all diesel 
formulations in the diesel fuel family is limited to 0.05 percent by 
weight, based on current EPA limits (55 FR 34120).
    The diesel fuel family includes both diesel #1 and diesel #2 
formulations.
    As originally proposed, the diesel baseline category includes 
diesel formulations containing no elements in addition to carbon, 
hydrogen, oxygen, nitrogen, and/or sulfur. Baseline diesel formulations 
must also possess, at the time of manufacture, all the physical and 
chemical properties of a diesel fuel as specified in the latest version 
of ASTM standard D 975. Oxygen content of baseline diesel formulations 
must be less than 1.0 percent by weight. The baseline diesel category 
includes all diesel fuels that meet the above criteria.
    The diesel baseline definition is consistent with existing 
information in EPA's F/FA registration data base, which indicates that 
most commercial diesel fuels, including their bulk additives, consist 
of carbon, hydrogen, oxygen, nitrogen, and/or sulfur. While some 
currently registered diesel fuels contain additives with additional 
elements, the objectives of the testing program are inconsistent with 
broadening the diesel baseline definition to include other elements 
with potentially different health effects from those of basic diesel 
formulations. A broader baseline definition would mean that the 
atypical diesel F/FAs would not be separately examined. Limiting 
baseline diesel F/FAs to those containing no elements in addition to 
carbon, hydrogen, oxygen, nitrogen, and/or sulfur ensures the conduct 
of separate health effects evaluations for the emissions of diesel 
formulations containing atypical elements.
    Similar to baseline gasoline, the baseline diesel category excludes 
fuels derived from synthetic crude oil sources. Thus, such formulations 
are included in the non-baseline category of the diesel family. The 
non-baseline diesel category also includes diesel formulations with 1.0 
percent or more oxygen by weight. Examples of non-baseline diesel 
formulations are alcohol blends and biodiesel formulations.
    c. Methanol. F/FAs in the methanol fuel family are defined as those 
containing at least 50 percent methanol by volume. The baseline 
methanol category is comprised of methanol and methanol-gasoline F/FAs 
that: (1) Contain at least 50 percent methanol by volume, (2) contain 
no more than 4 percent by volume of substances other than methanol and 
gasoline, and (3) contain no elements in addition to carbon, hydrogen, 
oxygen, nitrogen, sulfur, and/or chlorine. The sulfur content of 
baseline methanol formulations is limited to 0.004 percent by weight. 
Chlorine (as chloride) is limited to no more than 0.0001 percent by 
weight. Chlorine is allowed in methanol baseline formulations because 
it is a common contaminant remaining from methanol production.
    The baseline methanol category includes all methanol fuels meeting 
the above criteria and is divided into two groups: M100 group and M85 
group. The M100 group includes methanol-gasoline formulations 
containing at least 96 percent methanol by volume, while the M85 group 
consists of methanol formulations containing 50-95 percent methanol by 
volume.
    F/FAs within the baseline M100 group are required to contain 
odorants and bitterants. These formulations should have a distinctive 
and noxious taste, for purposes of preventing purposeful or inadvertent 
human consumption. The elemental composition of the odorant and 
bitterant is limited to carbon, hydrogen, oxygen, nitrogen, sulfur, and 
chlorine. Baseline methanol formulations in the M85 group must comply 
with the elemental composition specified above for all baseline 
methanol F/FAs, but need not have added odorants and bitterants.
    The non-baseline methanol category is comprised of methanol 
formulations (i.e., containing at least 50 percent methanol by volume) 
that meet the baseline limits on elemental composition, but contain 
more than 4 percent by volume of substances other than methanol and 
gasoline. a
    Atypical methanol F/FAs contain elements in addition to those 
allowed in the baseline methanol category or exceed the specified 
limits for sulfur or chlorine.
    d. Ethanol. Ethanol formulations in the ethanol fuel family are 
defined as those containing at least 50 percent ethanol by volume. The 
final rule defines a single group (represented by E85) for the baseline 
category of ethanol F/FAs. Although in the NPRM, EPA proposed two 
baseline groups for the ethanol fuel family (i.e., E100 and E85), EPA 
expressed its intention to establish a single group for baseline 
ethanol formulations in the Reopening Notice. As discussed in the 
Reopening Notice, the rationale behind this decision is that fuel 
ethanol is required to contain at least five percent denaturant, which 
means that, in actuality, E100 formulations contain only 95 percent 
ethanol (i.e., E95). Furthermore, gasoline is normally used as the 
denaturant for ethanol fuels. EPA judged that there was little 
incremental value in requiring tests of E95 in addition to E85. Thus, 
the final rule creates a single baseline ethanol group represented by 
E85. However, EPA retains the authority to require testing on other 
members of any F/FA group under Tier 3 (see Section IX.A).
    The baseline ethanol category is comprised of ethanol and ethanol-
gasoline F/FAs that: (1) Contain at least 50 percent ethanol by volume, 
(2) contain no more than 5 percent by volume of substances other than 
ethanol and gasoline, and (3) contain no elements in addition to 
carbon, hydrogen, oxygen, nitrogen, sulfur, chlorine, and/or copper. 
The sulfur content of ethanol baseline formulations is limited to 0.004 
percent sulfur by mass. Chlorine (as chloride) and copper are allowed 
in the baseline ethanol formulations at a maximum level of 0.0004 
percent by mass for chloride and 0.07 mg/l for copper. Chlorine and 
copper are permitted in the baseline ethanol formulations because they 
are common contaminants remaining from ethanol production. The baseline 
ethanol category includes all ethanol fuels meeting the above criteria.
    The non-baseline ethanol category is comprised of ethanol 
formulations (i.e., containing at least 50 percent ethanol by volume) 
that meet the baseline limits on elemental composition, but contain 
more than 5 percent by volume of substances other than ethanol and 
gasoline.
    Atypical ethanol F/FAs contain elements in addition to those 
specified in the baseline ethanol category or exceed the specified 
limits for sulfur, chlorine, or copper.
    e. Methane. Methane F/FAs are defined as those containing at least 
50 mole percent methane, including both compressed natural gas (CNG) 
and liquified natural gas (LNG). Baseline methane formulations must 
contain no elements in addition to carbon, hydrogen, oxygen, nitrogen, 
and/or sulfur, and must contain no more than 20 mole percent of non-
methane hydrocarbons. Sulfur content for baseline methane formulations 
(including additives) is limited to 16 parts per million (ppm) by 
volume. Methane formulations must contain added odorants with an 
elemental composition that satisfies the baseline methane definition. 
The baseline methane category includes all methane fuels (and 
associated additives) meeting the above criteria.
    Non-baseline methane formulations are those that exceed the limit 
of 20 mole percent non-methane hydrocarbons. Atypical methane 
formulations include products containing elements in addition to 
carbon, hydrogen, nitrogen, oxygen, and/or sulfur, or exceed the 
baseline sulfur limit of 16 ppm by volume.
    f. Propane. Propane formulations are defined as those containing at 
least 50 percent propane by volume. The baseline propane category 
includes LPG formulations containing no elements in addition to carbon, 
hydrogen, oxygen, nitrogen, and/or sulfur. Baseline LPG products are 
restricted to a maximum of 20 percent by volume for non-propane 
hydrocarbons. Sulfur content (including additives) is restricted to 123 
ppm by weight. LPG formulations must have a distinctive odor. The 
elemental composition of odorants added to LPG formulations is limited 
to carbon, hydrogen, oxygen, nitrogen, and sulfur. The baseline propane 
category includes all propane fuels (and associated additives) meeting 
the above criteria.
    Non-baseline propane formulations are those that exceed the 
specified limit for non-propane hydrocarbons. Atypical propane 
formulations include LPG products that contain elements in addition to 
carbon, hydrogen, nitrogen, oxygen, and/or sulfur, or exceed the 
baseline sulfur limit of 123 ppm by weight.
3. F/FA Groups
    The F/FA groups are subdivisions of the F/FA categories and 
represent the final level of product classification within the grouping 
system. The groups are the actual operating units of the grouping 
system. The objective underlying the group definitions is to sort F/FAs 
together when it is reasonable to assume that their emission products 
will be essentially the same on a qualitative basis.
    A summary table of the F/FA grouping system is included in the 
regulations (see Table F94-7 in Sec. 79.56). In this table, the fuel 
families serve as column headings and the categories define the rows. 
The resulting combination of fuel families and categories (i.e., the 
boxes in the table) contain the F/FA groups. Within each category, one 
or more groups are defined according to the presence of differing 
constituents in the raw fuel or additive/base fuel mixture. The number 
of groups in a particular F/FA category depends on the variability 
among the products in that category. For example, the atypical category 
for each fuel family potentially consists of many groups that are 
defined according to the atypical element(s) or constituent(s) 
specified for the particular family. Within each group, one formulation 
is chosen to represent all of the member products in compliance with 
the registration requirements. Related costs may be shared by 
participating F/FA manufacturers within each group.
    Groups within the Baseline Categories. The baseline category for 
each defined fuel family contains a single F/FA group, with the 
exception of the baseline methanol category. As discussed above, the 
baseline methanol category includes two groups: the M100 group and the 
M85 group. The representative to be used in required emission 
characterization and health effects tests for each baseline group is 
the designated base fuel for the respective fuel family (see Section 
V). For example, all gasoline formulations meeting the gasoline 
baseline criteria are sorted into one group, to be represented in 
testing by the designated gasoline base fuel. The same holds true for 
diesel, ethanol, methane, and propane fuel families. In the case of 
methanol, baseline formulations are divided into two groups and testing 
is performed on two representatives, one for each of the designated 
baseline groups, i.e., M100 base fuel and M85 base fuel.
    Groups within the Non-Baseline Categories. Non-baseline categories 
are defined for each fuel family. F/FAs in non-baseline groups include 
products that comply with the baseline elemental composition 
restrictions for the respective fuel family, but do not meet 
quantitative limits on certain baseline components (e.g., oxygen 
content). Non-baseline groups are defined according to the 
constituent(s) that differentiate the fuel or additive/base fuel 
mixture from the baseline products in the respective fuel family. The 
representative for each non-baseline group is the member of the group 
with the highest concentration of the non-baseline constituent.
    (a) Gasoline. Gasoline formulations which comply with the baseline 
elemental composition criteria, except that they have a total oxygen 
content of 1.5 weight percent or more, are designated as non-baseline. 
These products are grouped according to the specific oxygenate compound 
(e.g., any specific alcohol, ether, or methanol/co-solvent combination) 
used to increase the oxygen content of the gasoline formulation. Thus, 
separate non-baseline groups are defined for ethanol, methyl tertiary 
butyl ether (MTBE), ethyl tertiary butyl ether (ETBE), tertiary amyl 
methyl ether (TAME), diisopropyl ether (DIPE), di-methyl ether (DME), 
tertiary amyl ethyl ether (TAEE), etc.
    In the NPRM, EPA had proposed to define additional non-baseline 
groups for fuels containing combinations of oxygenate compounds (or for 
which the registration contained multiple oxygenate additives), with 
separate groups defined for each combination recorded in a 
registration. However, EPA decided not to require the testing of 
oxygenate combinations in this final rule during the routine Tier 1 and 
Tier 2 testing program. EPA believes that the testing of fuels with 
individual oxygenates will satisfy the main objectives of the program 
by providing basic information about the potential health effects of 
particular oxygenated compounds in gasolines. Requiring routine testing 
of every recorded combination was judged unreasonable, as it resulted 
in a number of groups that did not reflect actual formulations in use. 
If there is concern about the toxicity of specific mixtures of 
oxygenated compounds, EPA may require additional testing under Tier 3 
on a case-by-case basis.
    An exception to this treatment of oxygenate combinations occurs in 
the case of non-baseline formulations containing methanol. Existing 
``substantially similar'' criteria currently limit the use of methanol 
as an oxygenate in gasoline to 0.3 percent by volume (i.e., 0.1 percent 
by weight), unless the formulation contains appropriate alcohol co-
solvents. Thus, methanol-containing gasoline formulations with 1.5 
weight percent oxygen or more must also contain a co-solvent. 
Accordingly, in the grouping system, each methanol and co-solvent 
combination used in gasoline formulations defines a different non-
baseline group [e.g, methanol and isopropyl alcohol (IPA), methanol and 
tertiary butyl alcohol (TBA), methanol and butanol, etc.]. Those 
oxygenate compounds used as co-solvents for methanol need to be 
identified as such in a fuel's registration. If an oxygenate is not 
identified as a methanol co-solvent, even if it appears in a fuel 
registration that also includes methanol, then EPA will assume that it 
defines a gasoline/oxygenate group separate from the methanol/gasoline 
mixture.
    Within each non-baseline gasoline group, a formulation consisting 
of the base gasoline fuel blended with the highest weight percent of 
the oxygenate or methanol/co-solvent combination registered for any 
member F/FA product will serve as the group representative that will be 
tested to comply with the program's requirements. The selection of the 
group representative is to be based on the highest actual 
concentration-in-use or the highest recommended concentration-in-use, 
whichever is the greater, for the particular oxygenate or oxygenate/co-
solvent blend.
    EPA recognizes that current fuel registration procedures allow 
manufacturers to include in the original registration a list of all the 
potential additives that might be used in the marketed fuel, along with 
the applicable range of concentration-in-use for each alternative. 
Under these circumstances, this final rule makes the non-baseline 
producer responsible for the testing of each oxygenate listed in the 
registration. For example, if a gasoline fuel registration lists 
methanol/co-solvent, ethanol, MTBE, and ETBE, then the manufacturer is 
responsible for separately testing each of four gasoline/oxygenate 
blends: gasoline-methanol/co-solvent, gasoline-ethanol, gasoline-MTBE, 
and gasoline-ETBE. The multiple testing responsibility can be satisfied 
by the manufacturer individually or by participating in four applicable 
groups. In each group, a formulation consisting of the base gasoline 
fuel blended with the highest concentration of the oxygenate listed for 
any member fuel or additive/base fuel mixture would serve as the group 
representative to be tested to comply with the program's requirements.
    The existing fuel registration procedures also allow manufacturers 
to report a range of concentration-in-use for each bulk additive listed 
as a potential component of the registered fuel. Thus, it is possible 
for the same registration to include formulations under both baseline 
and non-baseline definitions. If so, the manufacturer is responsible 
for testing formulations in both categories covered by the indicated 
range listed in the registration. In other words, if the reported range 
of concentration-in-use of an added oxygenate could include gasoline 
formulations with less than 1.5 weight percent oxygen as well as 
formulations with 1.5 weight percent oxygen or more, then the 
manufacturer is responsible for testing formulations in both baseline 
and non-baseline categories. For example, suppose a gasoline 
registration includes two potential oxygenates with respective 
concentration-in-use (shown here in terms of the resulting oxygen 
content in the formulation), as follows: ethanol (0 to 3.5 percent 
oxygen by weight) and ETBE (0 to 2.7 percent oxygen by weight). Because 
the indicated ranges include both baseline and non-baseline 
formulations, the manufacturer would be responsible for the testing of 
three formulations: baseline gasoline, a non-baseline gasoline-ethanol 
blend, and a non-baseline gasoline-ETBE blend. If the manufacturer 
chooses to participate in grouping arrangements, then he/she would be 
sharing the cost of the testing for the representative of each of these 
three groups.
    (b) Diesel. Non-baseline diesel formulations contain at least 1.0 
percent oxygen by weight. Non-baseline formulations include alcohol 
blends, ether blends, biodiesels (e.g., diesel-soy methyl ester blend), 
and other formulations containing oxygenating compounds. Separate non-
baseline groups are defined for each added alcohol or ether (e.g., 
methanol, ethanol, DME, etc.) and for other oxygenating compounds by 
class (e.g., peroxides, nitroso compounds, nitro compounds, alkyl 
nitrites, alkyl nitrates, animal-source alkyl esters, vegetable-source 
alkyl esters, furans, etc.).
    Diesel fuel manufacturers are responsible for the testing of each 
added alcohol, ether, or oxygenate class included in their fuel 
registration. For example, if the registration includes added methanol 
and soy methyl ester, the manufacturer will be responsible for testing 
two non-baseline formulations: (1) A diesel-methanol blend and (2) a 
diesel formulation containing a vegetable-source alkyl ester. In order 
to satisfy the testing requirements, the manufacturer may perform the 
tests individually or take advantage of the grouping provisions to 
share the testing costs with other manufacturers of similar products. 
In the above example, the manufacturer will be able to group with other 
manufacturers of diesel formulations containing methanol and with other 
manufacturers of formulations containing other vegetable-source alkyl 
esters (e.g., rape methyl ester).
    For each diesel non-baseline group defined by the presence of an 
alcohol, ether, or class of oxygenating compound, the representative to 
be used in testing will be a formulation consisting of the diesel base 
fuel blended with the highest actual or recommended concentration-in-
use of the particular alcohol, ether, or class of oxygenating compound, 
as recorded for any member of the group. For example, if manufacturers 
form a group of non-baseline diesel formulations containing vegetable-
source alkyl esters, the group representative will be a diesel 
formulation containing the highest volume percent of any of the 
vegetable-source alkyl esters represented in the group. The alkyl ester 
is to be added to the base diesel fuel for conducting the required 
emission characterization and toxicity tests.
    EPA recognizes that current registration procedures allow 
manufacturers to include in the original diesel fuel registration a 
list of all the potential oxygenating compounds that might be used in 
the marketed fuel, along with the applicable range of concentration-in-
use for each alternative. As with gasoline formulations, this final 
rule requires the diesel fuel producer to test each alcohol, ether, or 
class of oxygenate listed in the registration. Also, if a registration 
lists a range of oxygen content that defines both baseline and non-
baseline formulations, then the manufacturer is required to test both a 
baseline formulation and a non-baseline formulation.
    (c) Methanol. Non-baseline methanol formulations conform with the 
baseline limits in terms of elemental composition, but contain more 
than 4 percent by volume of substances other than methanol and 
gasoline. Individual groups are defined for each non-methanol, non-
gasoline component, and for each unique combination of such components. 
The representative of each non-baseline methanol group will be the 
group member with the highest concentration (i.e., percent by volume) 
of the non-methanol, non-gasoline component(s).
    (d) Ethanol. Non-baseline ethanol formulations conform with the 
baseline limits in terms of elemental composition, but contain more 
than 5 percent by volume of substances other than ethanol and gasoline. 
Individual groups are defined for each non-ethanol, non-gasoline 
component, and for each unique combination of such components. The 
representative of each non-baseline ethanol group will be the group 
member with the highest concentration (i.e., percent by volume) of the 
non-ethanol, non-gasoline component(s).
    (e) Methane. There is only one non-baseline methane group. This 
group contains all methane formulations conforming with the baseline 
criteria except that they exceed the allowable limit for non-methane 
hydrocarbons (i.e., 20 mole percent). The representative for the non-
baseline methane group will be the member formulation containing the 
highest concentration of non-methane hydrocarbons.
    (f) Propane. Non-baseline propane formulations are those which 
conform with the baseline criteria except that they exceed the 
allowable limit for non-propane hydrocarbons (i.e., 20 percent by 
volume). All non-baseline propane formulations are sorted into a single 
group. The representative for the non-baseline propane group will be 
the member formulation containing the highest concentration of non-
propane hydrocarbons.
    Groups within the Atypical Categories. Atypical groups within each 
fuel family are defined according to the distinctive atypical 
constituent(s). Separate groups are established for any single atypical 
constituent and any unique combination of atypical constituent(s) which 
occurs among the products in each category. For example, if a gasoline 
fuel contains sodium, and no other atypical element, then this atypical 
fuel will group with other gasoline fuels or additive/base fuel 
mixtures containing sodium as their only atypical constituent. However, 
if a gasoline fuel contains sodium and potassium, then this fuel will 
define a separate group for formulations containing both sodium and 
potassium. As explained previously, EPA believes that this approach is 
reasonable because different atypical elements may have distinct 
toxicological effects. Thus, while similarly composed F/FAs may group 
together, EPA believes that testing distinct F/FAs separately best 
effectuates CAA Section 211(e), which states that ``each'' F/FA shall 
be tested.
    Groups are further subdivided according to the presence of polymers 
containing atypical element(s) in their molecular structure. F/FAs 
containing polymers are considered atypical for a respective fuel 
family only if the F/FA product as a whole contains one or more 
atypical elements. If the polymer contains an atypical element as part 
of its molecular structure, then the atypical polymer defines a 
separate atypical group. For example, the presence of polyethylene in a 
gasoline product does not in itself make that product atypical because 
polyethylene contains no elements in addition to carbon, hydrogen, 
oxygen, nitrogen, and sulfur. On the other hand, if the gasoline 
product contains chlorinated polyethylene, then the product is 
considered atypical because of the chlorine content (chlorine is an 
atypical element for the gasoline fuel family). Such product could 
group with other atypical gasoline products containing chlorinated 
polymers. However, if the atypical gasoline product contains 
polyethylene and chlorine as two different components of the 
formulation, the product will group with other atypical gasoline 
products containing chlorine in non-polymer constituents.
    For groups defined by a single atypical constituent, the 
representative to be used in satisfying the group's testing 
requirements will be the member fuel or additive/base fuel mixture with 
the highest actual or recommended concentration-in-use of the atypical 
constituent. Within a group of such products containing a unique 
combination of two or more atypical elements, the representative shall 
be the product which has the highest total concentration of atypical 
elements. In the case that two or more products within such a group 
contain the same and highest concentration of atypical constituents, 
the process specified for selecting the representative gives precedence 
to the highest total concentration of the atypical constituents in the 
following priority order: (1) Total concentration of metals, (2) total 
concentration of halogens, (3) total concentration of other atypical 
elements (including sulfur, if applicable), (4) total concentration of 
polymers containing atypical elements, (5) total concentration of 
oxygen.
    As discussed previously, current fuel registration procedures allow 
manufacturers to include in their registration a list of potential bulk 
additives to be used in the fuel. As a result, registrations could 
include several additives containing one or more atypical constituents 
with the same purpose-in-use, but which are not intended by the fuel 
manufacturer to be used at the same time. If several additives for the 
same purpose-in-use are listed in a single registration, and if these 
additives contain different atypical elements, the manufacturer is 
responsible for testing each individual atypical additive separately. 
This means that each unique atypical additive listed in a registration 
for the same purpose-in-use will define a different testing group. On 
the other hand, if a fuel registration includes additives with 
different functions and different atypical elements, and if these 
additives are normally blended together in the same formulation, then 
the manufacturer is allowed to test them together (or to participate in 
an applicable group). For example, if a diesel fuel registration lists 
two atypical biocide additives, one containing boron and the other 
containing chlorine, the fuel manufacturer would then be responsible 
for testing two formulations (one diesel formulation containing boron 
and one diesel formulation containing chlorine). However, if the 
registration includes a boron-containing biocide and a chlorine-
containing detergent, then the manufacturer may test the two additives 
together.

C. Implementation of Grouping System and Cost-Sharing Provisions

    The grouping system included in this final rule allows 
manufacturers of similar F/FAs, on a voluntary basis, to pool their 
resources and efforts to satisfy the registration requirements. The 
primary objectives of the grouping system and cost-sharing provisions 
are to reduce the overall costs of the registration program and 
maximize the efficiency of the program by avoiding duplication of 
effort. The grouping and cost-sharing provisions included in today's 
rule are supported by CAA section 211(e)(3)(B), which permits 
manufacturers of similar F/FAs to share the testing costs of the 
program so that requirements can be met without duplication. Although 
this rule allows manufacturers to comply with the program's 
requirements by participation in a group, each manufacturer continues 
to be individually subject to this rule and responsible for testing 
under this rule.
    The practical implementation of the grouping system involves two 
major tasks: (1) The organization and administration of group 
functions, and (2) the development of equitable arrangements for cost-
sharing. Backed by its experience with respect to the TSCA testing 
program, EPA judges that the F/FA industry, under the aegis of its 
various trade associations or other third parties, is capable of 
accomplishing these tasks with little or no Agency assistance and 
interference. EPA's experience with cost-shared testing under TSCA 
regulations (40 CFR part 791) indicates that manufacturers prefer to 
work out their own cost-sharing arrangements, and EPA anticipates that 
F/FA registration applicants will likewise prefer to work out their own 
cost-sharing agreements. Public comments from the regulated industry 
support this assumption. Thus, EPA intends for manufacturers to work 
out cost-sharing agreements by themselves. However, if F/FA 
manufacturers cannot work out cost reimbursement, this rule allows F/FA 
manufacturers to use procedures similar to existing TSCA procedures 
[see Sec. 79.56(c) of this rule] for resolution of disputes.
    In addition to establishing cost-sharing mechanisms, F/FA 
manufacturers will also need to develop agreements concerning the 
division of responsibilities among group members for meeting the 
specific requirements of the registration program. EPA expects the 
participation of industry trade associations in the formation of groups 
and management of these activities. These associations should be able 
to establish ``third-party'' mechanisms whereby individual 
manufacturers can enroll their products in appropriate groups while 
minimizing the extent to which confidential data must be revealed. Each 
manufacturer needs to determine whether the grouping and cost-sharing 
advantages outweigh the possible competitive risks involved.
    In general, F/FA manufacturers should be able to determine the 
appropriate groups for their products without EPA involvement, 
according to the grouping criteria specified in this final rule, and to 
enroll their products into those groups. However, EPA recognizes that 
some Agency involvement might be needed in some special cases. When 
appropriate, based on EPA's discretion, the Agency will provide limited 
guidance for those manufacturers needing assistance with the 
application of the grouping criteria to their specific products.
    Manufacturers of F/FAs registered prior to the effective date of 
this rule are required to notify EPA within six months after the 
effective date of this rule if they intend to comply with the rule as 
part of a group and, if so, to identify the person or entity which is 
organizing the testing (see Section XII.A). In this case, groups of 
producers would organize prospectively to complete the same program 
requirements for their similar products and cost-sharing arrangements 
could be reached in advance of testing.
    Manufacturers of F/FAs not registered prior to the effective date 
of this rule are expected to conduct the required testing individually, 
unless they certify to EPA that they intend to rely on data to be 
submitted (and/or previously submitted) by an existing group or 
individual manufacturer of a similar registered product. The 
certification needs to include assurances that the original submitter 
has been notified (see Section XII.A for notification requirements) and 
that the manufacturer intends to comply with reimbursement as provided 
in this rule.
    Under the reimbursement provisions in this rule, there will be a 
fifteen years ``reimbursement period'' for the original submitter 
(individual or group) to obtain reimbursement from those manufacturers 
that rely on previously submitted data. This period has been lengthened 
from the originally proposed five years in response to public comments.

V. Base Fuel Specifications and Formulation Requirements

    In this final rule, EPA is establishing chemical and physical 
specifications to represent base fuel formulations for each defined 
fuel family. EPA has adopted the method proposed in the reformulated 
gasoline rulemaking (56 FR 31176), which uses sales-weighted averages 
of fuel survey data to determine national average chemical and physical 
parameters, to establish base fuel specifications for gasoline and 
diesel. Because comparable survey data are not available for 
alternative fuels, the base fuels for the alternative fuel families are 
based on CARB definitions and limited survey information.
    The generic base fuel formulations will function as archetypes of 
the F/FAs in each fuel family and will serve as the test substance or 
group representatives for the baseline group(s) for the respective fuel 
family. The use of consistently formulated base fuels will facilitate 
the comparison of the emission and health effect test results from the 
many fuel and fuel additive products within each fuel family. The base 
fuels will also serve as the fuel substrates into which additives 
undergoing evaluation will be mixed prior to emission generation and 
testing. Tests conducted on the emissions of the base fuel will then 
serve as controls against which tests on the emissions of the additive/
base fuel mixture will be compared.
    In addition to defining chemical and physical parameters for each 
base fuel, EPA is also specifying the allowable additive(s) to be 
included in the base fuel. EPA recognizes that commercial fuels 
typically contain additives to control fuel quality and enhance fuel 
performance, as well as to help in fuel production and distribution. 
Ideally, in order to better isolate the health effects associated with 
a particular additive or fuel, the base fuel would not contain 
additives unless they were the actual test subjects. However, several 
bulk additive types are common to most of the fuels within a given fuel 
family, and these should arguably be included as part of the base fuel. 
As a practical matter, it would be difficult in some instances to find 
a fuel that did not contain certain additive types used by refiners to 
facilitate production or distribution. EPA is thus requiring that base 
fuels contain a limited complement of the additives which are essential 
for the fuel's production or distribution and/or for the successful 
operation of the test vehicle/engine throughout the mileage 
accumulation and emission generation periods required under this rule. 
Since additives may have a substantial effect on emissions, for 
purposes of standardization it is important to specify the additive 
types which are to be contained in the base fuels. However, the 
selection of the specific product within each specified additive 
functional category is left to the formulator of the base fuel and/or 
the manufacturer responsible for the testing. Unless otherwise 
restricted, the presence of trace contaminants does not preclude the 
use of a fuel or fuel additive as a component of a base fuel.
    Additive requirements for each defined base fuel are discussed in 
the following sections. Additives used as base fuel components are to 
be added at the minimum treatment rate needed for effective 
performance. In contrast, additives to be tested must be mixed in the 
base fuel at the maximum in-use concentration recommended by their 
manufacturers.\19\ When a fuel additive is tested, any additive 
normally contained in the base fuel which serves the same function as 
the test subject additive must be removed from the base fuel 
formulation. For example, if a corrosion inhibitor is to be tested, 
this test additive would replace the corrosion inhibitor normally 
included as a component in the base fuel. This substitution requirement 
may preclude the use of certain multi-functional additives as base fuel 
components (in the case where the subject additive serves one of the 
functions of the multi-functional additive), since it would not be 
possible to replace a portion of a multi-functional additive with the 
test subject additive.
---------------------------------------------------------------------------

    \19\Special provisions related to the testing of additives are 
discussed in Section VI.F.

    Note: The specifications in the following sections describe the 
base fuel(s) for each fuel family, which serve the test fuel 
functions discussed above. These base fuel specifications are not 
the same as the criteria which permit F/FAs to join the baseline 
group within a fuel family. The baseline group criteria are provided 
in the preceding section of this preamble.

A. Gasoline

    For the gasoline base fuel, EPA is requiring the use of the 
reformulated gasoline summer baseline fuel as specified in CAA Section 
211(k)(10)(B)(i). This unleaded gasoline fuel, which is free of 
oxygenates, was determined from fuel survey data and will be used to 
represent all grades of conventional gasoline. This base fuel has the 
same specifications as the industry average gasoline used in many 
recent fuel emission studies, including the Auto/Oil Program\20\ and 
EPA's reformulated gasoline testing program. Selecting this formulation 
as the base gasoline fuel allows the comparison of emission 
characterization results from the F/FA testing program with a larger 
body of current emission data. The blending tolerances for the gasoline 
base fuel are consistent with certain blending tolerances specified in 
the RFG rule (59 FR 7716).
---------------------------------------------------------------------------

    \20\Auto/Oil Air Quality Improvement Research Program, Technical 
Bulletin #1, December 1990; available in Docket A-90-07, Item No. 
IV-A-08.
---------------------------------------------------------------------------

    The gasoline base fuel must contain the following additives: 
deposit control, corrosion inhibitor, demulsifier, anti-oxidant, and 
metal deactivator. In addition to the above required additives, the 
final rule allows manufacturers to use anti-static additives in the 
gasoline base fuel, if needed. Anti-static additives are not required 
in gasoline base fuel because this type of additives is not considered 
essential for the fuel's production, distribution, or the vehicle 
operation. Thus, anti-static additives should be used only as a safety 
measure on a case-by-case basis, as needed (e.g., when static problems 
present a risk of explosion). The required and permissible gasoline 
base fuel additives may contain no elements in addition to carbon, 
hydrogen, oxygen, nitrogen, and/or sulfur.
    In the Reopening Notice, EPA proposed to preclude the use of 
sulfur-containing additives in the gasoline base fuel. However, in 
response to a number of comments from the regulated industry, this 
final rule permits up to 15 ppm sulfur to be included in the additives. 
The total sulfur content in the base fuel, including any sulfur 
contributed by the additive components, must equal 339 ppm (within a 
tolerance of  25 ppm). A summary of the gasoline base fuel 
specifications and its additive components is provided in the 
accompanying regulations [see Table F94-1 in Sec. 79.55(b)].

B. Diesel

    Reflecting its predominant usage, #2 diesel is selected in this 
final rule as the base fuel for diesel. The specifications for the 
diesel base fuel were determined by calculating an industry average 
diesel fuel from 1990 industry and government diesel fuel survey data. 
The sources of data and methods of calculations are contained in the 
docket for this rulemaking.\21\ The blending tolerances for the diesel 
base fuel have been set to be comparable to those used in the gasoline 
base fuel. An exception to this general methodology is the base fuel 
specification for sulfur level. The required sulfur level (0.05 weight 
percent) reflects current on-road diesel fuel sulfur limits (55 FR 
34120).
---------------------------------------------------------------------------

    \21\See memorandum from James Greaves to Docket A-90-07 (Item 
No. IV-B-01) regarding ``Revised Base Diesel Fuel Determination 
Procedures for the Fuels and Fuel Additives Rulemaking.''
---------------------------------------------------------------------------

    The additives required as diesel base fuel components are: 
corrosion inhibitor, demulsifier, anti-oxidant, and metal deactivator. 
In addition to the above required additives, the final rule allows the 
use of anti-static and flow improver additives in the diesel base fuel, 
as needed. As with gasoline, anti-static additives are not required 
because they should only be used in the case of static accumulation 
problems. Similarly, flow improvers may be used on a need basis to 
improve cold weather handling.
    As in the gasoline base fuel, the diesel base fuel additives may 
contain sulfur, as well as carbon, hydrogen, oxygen, and nitrogen. The 
total sulfur content in the diesel base fuel formulation, including any 
sulfur contributed by the additives, may not exceed 0.05 percent by 
weight. A summary of the diesel base fuel specifications and allowed 
additive components is provided in the regulatory text [see Table F94-2 
in Sec. 79.55(c)].

C. Alternative Fuels

    EPA has used CARB definitions and other available information to 
establish base fuel specifications for each alternative fuel family 
(see Tables F94-3--F94-6 in Sec. 79.55). However, due to rapidly 
developing technology, the fuel additive package requirements for these 
fuels are not as well established as for gasoline and diesel. In fact, 
there is only limited information available on the additive 
requirements for the successful long-term operation of each alternative 
fuel/vehicle combination. Hence, it is the responsibility of the F/FA 
manufacturers who are required to test such base fuels (in consultation 
with EPA), to comply with the additive requirements of the manufacturer 
of the particular vehicle/engine used for the testing of alternative F/
FAs. If the manufacturer of an alternatively-fueled vehicle or engine 
specifies that additives (beyond those specified in the regulations), 
are essential for operation, then the F/FA manufacturer should submit a 
request to EPA to use those additional additives as components of the 
base fuel at the minimal effective level. EPA will publish a document 
in the Federal Register whenever approving such a request to modify a 
base fuel.
1. Methanol
    The methanol fuel family contains two fuel groups, one for M100 
fuels and one for M85 fuels. Each of these methanol groups has its own 
base fuel. These base fuels may only contain the elements carbon, 
hydrogen, oxygen, nitrogen, sulfur, and chlorine. The chlorine (as 
chloride) is permitted as a contaminant remaining from methanol 
production, and is limited to no more than 0.0001 percent by mass. The 
sulfur content may not exceed 0.002 percent by mass in the base M100 
fuel and may not exceed 0.004 percent by mass in the base M85 fuel.
    The M100 base fuel must consist of 100 percent chemical grade 
methanol by volume. The M85 base fuel is to contain 85 percent chemical 
grade methanol by volume, blended with 15 volume percent base gasoline 
fuel (meeting the gasoline base fuel specifications outlined in Section 
V.A., above). Specifications for the methanol base fuels are listed in 
Table F94-3 in Sec. 79.55(d) of the regulations.
    Some gasoline detergents have been shown to cause intake system 
deposits when used in M85 applications. Likewise, lubricating oils 
containing calcium have been shown to cause injector tip deposits in 
M100 applications. Therefore, EPA recommends that F/FA manufacturers 
determine the methanol compatibility of lubricating oils as well as 
fuel additives used in the gasoline portion of the M85 base fuel.
2. Ethanol
    The ethanol fuel family contains one group, represented by E85 base 
fuel. The E85 base fuel is to contain 85 percent chemical grade ethanol 
by volume, blended with 15 volume percent base gasoline. The ethanol 
base fuel may only contain the elements carbon, hydrogen, oxygen, 
nitrogen, sulfur, chlorine, and copper. The chlorine (as chloride) is 
permitted as a contaminant remaining from ethanol production, and is 
limited to no more than 0.0004 percent by mass. The sulfur content may 
not exceed 0.004 percent by mass. Copper, also a contaminant from 
ethanol production, is limited to 0.07 mg/L.
    Additives used in the gasoline component of E85 base fuel must be 
ethanol-compatible. The base fuel specifications for E85 are summarized 
in Table F94-4 in Sec. 79.55(e) of the regulatory text.
3. Methane
    The methane fuel family is represented by a natural gas base fuel 
whose specifications are within the proposed ranges for natural gas 
certification fuel (as proposed in 57 FR 52912). This base fuel may 
only contain the elements carbon, hydrogen, oxygen, nitrogen, and 
sulfur, with the sulfur limited to 16 parts per million (by volume). 
The methane base fuel must contain added odorant for leak detection 
purposes, used at a level such that at ambient conditions the fuel has 
a distinctive odor potent enough for its presence to be detected down 
to a concentration in air of not over \1/5\ (one-fifth) of the lower 
limit of flammability.
    In the Reopening Notice, EPA proposed that any sulfur in the 
methane base fuel be limited to that contained in the odorant additive. 
In response to public comment, this restriction has been removed; 
however, the total sulfur in the methane base fuel formulation, 
including that contributed by any additives, may not exceed 16 parts 
per million. The methane base fuel specifications are listed in Table 
F94-5 in Sec. 79.55(f) of the accompanying regulations.
4. Propane
    The propane fuel family is represented by a commercial LPG base 
fuel. The propane base fuel may only contain the elements carbon, 
hydrogen, oxygen, nitrogen, and sulfur, with the sulfur limited to 123 
ppm (by weight). The propane base fuel must contain added odorant, for 
leak detection purposes, at a level such that at ambient conditions the 
fuel has a distinctive odor potent enough for its presence to be 
detected down to a concentration in air of not over \1/5\ (one-fifth) 
of the lower limit of flammability. As in the case of the methane base 
fuel, the final rule does not require the sulfur in the formulation to 
be contained only in the odorant additive. Rather, the sulfur 
limitation applies to the fuel/additive mixture in combination. The 
propane base fuel specifications are listed in Table F94-6 in 
Sec. 79.55(g) of the regulatory text.

VI. Emission Generation

A. General Approach

    As part of the registration requirements, F/FA manufacturers are 
required to conduct a detailed characterization of the combustion and 
evaporative emissions of their products, as well as biological tests in 
which animals are exposed to these emissions. The next sections 
describe the methods specified in the rule for generating the emissions 
to be used in these chemical and biological tests.
    As proposed in the NPRM, combustion emissions are to be generated 
using applicable portions of the FTP.\22\ To control some of the 
inherent variability of FTP emissions generated under transient engine 
operation,\23\ this final rule requires the use of a mixing chamber or 
other apparatus (see Section VI.B.2). This is one of the approaches 
discussed for consideration in the Reopening Notice. EPA is permitting 
the use of either the engine dynamometer or the chassis dynamometer for 
emission generation during biological testing using FTP or FTP-
equivalent cycles. For the reasons discussed in the Reopening Notice, 
EPA has decided to require the use of non-catalyzed emissions (i.e., 
untreated exhaust emissions)\24\ for biological testing in order to 
assure that the test animals are exposed to the full range of emission 
species potentially resulting from the combustion of F/FAs. A brief 
summary of the rationale behind this decision is included below.
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    \22\Federal Test Procedure (FTP) are the standard exhaust and 
evaporative emissions test procedures described in 40 CFR part 86 
and used by EPA to certify new vehicles.
    \23\Transient engine operation is achieved by varying the engine 
speed and/or engine load, which typically results in an emission 
stream varying in quantity and composition over time.
    \24\Exhaust emission not subject to an aftertreatment device 
such as a functional catalyst or particulate trap.
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    It is important to keep in mind that the purpose of this program is 
not to test the effectiveness of emission control devices or to 
directly evaluate the emission performance of various vehicles and 
engines. Rather, it is to examine the potential toxicologic effects of 
the emissions produced by F/FAs in use. With modern emission control 
technology in place, most of the ambient air pollutant species 
attributable to automobile exhaust come from two sources: 
Malfunctioning vehicles (``high emitters'') and normal vehicles during 
their cold start period, when their engines run rich and their 
catalytic converters have not yet reached effective operating 
temperatures. The variety of emissions from these two important sources 
are not well represented by hot, catalyzed exhaust generated from well-
maintained, modern vehicles. Emissions during the cold-start include 
hundreds of organic chemical species which are generated before the 
catalytic converter reaches its effective temperature. Once the 
catalytic converter is warmed-up, its efficiency increases to the point 
where only a dozen or so simple compounds remain in readily measurable 
amounts in the catalyzed exhaust. Thus, the use of catalyzed exhaust in 
the biological testing program would exclude from the tests relevant 
emission species that could potentially be harmful to human health or 
the environment. In fact, laboratory animals would be exposed to only 
very few of the organic emission species associated with the combustion 
of the fuel or additive of interest. In contrast, the ambient air 
normally contains the full range of combustion emissions, since cold-
start emissions are continuously reintroduced and some ``high 
emitters'' are always in operation. Since humans experience long-term 
exposure to these emissions, EPA believes it is important that they be 
included in the test exposure atmosphere. EPA's analysis\25\ of non-
catalyzed emission data demonstrates that emissions that receive no 
aftertreatment represent a comprehensive aggregate of characteristic 
combustion products at enriched concentrations, including the species 
which may otherwise be emitted only during the cold start or by high-
emitting vehicles. In order to simulate emissions that include the full 
range of potential species produced in the combustion of F/FAs, EPA is 
requiring the use of non-catalyzed emissions for biological testing in 
this program.
---------------------------------------------------------------------------

    \25\See memorandum from Stephen Mayotte to Docket A-90-07 (Item 
No. IV-B-02) regarding ``Engine-out versus Tailpipe Emissions in 
Light-duty Vehicles.''
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    With the exception of exhaust after-treatment devices, this final 
rule requires that all normally required emission control equipment be 
present and fully operational on all test vehicles and heavy-duty 
engines used in the generation of non-catalyzed emissions. In order to 
maintain the appropriate operation of the exhaust system while 
obtaining non-catalyzed emissions, EPA requires the use of non-
functional aftertreatment devices (e.g., a blank catalyst with no 
catalytic wash coat) in order to simulate the back pressure, residence 
time, and mixing characteristics usually provided by normally 
functioning aftertreatment devices. Special emission generation 
allowances for the testing of specific additives which are introduced 
for use in conjunction with certain aftertreatment devices are 
discussed in Section VI.F.

B. Combustion Emission Generation

1. For Emission Characterization
    Manufacturers are required under Tier 1 to characterize the 
combustion emissions of their F/FAs. Depending on the fuel family in 
question, vapor-phase, semi-volatile, and particulate emissions may be 
required to be characterized.\26\ As discussed in Section VII.B., the 
emission characterization requirements include the measurement of 
hydrocarbons, carbon monoxide, oxides of nitrogen, particulates, 
aldehydes, ketones, alcohols, ethers, polycyclic aromatic compounds, 
and atypical products, as applicable.
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    \26\Examples of general sampling procedures for vehicle 
emissions are discussed in Schuetzle, D., ``Sampling of Vehicle 
Emissions for Chemical Analysis and Biological Testing,'' 
Environmental Health Perspectives, Volume 47, pp. 65-80, 1983.
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    Both untreated (non-catalyzed) and treated (tailpipe)\27\ emissions 
generated using FTP conditions are to be characterized. 
Characterization of the tailpipe emissions will allow comparison of 
emissions from the test F/FA product with results from other studies. 
Characterization of the non-catalyzed emissions will be used to 
identify the emissions to which animals will be exposed in the 
biological tests.
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    \27\Tailpipe emissions are emissions downstream from all 
normally present emission aftertreatment devices, i.e., catalytic 
converters and/or particulate traps.
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    Applicable FTP procedures to be used in generating emissions are 
specified in 40 CFR part 86. The Urban Dynamometer Driving Schedule 
(UDDS)\28\ and the Engine Dynamometer Driving Schedule (EDS)\29\ cycles 
of the FTP shall be used in the emission generation for light-duty 
vehicles and heavy-duty vehicles, respectively. The motoring portion of 
the heavy-duty test cycle can be eliminated, at the manufacturer's 
option, for the generation of emissions. This will allow the use of 
relatively inexpensive dynamometer equipment without compromising the 
value of the test.
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    \28\UDDS is a 1372 second transient speed driving sequence used 
by EPA to simulate typical urban driving. The UDDS for light-duty 
vehicles is described in 40 CFR part 86, Appendix I(a).
    \29\EDS is the transient engine speed versus torque time 
sequence commonly used in heavy-duty engine evaluation. The EDS for 
heavy-duty diesel engines is described in 40 CFR part 86, Appendix 
I(f)(2).
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    As discussed earlier, this final rule allows the use of a vehicle 
or engine for emission generation using FTP procedures. In the case of 
F/FAs normally used in light-duty vehicle applications, if an engine is 
to be used, the appropriate speed versus torque trace for the UDDS must 
be determined in a vehicle on a chassis dynamometer prior to emission 
generation. The engine used for emission generation in this testing 
program must then be operated under specific speed and torque 
conditions that simulate the UDDS.
    In light-duty vehicle testing, vapor phase emission samples are to 
be collected for each segment of the FTP cycle (i.e., Bag 1, Bag 2, and 
Bag 3). In addition, a semi-volatile sample and a particulate sample 
are to be collected during the driving cycle for light-duty vehicles. 
The heavy-duty testing procedure includes two tests: a cold-start test 
and a hot-start test. All three emission phases (i.e., vapor, semi-
volatile, and particulate) are to be collected for each heavy-duty 
test. Some modifications to the standard FTP may be required for 
collection of semi-volatile and particulate emissions, which are 
required for emission characterization and in-vitro biological testing 
(see next section). Special procedures may also be necessary in order 
to characterize emissions from F/FAs containing atypical elements. Good 
engineering and analytical chemistry practices should be followed while 
modifying the applicable test cycle for the collection of fractions not 
specified in 40 CFR part 86. Such modifications must be described in 
detail in the discussion of emission generation procedures to be 
included in the report provided to EPA, as discussed in Section XII.B.
    Vapor-phase emissions are to be collected and stored in Tedlar bags 
for subsequent chemical analysis. These emissions can be stored for 
only a limited period of time before chemical changes may occur. The 
critical time period is a function of the composition of the emissions, 
storage temperature and pressure, type of storage container, exposure 
to ultraviolet light, and the amount of deterioration that is 
considered acceptable. The maximum allowable storage times for 
emissions which are to be subjected to chemical analysis will vary 
depending on the speciation protocol, and are identified in relevant 
parts of the regulatory text.
    The particulate fraction may be collected on a single filter 
instead of on multiple filters as prescribed in the FTP. Although the 
filter collection procedures outlined in the CFR were designed for 
heavy-duty emission testing, these methods are applicable and can be 
used in light-duty applications as well. Similarly, semi-volatile phase 
emissions are to be collected on one apparatus for the entire driving 
cycle. Semi-volatile emissions are collected immediately downstream 
from the particulate collection filters using porous polymer beds or 
other equipment designed for their capture.\30\ After collection, the 
soluble organic fractions of the particulate and semi-volatile 
emissions are to be separately extracted using appropriate laboratory 
procedures.\32\ Because the extracted materials are much more stable 
than gaseous combustion emissions, they can be stored up to six months 
if protected from ultraviolet light and maintained at or below -20 
deg.C. Particulate phase emissions can be stored either on the 
collection filter or after extraction. Semi-volatile phase emissions 
must be extracted immediately after collection. The duration of the 
collection process which will be needed to obtain sufficient quantities 
of the test substance will vary depending on the emission 
characteristics of the engine and fuel or additive/base fuel mixture, 
and on the requirements of the biological test protocol. If an 
insufficient amount of particulate or semi-volatile material is 
obtained during a single driving cycle, the FTP may be repeated as 
required and the extracted organic fractions combined.
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    \30\An example procedure using a porous polymer resin as a 
trapping medium is described in Stump, F. et al., ``Trapping Gaseous 
Hydrocarbons for Mutagenic Testing,'' SAE Technical Paper Series No. 
820776, 1982; Available in Docket A-90-07 (Item No. II-J-14).
    \31\Examples of particulate and semi-volatile emission 
collection and analysis methods are described in 40 CFR 
Sec. 86.1301-1344 and in Coordinating Research Council Report No. 
551 (entitled ``Chemical Methods For The Measurement Of Unregulated 
Diesel Emissions--Carbonyls/Aldehydes, Particulate Characterization, 
Sulfates, PAH/NO2PAH,'' August 1987; available in Docket A-90-
07, Item No. II-J-15).
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2. For Biological Testing
    Non-catalyzed emissions are to be generated for conducting 
biological tests, following the same procedures described above for 
emission characterization.
    In vitro biological testing (i.e., the Salmonella assay) is to be 
conducted on extracts of the particulate and semi-volatile emission 
phases separately. Particulate and semi-volatile emissions are to be 
collected in a manner identical to the procedure used for particulate 
and semi-volatile emission characterization.
    The in vivo biological testing requires the generation of whole 
untreated emissions for a minimum of six hours per day, five days per 
week, for 13 weeks. To generate these emissions, light-duty vehicles 
(or engines) with non-functional after treatment devices (e.g., blank 
catalyst with no catalytic wash coat) are to be operated under FTP or 
FTP-equivalent engine conditions. The continuous generation of 
emissions throughout the required exposure period requires light-duty 
vehicles/engines to be driven through repeated UDDS cycles and heavy-
duty engines to be operated over repeated EDS cycles. If desired, 
registrants may automate their emission generation system.
    As discussed in the Reopening Notice, EPA was concerned about the 
inherent variability of FTP-generated emissions. To accommodate the FTP 
transient cycle within the biological testing program, this rule 
requires the use of an apparatus to provide a more stable exposure 
environment for biological testing. For this purpose, EPA recommends 
the development and use of a large dilution/mixing/integration chamber 
located between the constant volume sampling (CVS) system and the final 
dilution apparatus, just prior to the exposure chamber containing the 
test animals. The mixing chamber will allow the necessary adjustment of 
the exhaust concentrations and integration of the large concentration 
swings typical of FTP exhaust, prior to exposing the test animals. This 
chamber must meet certain performance specifications based on the 
average concentration of total hydrocarbons in the exhaust. That is, 
the average concentration of total hydrocarbons leaving the mixing 
chamber must be within ten percent of the average concentration of 
total hydrocarbons entering the chamber. Much of the CVS system 
concentration variability is associated with the rapidly changing 
dilution ratios that result from rapidly changing exhaust flow rates. 
EPA recognizes that vehicle exhaust sampling devices, such as mini- 
diluters,\32\ are being developed to maintain constant dilution ratios 
during transient testing. These systems will eliminate much of the 
concentration variability of classical CVS exhaust. As discussed in the 
Reopening Notice, these systems are currently under development and 
their use at this time is limited. However, today's rule will allow 
their use if they can meet the performance specifications defined above 
as well as other requirements of the testing program.
---------------------------------------------------------------------------

    \32\A discussion on mini-diluter technology can be found in: 
American Industry/Government Emissions Research (AIGER) Cooperative 
Research and Development Agreement, ``Specifications for Advanced 
Emissions Test Instrumentation,'' AIGER PD-94-1, Revision 5.0, 
February 1994; available in Docket A-90-07, Item No. IV-A-09.
---------------------------------------------------------------------------

    The combustion emissions generated for animal testing are to be 
diluted prior to delivery to the test animals. The CVS system, commonly 
used to condition exhaust for sampling and analysis, provides for 
controlled ambient air dilution of the combustion emissions. However, 
water condensation can be a problem during CVS system sample 
conditioning, depending upon vehicle fuel consumption and fuel economy, 
dilution air humidity, and exhaust/diluent ratio.\33\ The use of pre-
dried dilution air will lower the sample humidity, thus permitting 
lower dilution ratios and higher concentration of hydrocarbons to be 
achieved without condensation of water vapor. The minimum dilution 
ratio will vary with fuel composition. For example, a minimum dilution 
ratio of about 1:5 raw exhaust (dewpoint about 125  deg.F) with dry, 
clean filtered air is expected for gasoline fuels to reduce the water 
concentration to a dewpoint of about 68  deg.F. The minimum dilution 
ratio (maximum exhaust flow rate) occurs at about 200 seconds into the 
UDDS transient driving cycle. The dilution ratio is expected to be 
greater for methanol, ethanol, and natural gas fuels than for gasoline 
fuels because the exhaust water concentrations are greater with these 
alternative fuels. Heated transfer ducts or tubing can be used to avoid 
water condensation in much of the system, but the dilution/mixing/
integration chamber will generally be at or near laboratory temperature 
(about 70  deg.F), and CVS dilution will have to be adequate to assure 
that the cumulative integrated chamber dew point remains below 
laboratory temperature at all times.
---------------------------------------------------------------------------

    \33\An example procedure on how to deal with water vapor 
condensation problems is found in Black and Snow, ``Constant Volume 
Sampling System Water Condensation,'' SAE 940970, 1994. This paper 
describes a ``spreadsheet'' procedure for detailed, second by 
second, determination of diluted exhaust dew point and the necessary 
CVS system flow rates to avoid water vapor condensation.
---------------------------------------------------------------------------

    After initial dilution to preserve the character of the emissions, 
the exhaust stream may be further diluted to achieve the desired 
biological exposure concentrations. In testing the emissions of a 
particular fuel or additive/base fuel mixture, a manufacturer shall 
determine an optimum range of dilutions with which to characterize the 
health effects of the test substance. The range of dilutions shall 
include, at a minimum, an overtly or highly toxic concentration, a 
minimally toxic or non-toxic concentration, and a concentration of 
emissions having an intermediate level of toxicity. The selected 
concentrations must allow the determination of a concentration-response 
relationship (see Section VIII.A.3). EPA recommends that manufacturers 
review available literature for information on the design of inhalation 
studies.\34\
---------------------------------------------------------------------------

    \34\An example reference is Phalen, R. F., ``Inhalation Studies: 
Foundations and Techniques,'' CRC Press, Inc., Boca Raton, Florida, 
1984.
---------------------------------------------------------------------------

    One important factor to consider in determining the exposure 
concentrations or dilutions is the effect of carbon monoxide (CO) 
concentration in test animals. The CO concentration in the emissions is 
expected to be a limiting factor in establishing the appropriate 
dilutions for the testing of F/FAs. Anoxia, among other negative health 
effects from this combustion product, may mask the more subtle health 
effects of F/FA emissions. EPA recommends that manufacturers review 
available literature on previous toxicity studies for information on 
appropriate CO concentrations that have been used in the exposure of 
laboratory animals to automobile emissions.\35\\36\\37\
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    \35\Stara et al., ``Long-Term Effects of Air Pollutants in 
Canine Species,'' EPA/600/8-80/014, 1980.
    \36\Brightwell, J. et al., ``Neoplastic and Functional Changes 
in Rodents after Chronic Inhalation of Engine Exhaust Emissions,'' 
In: Ishinishi, N. et al., (eds), Carcinogenic and Mutagenic Effects 
of Diesel Engine Exhaust, Elsevier Science Publishers, Amsterdam, 
pp. 471-485, 1986; available in Docket A-90-07, Item No. IV-A-17.
    \37\Pepelko, W. E. et al., ``Effect of 90 Days Exposure to 
Catalytically Treated Automobile Exhaust in Rats,'' Environmental 
Research, Volume 19, pp. 91-101, 1979.
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3. Verification Testing
    A number of mechanisms can cause emissions to be captured in the 
dilution and sampling system before they can be characterized or used 
for animal exposures. Verification testing is required to determine the 
ratio (``recovery factor'') of emissions that exit the sampling system 
to those that enter the system. This ratio must be high in order for 
subsequent emission testing to be meaningful.
    EPA requires testing to verify the exposure atmosphere and to 
monitor the performance of the dilution/sampling system and mixing 
chamber, ensuring the repeatability of test results. Verification 
testing of the dilution/sampling system must be accomplished by 
injecting a known sample at the inlet and measuring the amount that 
exits the sample probe. For example, an injected hydrocarbon sample 
could be detected with a gas chromatograph and flame ionization 
detector to estimate the recovery factor. Similar verification 
procedures apply to the verification testing of the mixing chamber. 
Verification procedures for the dilution/sampling system and mixing 
chamber are included in Sec. 79.57(e)(2)(v) of the accompanying 
regulations. Additional requirements include the monitoring of 
conditions (e.g., air flow, CO levels, etc.) in the inhalation exposure 
chamber and verification of test animal exposure levels (see 
Sec. 79.61).

C. Evaporative Emission Generation

    Section III.A.2 discusses the RVP criteria which determine the 
applicability of evaporative emission testing to specific fuels and 
additive/base fuel mixtures. Evaporative emissions from in-use vehicles 
include diurnal, hot soak, resting and running loss emissions, and 
refueling emissions. However, to simplify the generation and collection 
procedures and to supply evaporative emissions of sufficient 
concentration for biological exposure testing, today's rule requires 
that evaporative emissions be generated using an evaporative emission 
generator (EEG). Emissions to be used both for characterization tests 
and biological exposure tests are to be generated in this way. The EEG 
is a fuel tank or vessel to which heat is applied to cause a portion of 
the fuel or additive/base fuel mixture to evaporate at a desired rate. 
Manufacturers will have flexibility in the design of the EEG used to 
test their particular F/FA. The size and/or number of EEG units to be 
used for evaporative emission testing will depend on the rate of 
emissions needed for the inhalation study. The vapor pressure of the F/
FA product may influence the required tank size, as well. Emission rate 
modifications shall not be adjusted by temperature control, since 
emission composition is sensitive to temperature changes.
    In general, the composition of evaporative emissions from vehicles 
does not resemble fully-evaporated whole samples of raw fuels or fuel 
additives. This phenomenon is due to differences in the vapor pressure 
of the fuel or fuel additive components and the effects of evaporative 
emission control equipment. To simulate this phenomenon with the EEG, 
procedures are to be followed to ensure that the evaporated fraction 
contain a reasonable representation of potential evaporated emission 
compounds. The EEG will be run at 1305  deg.F and will be 
equipped with a drain. The fuel will be drained and replenished 
periodically in order to maintain a constant composition and prevent 
the build-up of heavier compounds in the non-evaporated portion. The 
concentration of emissions of the evaporated fuel or additive/base fuel 
mixture in the vapor space of the EEG during the time emissions are 
being withdrawn for testing shall not vary more than ten percent from 
the equilibrium concentration in the vapor space of emissions generated 
from fresh fuel or additive/base fuel mixture in the evaporative 
chamber.
    EPA recognizes that other methods may also be suitable for 
generating F/FA evaporative emission mixtures for the testing purposes 
of this program. One possible alternative method was suggested in a 
comment received by EPA in response to the Reopening Notice.\38\ Based 
on the distillation properties of the test formulation, the suggested 
method would involve the distillation, condensation, and storage of the 
light-end components of the test fuel mixture, with revaporization of 
this whole fraction to generate test atmospheres. Other alternatives 
may also be valid. To accommodate these potential alternatives, the 
final rule contains a provision (see Sec. 79.57(f)(5) of the 
accompanying regulations) which permits manufacturers to request 
approval for methods other than the EEG for generating evaporative 
emission test atmospheres. To be granted, such requests must include 
supporting information which demonstrates (among other requirements) 
that the proposed procedures will generate emissions reasonably similar 
to in-use evaporative emission mixtures and that the generated 
emissions will be sufficiently concentrated to be useful in the context 
of toxicology tests. Approved procedures will be placed in the public 
docket.
---------------------------------------------------------------------------

    \38\Comments of the American Petroleum Institute on U.S. EPA's 
Fuels and Fuel Additives Registration Regulations, March 28, 1994 
(Item IV-D-49 in Docket A-90-07).
---------------------------------------------------------------------------

    For applicable F/FAs, evaporative emission characterization 
requirements include the measurement of total volatile organic 
compounds with speciation of the hydrocarbon compounds, alcohols, 
ethers, and atypical compounds. Characterization requirements are 
discussed in Section VII.B.
    For biological testing, evaporative emissions will be diluted and 
routed to the animal chambers in a manner similar to the method used 
for combustion emissions health effects testing, as described in the 
previous section, except that a mixing chamber is not required. The 
rate of emission generation shall be high enough to supply the 
biological exposure chamber with sufficient emissions to allow for a 
minimum of fifteen air changes per hour.
    The concentration of total hydrocarbons in the evaporative emission 
stream routed to the biological exposure chambers is to be diluted to 
three separate concentrations to establish a range of responses similar 
to combustion emission testing (see previous section). Evaporative 
emissions are not constrained by CO, NOX, or CO2 levels, and 
hence can be used at higher concentrations than combustion emissions. 
Verification testing is required for evaporative emissions in a manner 
analogous to the verification testing performed for combustion 
emissions.

D. Vehicle Selection

    EPA is requiring that new vehicles or engines be used for the 
combustion emission generation and testing of F/FAs to avoid the carry-
over effects from previously used fuels. All F/FAs must be tested in 
vehicles or engines (corresponding to chassis or engine dynamometer 
testing, respectively) that have been operated exclusively on the fuel 
or additive/base fuel mixture to be tested.
    EPA is also requiring that vehicles and engines used for the 
testing of
F/FAs be unaltered from original equipment manufacturer (OEM) 
specifications (with the exception of modifications in aftertreatment 
devices as described in Section VI.A). Rebuilds and alteration kits 
will only be allowed upon EPA's approval, when a F/FA manufacturer 
demonstrates to EPA that OEM equipment suitable for their F/FA 
product's testing is unavailable.
    As proposed in the NPRM (57 FR 13192-13193), vehicle and engine 
selection must follow the criteria outlined in Sec. 79.57(a) of the 
accompanying regulations. The selection method is described in detail 
in a memorandum entitled, ``Vehicle Selection Procedures for the 
Proposed Fuels and Fuel Additives Rulemaking'' (see Item No. II-B-6 in 
the public docket of this rule). As proposed, the final rule does not 
differentiate between light-duty vehicles and light-duty trucks. Thus, 
vehicles/engines are separated into two classes: light-duty and heavy-
duty. The vehicle or engine selected must be a new vehicle or engine of 
the model year in which testing begins. However, vehicle selection 
criteria are to be based on technology characteristics of the previous 
model year. Any one of the top five selling models (based on sales 
figures from the year prior to testing) with the appropriate technology 
in a fuel group may be chosen. Each test vehicle or engine must be 
equipped with all of the normally required and functioning emission 
control equipment, with the exception of aftertreatment devices, when 
applicable (see Section VI.A).
    Considering the practical constraints of the rule, EPA is requiring 
that only one vehicle or engine model be used to generate emissions for 
these tiers. Although EPA recognizes that emission composition is 
somewhat dependent on vehicle models and may even vary in replicate 
tests of the same vehicle/fuel combination, the use of untreated 
exhaust in the testing program will greatly reduce the significance of 
these potential sources of variability. The purpose of the testing 
program is to determine potential health effects of
F/FA emissions and not to establish in-use fleet average emission 
levels for different types of vehicles. However, EPA reserves the right 
to require the testing of F/FAs in additional vehicles or engines, 
under Tier 3, if there is concern for technology-based differences in 
toxicological effects. Furthermore, EPA could require the use of 
catalyzed exhaust to perform tests under Tier 3.
    Although EPA is routinely requiring only one vehicle or engine for 
the testing of F/FAs, EPA foresees that at least one backup vehicle/
engine of the identical model may be needed to replace vehicles/engines 
that wear out or malfunction during the course of testing. The 
probability of needing a replacement vehicle or engine increases in the 
case of testing F/FAs containing atypical elements that require 
additional mileage accumulation (see next section). The decision 
concerning the timing of vehicle and engine replacements is the 
responsibility of the F/FA manufacturer seeking registration. EPA 
recommends that backup vehicles/engines (if present) accumulate mileage 
along with the primary test vehicle, so as to minimize testing 
interruptions if the backup vehicle/engine is needed. Manufacturers 
may, at their own discretion, alternate between backup vehicles (or 
engines) during testing to further decrease the probability of problems 
or interruptions. Similarly conditioned vehicles/engines (i.e., primary 
and backup vehicle/engine) would be expected to generate comparable 
emissions. Emissions from backup vehicles/engines must have their 
emissions characterized prior to use in the biological studies. Wide 
discrepancies between the emissions of primary and backup vehicle/
engine emissions may be cause to void a test.
    During emission generation, vehicles and engines must be maintained 
in good condition by following the OEM recommendations for service 
schedule and parts replacement. If unscheduled maintenance becomes 
necessary, the vehicle or engine must be repaired to OEM 
specifications, using OEM or OEM-approved parts. In addition, the 
manufacturer is required to measure the basic emissions (as described 
in Section VII.B.2.a) after the unscheduled maintenance and before 
resuming testing, to demonstrate that the post-maintenance emissions 
are within 20 percent of pre-maintenance emission levels. If the basic 
emissions cannot be brought within 20 percent of their previous levels, 
then the manufacturer must restart testing using a new vehicle or 
engine. Provisions in the regulations allow for a limited amount of 
emission generation disruption without voiding the biological test.

E. Mileage Accumulation

    New vehicles (or engines) to be used in emission generation for the 
testing of F/FAs are required to undergo a break-in period in which the 
vehicle (or engine) is run exclusively on the fuel or additive/base 
fuel mixture to be tested. The mileage accumulation requirements of 
this final rule follow the approaches discussed in the Reopening 
Notice. These requirements serve the purpose of stabilizing the 
emissions from the new vehicle or engine.
    Vehicles to be used in the evaluation of baseline and non-baseline 
F/FAs are required to accumulate at least 4,000 miles prior to emission 
testing. For engines operated on an engine dynamometer, the minimum 
break-in requirement is 125 hours of operation for testing baseline and 
non-baseline F/FAs. The 4,000 mile/125 hour mileage accumulation 
requirements are consistent with the emission stabilization procedures 
used in EPA's new vehicle certification program.\39\ An intact 
aftertreatment device must be used when accumulating mileage in the 
evaluation of baseline and non-baseline F/FAs. Mileage can be 
accumulated in a number of ways, i.e., on a test track, on a 
dynamometer, on the street, or as part of a manufacturer's fleet. No 
specific driving cycle is required, but it must include a reasonable 
amount of transient operation.
---------------------------------------------------------------------------

    \39\40 CFR 86.094-26. Mileage and service accumulation; emission 
requirements.
---------------------------------------------------------------------------

    For atypical F/FAs, the minimum mileage accumulation required prior 
to testing is also 4,000 miles for test vehicles or 125 hours for test 
engines. After completion of the 4,000 mile/125 hour minimum mileage 
accumulation, an attempt should be made to identify and measure the 
atypical element(s) in the emissions. Mileage accumulation must 
continue until either: (1) 50 percent or more of the input mass of each 
atypical element is measured in the emissions (i.e., vapor, semi-
volatile, and particulate combined), or (2) a maximum mileage 
accumulation equivalent to 40 percent of the average useful life of the 
applicable vehicle/engine (e.g., 40,000 miles for light-duty vehicles, 
116,000 miles for heavy-duty vehicles, or engine equivalent) has been 
reached. When either of these conditions has been met, emission 
generation can begin for purposes of emission characterization or 
biological testing. Because the presence of atypical species in 
specific emission fractions will be dependent on the nature of the 
particular atypical element, EPA recommends examination of all emission 
fractions (i.e., vapor, semi-volatile, and particulate). The 
determination of the appropriate intervals for conducting emissions 
measurements is left to the manufacturer's discretion.
    Manufacturers of atypical F/FAs may choose to accumulate the 
required mileage using a vehicle/engine equipped with either an intact 
aftertreatment device or with a non-functional aftertreatment device 
(e.g., a blank catalyst without its catalytic wash coat).\40\ However, 
the sampling and analysis of emissions for detecting the atypical 
element(s) of interest, prior to emission characterization or 
biological testing, must be done with a non-functional aftertreatment 
device. A brief period of warm-up driving (i.e., 10 miles or equivalent 
time) needs to precede the sampling for the detection of atypical 
element(s).
---------------------------------------------------------------------------

    \40\If the manufacturer chooses to accumulate mileage without a 
functional aftertreatment device, and if the manufacturer wishes to 
do this outside of a laboratory/test track setting, then a 
memorandum of exemption for product testing must be obtained by 
applying to the Director of the Field Operations and Support 
Division [see Sec. 79.51(e)(6)(iv) of this rule].
---------------------------------------------------------------------------

    During the mileage accumulation period for the testing of any fuel 
or fuel additive, vehicles and engines used for emission generation 
must be maintained in good condition by following the recommended 
maintenance practices in the appropriate vehicle or engine owner's 
manual. Maintenance requirements were described in the previous 
section.

F. Special Requirements for Additives

    This section describes provisions for the testing of specific types 
of additives that might require modification in emission generation 
requirements. These include additives used in conjunction with 
aftertreatment devices, additives used infrequently, and diesel 
additives produced exclusively for use in diesel #1 fuels.
    As explained in the above sections, EPA is requiring that emissions 
used for Tier 1 and Tier 2 testing be generated from vehicles/engines 
with non-functional aftertreatment devices. In response to comments 
received on the Reopening Notice, however, EPA is including in this 
final rule a special allowance for specific types of additives that are 
designed to work in conjunction with aftertreatment devices. In the 
case of fuel additives specifically intended to enhance the 
effectiveness of exhaust aftertreatment devices, the related 
aftertreatment device may be used on the emission generation vehicle/
engine during all mileage accumulation and testing.
    Regarding infrequently used additives, EPA recognizes that some 
aftermarket additives are intended by the manufacturer (as stated in 
the additive's instructions for use) to be added to the fuel tank only 
at infrequent intervals. During mileage accumulation, these types of 
additives may be applied according to the manufacturer's 
specifications. However, during emission generation and testing, EPA 
requires that each tankful of fuel used contain the fuel additive at 
its maximum recommended level. In the case of bulk additives used 
intermittently for the direct purpose of conditioning or treating a 
fuel during storage or transport, or for treating or maintaining the 
storage, pipeline, and/or other component of the fuel distribution 
system (and not the vehicle/engine for which the fuel is ultimately 
intended), EPA also requires that the additive be added to the base 
fuel at the maximum concentration recommended by the additive 
manufacturer for treatment of the fuel or distribution system 
component. If the manufacturer of infrequently used aftermarket or bulk 
additives (as described above) is concerned that the test vehicle/
engine may be adversely affected and/or the emissions may be subject to 
artifacts due to overuse of these types of additives, then that 
manufacturer may submit a request for a modification in test procedure 
requirements. Any such request must include test data (e.g., emission 
characterization data) to support the claim that procedural 
modification is needed, as well as a suggested substitute procedure.
    In order to simplify diesel additive testing and allow 
comparability between the test results of all diesel additives 
evaluated in this program, EPA requires that all diesel additives 
(including those produced exclusively for use in #1 diesel fuels) be 
tested on the #2 diesel base fuel (specified in Section V). If a 
manufacturer is concerned that the emissions generated using a blend of 
their #1 diesel fuel additive with the #2 diesel base fuel may be 
subject to artifacts due to this blending, then that manufacturer may 
submit a request for a modification in test procedures. Any such 
request must include supporting data (e.g., emission characterization 
data) and suggested test modifications.

VII. Tier 1 Requirements

    The scope of Tier 1 encompasses: (1) a literature search for 
available information on the composition and effects of F/FA emissions 
on public health and welfare, (2) a chemical analysis to characterize 
the emissions of fuels or additive/base fuel mixtures, and (3) a 
qualitative discussion of potential exposures using information on 
total production volume and market distribution patterns of the 
particular fuel(s) or additive/base fuel mixture(s).

A. Literature Search

1. Scope
    The registration program requires
F/FA manufacturers to conduct a comprehensive data search that will 
include all relevant existing information concerning previous emission 
characterization and health effects and welfare studies. The data 
search must address the chemical composition and potential adverse 
effects of whole combustion emissions, relevant combustion emission 
fractions (e.g., particulate phase), and whole evaporative emissions, 
as applicable. The literature search must also address each of the 
individual combustion and evaporative (where different) emission 
products identified by the required emission speciation procedures, 
with the exception of carbon monoxide, carbon dioxide, nitrogen oxides, 
benzene, 1,3-butadiene, acetaldehyde, and formaldehyde. Special 
literature search requirements for non-baseline and atypical F/FAs are 
described in Section VII.A.3.
    Information considered applicable to a given fuel or additive 
includes data obtained from the testing of emissions from the fuel or 
additive in question or from other similar products. For this purpose, 
``similar'' products are those which meet the criteria for enrollment 
in the same F/FA group as the subject fuel or additive, pursuant to the 
grouping system criteria discussed in Section IV. F/FA manufacturers 
who choose to participate in the grouping system may pool information 
about all member products for purposes of their joint submission and 
may also make use of available data on other products which are not 
enrolled in the group but share the designated formulation 
characteristics of group members. Similarly, a manufacturer who chooses 
not to participate in the grouping system could include any test 
results which may be available for products which could theoretically 
be assigned to the same group as the manufacturer's own product.
    The survey on health effects studies is not restricted to the 
particular endpoints and experimental protocols included in Tier 2. 
Studies using other scientifically acceptable methods or protocols 
addressing all health effects of F/FA emissions must also be included 
in the Tier 1 report. Most often, data will be available from 
experiments conducted with laboratory animals, but other applicable 
studies must also be considered. Evidence for potential toxicity or 
lack of toxicity in exposed humans may be available from 
epidemiological studies, clinical studies, occupational exposures, or 
case reports. In general, referenced experiments must be concerned with 
the health effects of inhalation exposure to F/FA emissions (combustion 
and evaporative). However, data collected from relevant studies using 
other routes of exposure must also be included. Available results from 
in vitro tests, comparative metabolism studies, and structure-activity 
analyses are also considered relevant and must be included in the 
summary report for health effects of F/FA emissions.
    The data search must include available literature on welfare 
effects, including, but not limited to, the exposure and response of 
plants and animals to whole emissions and individual components of 
emissions, the potential for bioaccumulation, and the concentration and 
persistence of emission products in the air, soil, and water. Available 
results of exposure modeling analyses, environmental and atmospheric 
fate modeling studies, field studies, monitoring studies, accident 
evaluations, or environmental simulation experiments must be included 
to characterize potential exposures and the environmental impact of F/
FA emissions. Specific ecological studies addressing the potential 
environmental effects of F/FA emissions on vegetation, livestock, 
wildlife, aquatic species, and soil organisms must be included. In 
addition, the data search must address the welfare effects of F/FA 
emissions concerning their contribution to odor and visibility 
nuisances.
    Both public and in-house available sources must be included in the 
literature survey. Information on the health and environmental effects 
of
F/FAs is to be compiled from peer- reviewed scientific journals and 
other literature as well as internal industry studies, government-
sponsored reports, proceedings of scientific meetings, and other 
documented sources. In general, EPA will place greater confidence in 
studies that have been subject to peer review. A search of appropriate 
commercially available chemical, toxicological, and environmental data 
bases must be conducted to obtain information from published sources. 
An example list of commercially available data bases that may be used 
to obtain information on potential health and environmental effects, as 
well as environmental fate data, is available in the public docket of 
this rule.\41\
---------------------------------------------------------------------------

    \41\See memorandum from Ines del C. Figueroa to Docket A-90-07 
(Item No. IV-B-03) regarding ``List of Data Bases.''
---------------------------------------------------------------------------

    In the NPRM, EPA proposed that literature searches cover at least 
fifteen years. However, in response to public comments, EPA has 
increased this time period to cover at least thirty years prior to the 
date of submission, so that important information from earlier testing 
will not be omitted. In addition, literature searches must be current 
as of six months prior to the beginning of testing. The thirty years 
are not meant to be an absolute limit for data collection. EPA 
encourages F/FA manufacturers to do a comprehensive search that will 
include all relevant available information, regardless of the age of 
the data.
    The information to be submitted to EPA as a result of the data 
search includes the following items: (1) Brief text summary of the 
general findings and conclusions, including references, (2) a printed 
copy of the outputs from the data base searches, including reference 
list and associated abstracts, (3) complete documentation in scientific 
journal format of unpublished in-house or other privately-conducted 
studies, and (4) tables summarizing the protocols and results of all 
cited studies, organized by health or environmental endpoint and type 
of emissions (e.g., whole combustion emission, individual emission 
product). In addition, the person(s) or contractor(s) conducting the 
literature search and summary must be identified. Further discussion on 
the reporting requirements of this final rule is included in Section 
XII.
2. Adequate Existing Information
    The primary purpose of the literature search is to provide EPA with 
a comprehensive survey of the available data on health and welfare 
effects of
F/FAs. A secondary function of the literature search is to enable F/FA 
manufacturers to document the extent to which the emission 
characterization in Tier 1 and/or the evaluation of health effects 
included in Tier 2 have already been addressed by previous adequate 
testing and/or analysis. If adequate testing/analysis exists, F/FA 
manufacturers may submit such previous data in compliance with the 
requirements of the registration program. For example, if previous 
emission characterization studies addressing the speciation 
requirements of this program are available in the literature, then F/FA 
manufacturers may submit those studies in lieu of new characterization 
tests. Similarly, F/FA manufacturers could use the literature search to 
determine the availability of adequate biological tests in compliance 
with Tier 2 requirements.
    To satisfy the testing requirements of Tier 1 and/or 2 with 
previously conducted studies, reports of such previous tests must be 
sufficiently detailed to allow EPA to judge the adequacy of protocols, 
techniques, experimental design, statistical analyses, and data 
interpretation. Documentation must be sufficient to determine if the 
previously conducted studies were performed in a manner consistent with 
generally accepted scientific principles, good laboratory practices, 
and the specific testing guidelines in question. The age of the data 
will be considered but will not be the ultimate determining factor in 
deciding if an existing study is adequate. Although changes in 
technological approaches and methodology might preclude the use of some 
older studies, EPA recognizes that older literature can be useful for 
the purposes of this program. Thus, the quality of the study will be 
the deciding factor in determining the adequacy of existing studies, 
not the age per se. Additional criteria to be used in determining the 
adequacy of existing data/studies in relation to Tier 2 compliance are 
provided in Section VIII.C.
3. Special Requirements for Non-Baseline and Atypical F/FAs
    EPA recognizes that many of the individual chemical species that 
will be present in the emissions of non-baseline and atypical F/FAs 
will also be present in the emissions of baseline products in the same 
fuel family. Non-baseline formulations, as defined in this rule, 
contain the same elements as baseline formulations. Thus, on a 
qualitative basis, the emission products from non-baseline F/FAs are 
expected to overlap with those of baseline F/FAs in the same fuel 
family. For atypical F/FAs, the main differentiating characteristic is 
the presence of atypical element(s) which are not included in the 
baseline category for a particular fuel family. The composition of the 
emissions for atypical products, therefore, is expected to consist 
mainly of those species present in the emissions of baseline
F/FAs (for the same fuel family), with the addition of compounds which 
host the specific atypical element(s) of interest.
    In addition to requiring literature data on the potential health 
and welfare effects of the whole combustion and evaporative (where 
different) emissions of the particular F/FA product, Tier 1 also 
specifies that a literature search be conducted on each of the emission 
products of the tested fuel or additive/base fuel mixture. Because of 
the substantial overlap in the emission species of F/FAs in different 
categories within the same fuel family, however, this requirement could 
result in significant duplication of effort and waste of resources. To 
avoid this outcome, as authorized under CAA section 211(e)(3)(C), this 
final rule allows manufacturers of non-baseline and atypical F/FAs to 
limit the literature search done for individual emission species to 
only those compounds which are different from the compounds typically 
present in the emissions of baseline F/FAs for the same fuel family.
    In order to take advantage of this reduction in requirements, 
manufacturers of non-baseline or atypical F/FAs must compare the 
emission characterization results of their products with emission 
characterization data for baseline F/FAs. Such data may be available 
from private sources, in-house testing, or from publicly available 
literature or data bases. For example, emission characterization data 
for baseline gasoline are expected to be available in published 
literature from studies sponsored by the Auto/Oil Program.\42\ The data 
base ``SPECIATE'' might also be useful in identifying baseline 
emissions species for gasoline.\43\ Other applicable literature on 
gasoline and diesel emissions can be obtained in the NRC Report on 
``Feasibility of Assessment of Health Risks from Vapor-Phase Organic 
Chemicals in Gasoline and Diesel Exhaust.''\44\ Emission 
characterization data for alternative fuels is available in a variety 
of CARB reports.45,46
---------------------------------------------------------------------------

    \42\An example reference is ``The Auto/Oil Air Quality 
Improvement Research Program SP-920,'' (published by SAE, Inc., 
February 1992). Similar information may be obtained from other Auto/
Oil publications.
    \43\``SPECIATE--VOC/PM Speciation Data Base Management System,'' 
Version 1.5, EPA-454/C-93-013, October 1992. This data base can be 
obtained electronically from the CHIEF Bulletin Board System (modem 
phone no. 919-541-5742). For information on this data base, call 
919-541-5285 (INFO CHIEF).
    \44\Published by National Academic Press, Washington, DC, 1983 
(see Appendix A of Report).
    \45\``Definition of Low-Emission Motor Vehicle in Compliance 
with the Mandates of Health and Safety Code Section 39037.05,'' 
CARB, May 19, 1989.
    \46\``Proposed Reactivity Adjustment Factors for Transitional 
Low-Emission Vehicles,'' Technical Support Document, CARB, September 
27, 1991.
---------------------------------------------------------------------------

B. Characterization of Emissions

1. Scope
    The chemical analysis requirements of Tier 1 satisfy the provision 
in CAA section 211(b)(2)(B) requiring information ``to determine the 
emissions resulting from the use of the fuel or additive contained in 
such fuel.'' The characterization of emissions in Tier 1 will provide a 
useful inventory of potentially harmful F/FA emission products for 
further study and evaluation in support of the F/FA testing program, 
risk assessments, and future regulatory actions.
    F/FA manufacturers are responsible for the generation, collection, 
and sampling of the combustion and, if applicable, the evaporative 
emissions of their F/FAs, and for the conduct of tests to determine the 
identity and concentration of individual emission products. In general, 
the required procedures are directed toward the detection and 
measurement of selected chemical classes and compounds. The analyses 
include: (1) the measurement of basic emissions (i.e., total 
hydrocarbons, carbon monoxide, oxides of nitrogen, and particulates), 
(2) the speciation of volatile hydrocarbon compounds, aldehydes, 
ketones, alcohols, ethers, and polycyclic aromatic compounds, and (3) 
the speciation of atypical emission products (when atypical elements 
are known to be present in the raw fuel or additive formulation). 
Speciation requirements are summarized in Table 1.

                          Table 1.--Emission Characterization/Measurement Requirements                          
----------------------------------------------------------------------------------------------------------------
                                                                    Speciated Emissions                         
                                           ---------------------------------------------------------------------
       Emission Type              Basic                                                Polycyclic               
                               Emissionsa   Hydrocarbons   Ketones and  Alcohols and    Aromatic     Atypicalsd 
                                                           Aldehydes       Ethersb     Compoundsc               
----------------------------------------------------------------------------------------------------------------
Combustion emissions:                                                                                           
  Vapor phase...............            X             X             X             X   ............            X 
  Semivolatile phase........  ............  ............  ............  ............            X             X 
  Particulate phase.........            X   ............  ............  ............            X             X 
Evaporative emissions:e                                                                                         
  Evaporative emission                                                                                          
   generatorf...............           Xg             X   ............            X   ............           X  
----------------------------------------------------------------------------------------------------------------
aBasic emissions=total hydrocarbons, carbon monoxide, oxides of nitrogen, and particulates (see Section         
  VII.B.2.a below).                                                                                             
bRequired if alcohols or ethers exist in the uncombusted fuel or additive/base fuel mixture.                    
cIncludes specific polycyclic aromatic hydrocarbons (PAHs), nitrated polycyclic aromatic hydrocarbons (NPAHs),  
  and poly-chlorinated dibenzodioxins/dibenzofurans (PCDD/PCDFs). PAH and NPAH speciation is not required for F/
  FAs in the methane (CNG, LNG) and propane (LPG) families, or for F/FAs in the atypical categories of other    
  fuel families. Chlorine-containing atypical F/FAs are subject to the dioxins/furans speciation requirements.  
dManufacturers of atypical products must examine all emission fractions for the measurement and identification  
  of potential atypical species.                                                                                
eOnly applicable to F/FAs required to measure evaporative emissions.                                            
fEvaporative emissions are to be generated using an evaporative emission generator as described in Section VI.C.
                                                                                                                
gThe only basic emission required to be measured for evaporative emissions is total hydrocarbons.               


2. Speciation Procedures
    Section VI describes the required procedures for the generation of 
both combustion and evaporative emissions. Characterization of 
combustion emissions must be done both for non-catalyzed emissions and 
for tailpipe emissions. As discussed in Section III.A.2, the 
evaporative emissions of some F/FAs are also required to undergo 
emission characterization analysis. To provide an indication of the 
variability, the emissions must be generated and characterized three 
times on three different days. Collection and speciation of background 
samples is required.
    The CAA authorizes EPA to require information to characterize F/FA 
emissions, while giving EPA discretion to specify the particular 
protocols to be used for this purpose. The following sections identify 
the general emission product categories of interest and discuss 
currently available protocols which are suitable for their analyses. 
EPA recognizes that scientific methods can be expected to advance in 
the future. Thus, the use of the protocols referenced in this final 
rule is not mandated. Rather, EPA will hold F/FA manufacturers 
accountable for state-of-the-art methods and good analytical chemistry 
and laboratory practices, such as those described in the article 
``Principles of Environmental Analysis.''\47\
---------------------------------------------------------------------------

    \47\Keith et al., ACS Committee on Environmental Improvement, 
``Principles of Environmental Analysis,'' The Journal of Analytical 
Chemistry, Volume 55, pp. 2210-2218, 1983; available in Docket A-90-
07, Item No. II-J-12.
---------------------------------------------------------------------------

    Today's rule does not discourage the use of any validated method to 
perform the characterization of emissions, or the submittal of existing 
speciation results obtained from validated methods, as long as the data 
address the speciation requirements of the F/FA registration program. 
EPA acknowledges the state-of-the-art methods of the Auto/Oil Air 
Quality Improvement Research Program (Auto/Oil Program) for the 
characterization of emissions. In fact, the speciation requirements 
included in this rule for fuels composed primarily of hydrocarbon 
compounds of twelve carbons (C12) or less (e.g., gasoline) are based on 
such methodology.48, 49, 50 Where applicable, EPA will accept 
results from the Auto/Oil Program as adequate data in lieu of new 
testing. However, the Auto/Oil Program might not address all the 
emission characterization requirements of today's rule, so additional 
procedures (e.g., for the analysis of polycyclic aromatic compounds) 
might be needed. EPA recognizes that characterization data have already 
been submitted to EPA in relation to the Auto/Oil Program. F/FA 
manufacturers need not resubmit this information, but are required to 
reference these data (e.g., report number, applicable page numbers, 
etc.) on the Tier 1 report so EPA can verify the adequacy of the 
information being used in compliance with the F/FA registration program 
for the particular
F/FA product or group representative. Although resubmission of the raw 
emission data is not required, manufacturers are still responsible for 
providing a summary discussion of the emission characterization results 
in the Tier 1 report as outlined in Section XII.B.
---------------------------------------------------------------------------

    \48\Jensen, T. E. et al., ``Advanced Emission Speciation 
Methodologies for the Auto/Oil Air Quality Improvement Program--I. 
Hydrocarbons and Ethers,'' SAE 920320 In: Auto Oil Air Quality 
Improvement Research Program, SP-920, February 1992.
    \49\Swarin, S. J. et al., ``Advanced Speciation Methodologies 
for the Auto/Oil Air Quality Improvement Research Program--II. 
Aldehydes, Ketones, and Alcohols,'' SAE 920321, In: Auto Oil Air 
Quality Improvement Research Program, SP-920, February 1992.
    \50\Siegl, W. O. et al., ``Improved Emission Speciation 
Methodology for Phase II of the Auto/Oil Air Quality Improvement 
Research Program--Hydrocarbons and Oxygenates,'' SAE 930142, 1993.
---------------------------------------------------------------------------

    For the characterization of diesel
F/FAs, EPA recognizes the procedures under the Air Pollution Research 
Advisory Council (APRAC) program. The work done by APRAC provides 
speciation guidelines for unregulated diesel emissions and addresses 
diesel combustion compounds of concern to EPA. As with the Auto/Oil 
Program studies, existing applicable APRAC speciation studies will be 
considered adequate data in lieu of new testing. However, today's rule 
requires manufacturers of diesel F/FAs to perform speciation procedures 
for hydrocarbons which might not be included in the APRAC program. 
These are discussed in the following sections.
    a. Characterization of Basic Emissions. EPA proposed to require the 
characterization of ``regulated emissions'' for fuel/vehicle types for 
which certification procedures existed at the time of the publication 
of the NPRM (i.e., gasoline, diesel, and methanol). Today, EPA is 
terming this requirement ``basic emissions,'' instead of ``regulated 
emissions,'' because certification requirements are not established yet 
for all the F/FAs included in this rule. To be consistent and avoid 
confusion, the term ``basic emissions'' is used for all F/FA families 
included in this rule. Based on the current regulated emissions and 
taking into consideration the objectives of this program, EPA selected 
four basic emissions for measurement, as follows: Total hydrocarbons, 
carbon monoxide, oxides of nitrogen, and particulates.
    The four basic emissions are to be measured in combustion emissions 
as a routine requirement for all F/FA families, as shown in Table 1. 
Only total hydrocarbons are required to be measured in evaporative 
emissions. Manufacturers are referred to the vehicle certification 
procedures in 40 CFR part 86 for general guidance on the measurement of 
the basic emissions of interest to this rule.
    b. Characterization of Hydrocarbons. As shown in Table 1, this rule 
requires the speciation of hydrocarbons for the vapor phase of 
combustion emissions and for evaporative emissions generated using an 
evaporative emission generator. The speciation is to be performed using 
methods that identify and determine the concentration of all 
hydrocarbon compounds containing twelve or fewer carbon atoms. The 
Auto/Oil Program procedures referenced above provide an acceptable 
speciation method for hydrocarbons.
    c. Characterization of Aldehydes and Ketones. Speciation of 
aldehydes and ketones containing a maximum of eight carbon atoms is 
required only for the vapor phase of combustion emissions. A test 
procedure for formaldehyde measurement is included in 40 CFR part 86 
for formaldehyde. F/FA manufacturers are also referred to the Auto/Oil 
Program procedures referenced above for the analysis of aldehydes and 
ketones. Additional applicable procedures are available in ASTM D 5197-
91, ``Standard Test Method for Determination of Formaldehyde and Other 
Carbonyl Compounds in Air (Active Sampler Methodology).''
    d. Characterization of Alcohols and Ethers. Alcohol and ether 
compounds containing six or fewer carbon atoms are to be characterized 
for both evaporative and combustion emissions, whenever the fuel or 
additive/base fuel mixture under evaluation contains alcohols or 
ethers. If a F/FA formulation contains an alcohol or ether with more 
than six carbon atoms, then this manufacturer is required to measure 
their presence in the emissions, as well as alcohols or ethers with 
fewer number of carbon atoms. For example, if an ether containing seven 
carbon atoms (e.g., isopropyl tertiary butyl ether) is part of a fuel 
formulation being tested in this program, then its manufacturer must 
characterize ethers with seven or fewer carbon atoms.
    In addition to the Auto/Oil Program procedures referenced above, a 
test procedure for the characterization of alcohols and ethers is 
described in 40 CFR part 80, Appendix F, entitled ``Test Method for 
Determination of C1-C4 Alcohols and MTBE in Gasoline by Gas 
Chromatography''. This procedure can be used for the identification of 
ethers in addition to MTBE, but will require appropriate modifications 
for application to gas phase samples.
    e. Characterization of Polycyclic Aromatic Compounds. In the NPRM, 
EPA proposed a broad requirement for the identification and measurement 
of polycyclic aromatic compounds. In the final rule, this requirement 
is narrowed to a limited number of specified compounds which are of 
significant concern in terms of their potential non-carcinogenic and/or 
carcinogenic effects. Included are specified polycyclic aromatic 
hydrocarbon (PAH) and nitrated polycyclic aromatic hydrocarbon (NPAH) 
compounds as well as individual compounds and classes of 
polychlorinated dibenzodioxins/dibenzofurans (PCDD/PCDFs).
    In addition to specifying particular polycyclic aromatic compounds 
for analysis, the final rule reduces the families and/or categories of 
F/FAs which are subject to these requirements. PAH and NPAH speciation 
need not be done for F/FAs in the methane and propane fuel families, 
nor for F/FAs in the atypical categories of other fuel families. 
Furthermore, speciation of dioxins/furans is required only for
F/FAs which contain chlorine as an atypical element. This is consistent 
with the requirement applicable to atypical F/FAs in general, that all 
emission species containing the relevant atypical elements be 
identified and measured (see section f, below).
    While EPA believes that characterization of dioxins/furans is also 
important in the case of baseline and non-baseline
F/FAs, the NPRM did not propose to require this procedure on a wider 
basis. EPA has thus refrained from including mandatory requirements for 
speciation of dioxins/furans in the case of baseline and non-baseline
F/FAs. Instead, for manufacturers of
F/FAs other than chlorine-containing atypical F/FAs, dioxin/furan 
characterization is included in the final rule only on a voluntary 
basis. EPA strongly encourages manufacturers of baseline and non-
baseline F/FAs to collect the necessary emission samples and conduct 
these voluntary procedures at the same time that mandatory emission 
characterization requirements are being fulfilled. The recent attention 
and concern about the potential health effects of dioxins/furans, 
combined with the current dearth of information on the specific sources 
and generation of these compounds, increases the likelihood that these 
procedures will be prescribed under EPA's discretionary Tier 3 
authority if the necessary data are not otherwise submitted on a 
voluntary basis. If so, the incremental costs are likely to be 
considerably higher than if the procedures were conducted in 
conjunction with the standard Tier 1 emission characterization tasks.
    As was proposed, the final rule requires the measurement and 
speciation of polycyclic aromatic compounds in both the semi-volatile 
phase and particulate phase of combustion emissions. While, in the 
past, these compounds have been analyzed primarily in the particulate 
phase, the quantity of these compounds in the semi-volatile phase at 
the temperatures encountered in dilute exhaust may also be important.
    Particulate and semi-volatile phase emissions are to be collected 
using methods described in Section VI.B.1. The soluble organic fraction 
(SOF) is to be extracted from the filter and polymer bed separately. 
The extracts of the two phases are to be tested separately for PAHs and 
NPAHs, but may be combined before testing for dioxins/furans. Examples 
of protocols suitable for characterizing polycyclic aromatic compounds 
are available in the literature.51, 52, 53, 54, 55, 56
---------------------------------------------------------------------------

    \51\Coordinating Research Council, ``Chemical Methods for the 
Measurement of Unregulated Diesel Emissions,'' CRC Report No. 551, 
1987; available in Docket A-90-07, Item No. II-J-15.
    \52\Tejada, S.B., ``Fluorescence Detection and Identification of 
Nitro Derivatives of Polynuclear Aromatic Hydrocarbons by On-Column 
Catalytic Reduction to Aromatic Amines,'' Analytical Chemistry, 
Volume 58, Number 8, pp. 1827-1834, July 1986.
    \53\Tejada, S.B. et al., ``Analysis of Nitroaromatics in Diesel 
and Gasoline Car Emissions,'' SAE Paper No. 820775, 1982.
    \54\Schuetzle D., ``Analysis of Nitrated Polycyclic Aromatic 
Hydrocarbons in Diesel Particulates,'' Analytical Chemistry, Volume 
54, pp. 265-271, 1982.
    \55\John J. H. et al., ``A review of diesel particulate control 
technology and emissions effects--1992 Horning Memorial Award 
Lecture,'' SAE Technical Paper Series No. 940233, 1994.
    \56\A protocol for identification and measurement of poly-
chlorinated dibenzodioxins and dibensofurans is provided in 40 CFR 
part 60, Appendix A, Method 23.
---------------------------------------------------------------------------

    f. Characterization of Emissions with Atypical Elements. F/FAs 
containing chemical elements other than those included in the baseline 
formulations for the respective fuel family are classified as atypical 
formulations (see Section IV.B.2). In addition to the emission 
characterization requirements described above, producers of atypical F/
FAs are required to identify and measure the emission products 
containing the associated atypical element(s). For example, if a 
gasoline additive product contains chlorine, then this manufacturer 
must identify and measure all emission compounds that contain chlorine. 
Due to the nature of atypical products, special procedures for the 
generation of emissions are required (see Section VI.E).
    The presence of atypical species in specific emission fractions 
will be dependent on the nature of the particular atypical element/
compound of concern. In view of this, EPA recommends that manufacturers 
of atypical products examine all emission fractions (i.e., vapor, semi-
volatile, and particulate) for the measurement and identification of 
potential atypical species. Because of the variety of potential 
elements and reaction products involved, all of the necessary chemical/
analytical procedures cannot be specified in this final rule. The 
selection of the particular method(s) for measuring atypical elements 
or compounds is left to the manufacturer. However, the procedures used 
must be state-of-the-art and based on sound analytical chemistry 
principles applicable to the atypical element or compound of concern.
3. Quality Assurance
    While today's rule requires emissions to be generated and 
characterized three times as a way to evaluate the repeatability of the 
test results, additional quality assurance procedures are needed to 
control variability during the characterization of emissions. 
Laboratories conducting emission characterization/speciation analyses 
are required to perform verification testing to examine the 
repeatability and accuracy of test procedures. For this purpose, a 
prepared mixture of chemical compounds, as appropriate for each 
particular procedure, should be subjected to the speciation protocols. 
The use of analytical standards and controls for calibration of 
instruments is also required to assure precision and accuracy of 
results.
    EPA reserves the right in this final rule to audit testing 
facilities involved in the generation and characterization of 
emissions, as well as the health effects testing of F/FAs. Such audits 
will be organized and administered by EPA at its own expense. The audit 
procedures could include a requirement that facilities submit a 
completed questionnaire in which equipment and procedural information 
is described. EPA might make recommendations based on the submitted 
information and/or might follow up with a visit to observe the 
performance of the protocols. The audit could also include EPA 
distribution of ``blind'' samples for analysis at participating 
laboratories (at their expense). The audit will not have the purpose of 
certifying that the laboratory is ``EPA approved''. Rather, it will 
have the purpose of determining the weaknesses of laboratories and the 
acceptability of the laboratory's current performance.

C. Exposure Analysis

    In the NPRM, EPA proposed to require modeling or other analytic 
methods to evaluate potential exposures, expected atmospheric 
reactivity, and environmental partitioning of emission products. 
However, as discussed in Section III.C., this final rule does not 
require modeling analyses to be performed as routine requirements under 
Tier 1. Instead, quantitative modeling efforts will be required on a 
case-by-case basis as needed under Tier 3.
    Nevertheless, EPA believes that exposure data are still critical 
for the assessment of the potential risks associated with the emissions 
of F/FAs in question. For this purpose, today's rule requires 
manufacturers to provide a qualitative discussion of potential 
population exposures based on the production and use of the particular 
fuel or additive (or group of F/FAs) in question. This qualitative 
analysis must consider the actual and/or projected total annual 
production volumes and the market distribution patterns (e.g., percent 
of sales by state or region) of the particular product or group of 
products. Group submissions must assess the cumulative exposure 
resulting from all members of the group. A quantitative analysis is 
encouraged when appropriate data are available, including any existing 
modeling data, to support the exposure analysis. As discussed earlier, 
EPA retains the authority to require from manufacturers more exhaustive 
exposure analysis for particular products of concern under Tier 3 
(including modeling), based on the EPA evaluation of Tier 1 and Tier 2 
results or other available information.

VIII. Tier 2 Requirements

    In the NPRM, EPA proposed short-term (42-day) tests under Tier 2 
for the evaluation of six health effects endpoints: carcinogenicity, 
mutagenicity, teratogenicity, reproductive toxicity, neurotoxicity, and 
pulmonary toxicity. EPA examined the proposed Tier 2 program and found 
that similar requirements among the various proposed tests (in regard 
to animal subjects, exposure scenarios, and general technical 
principles) provided the opportunity to combine several endpoint tests 
within the same exposure protocol. In view of this, EPA has modified 
the Tier 2 testing program to allow for concurrent test performance in 
a more cost-effective manner.
    The revised Tier 2 testing program enhances efficiency and 
feasibility, while providing better health effects information. In 
fact, the design of the Tier 2 testing program makes best use of 
animals (minimum number of animals used), laboratory capacity, and 
financial resources. The basic Tier 2 testing framework of this final 
rule consists of a 90-day subchronic inhalation study to examine 
general systemic and organ toxicity (including pulmonary effects), with 
the addition of ancillary tests that allow the assessment of several 
specific health effect endpoints (carcinogenicity, mutagenicity, 
teratogenicity, reproductive toxicity, and neurotoxicity) within the 
same exposure schedule. A fertility assessment is coordinated with the 
90-day study to examine reproductive and teratogenic effects.
    Brief descriptions of test guidelines for the evaluation of each 
health effect endpoint are provided in the sections below. Most of 
these testing guidelines are modified versions of guidelines previously 
published under TSCA (40 CFR part 798, revised as of July 1, 1992) and/
or the test guidelines which accompanied the NPRM. Detailed protocols 
for the Tier 2 testing program are included in Sec. 79.62-Sec. 79.68 of 
the accompanying regulations. Figure 4 shows a diagram of the suggested 
timing and organization of the Tier 2 studies within the general 90-day 
subchronic exposure schedule.

    BILLING CODE 6560-50-P

TR27JN94.002


    BILLING CODE 6560-50-C

 A. General Methodology

1. Exposure Duration
    As mentioned previously, EPA originally proposed a program that 
included six separate tests for the evaluation of the endpoints of 
concern. Because of cost considerations, a minimum six-week (42-day) 
exposure period was proposed in the NPRM, instead of the traditional 
90-day test. EPA requested comments on the adequacy of the proposed 
exposure period and exposure regimen, and on the possibility of 
extending these tests to 90 days for comparability to historical data. 
Comments were also requested on the possible use of an alternative 
approach for the testing program, the Screening Information Data Set 
(SIDS) protocol developed for use by the Organization for Economic 
Cooperation and Development (OECD). The standard SIDS protocol is 
designed as a single-study screen (45-day) for repeat dose, 
reproductive, and developmental effects.
    Public comments from industry supported the 90-day test over either 
the proposed 42-day test or the SIDS protocol. Commenters expressed 
concern regarding both of these protocols because relatively less 
scientific experience and historical comparison data are available for 
these shorter test scenarios. Upon reconsideration, EPA agrees that the 
90-day protocol is more suitable and more cost effective than either of 
the alternatives proposed in the NPRM for purposes of the F/FA 
registration program. EPA recognizes that the shorter tests might miss 
some adverse health effects that might be identified with the 90-day 
exposure. Apart from the study duration, the SIDS protocol includes 
somewhat less information for each endpoint, in comparison with the 
selected 90-day subchronic inhalation study, with relatively small 
savings in time and cost. For example, the SIDS protocol includes a 
fertility screen (i.e., one-generation reproductive study), but does 
not include an examination of the fetus for teratogenic effects.
    Thus, today's final rule requires F/FA manufacturers to use the 
more standard 90-day subchronic inhalation protocol for the evaluation 
of health effects, with an exposure regimen of at least six hours per 
day, five days per week. EPA judges the exposure regimen of five days 
per week for the 90-day protocol to be the minimum acceptable exposure 
period for the purposes of the Tier 2 evaluation. Also, the 90-day 
protocol provides a broad and efficient testing approach that allows 
the evaluation of several endpoints at the end of the same exposure 
period using the same exposed test animals. As a result, the modified 
Tier 2 program provides a reduction in the number of animals needed to 
perform the evaluation of the endpoints of concern and savings on 
emission generation and testing costs.
2. Animal Model and Laboratory Practices
    In general, the Tier 2 testing program requires the exposure of 
live laboratory animals to whole F/FA emissions. Rodent species are 
required and rats are specifically recommended. Animal facilities must 
be operated in compliance with the ``Guide for the Care and Use of 
Laboratory Animals'' (U.S. DHHS Publication (NIH) 86-23, 1985). To 
ensure the quality and integrity of test results, the performance of 
all studies will be required to conform with good laboratory practice 
(GLP) standards. GLP standards specific to this rule are included in 
Sec. 79.60. As proposed in the NPRM, the GLP standards are based on 
those published in 40 CFR part 792 (revised as of July 1, 1992) for 
conducting tests under TSCA, with modifications to accommodate the 
specific goals of this rule. The GLP standards address facility, 
equipment, organization, quality assurance, and personnel requirements, 
as well as specifications for proper care of laboratory animals, 
handling of test substances, instrumentation issues, conduct of 
studies, record keeping, and reporting of results.
3. Exposure Route and Concentrations
    With the exception of the Salmonella assay, the Tier 2 testing 
program is based on the inhalation exposure of laboratory animals to 
diluted whole emissions. Such studies require an exposure system 
designed to ensure the controlled generation, dilution, and delivery of 
F/FA emissions to the laboratory animals for prolonged periods. Section 
VI describes the methodology for the generation of F/FA combustion and 
evaporative emissions and the procedures to deliver the emissions to 
the test animals. Requirements for hardware, maintenance, and the use 
of emission generation and inhalation systems are included in 
Sec. 79.57 and Sec. 79.61 of this rule.
    Before testing the emissions of a particular fuel or additive/base 
fuel mixture, a manufacturer must determine an appropriate range of 
exposure concentrations to be used in the characterization of potential 
health effects. The objective is to select exposure concentrations to 
determine a reasonable concentration-response curve that may predict 
the potential health risks associated with a particular exposure. 
Concentrations should be spaced to produce test groups with a range of 
toxic effects. In order to accomplish this, EPA requires that at least 
three concentration levels be used to construct the concentration-
response curve. These levels should correspond to, at a minimum: (1) an 
overtly or highly toxic concentration, (2) a concentration having an 
intermediate level of toxicity, and (3) a minimally toxic or non-toxic 
level. The highest concentration should result in toxic effects but not 
produce a level of fatalities which would prevent a meaningful 
interpretation of the resulting data. The lowest concentration should 
produce minimal or no observable toxic effects. If more than one 
intermediate concentration level is used, the concentrations should be 
spaced to produce a gradation of toxic effects. Due to the inherent 
toxicity of most vehicle emissions, it might be impossible to precisely 
select an exposure level which results in no observable evidence of 
toxicity, or a no- observed-adverse-effect-level (NOAEL). Thus, EPA is 
not requiring manufacturers to specifically achieve a NOAEL. Instead, 
EPA recommends that manufacturers use available scientific approaches 
(e.g., range-finding test and extrapolation of data results) to design 
the study with reasonable concentration spacing so as to improve the 
probability of achieving a NOAEL. In recognition of the possibility 
that the highest achievable exposure concentration (considering the 
limiting CO concentration) may also be non-toxic, provisions are made 
for ``limit tests,'' where appropriate. If a test at the highest 
achievable concentration produces no observable toxic effect(s), then a 
full study using three concentration levels might not be necessary (see 
regulatory text for specific endpoint tests).

B. Subchronic Inhalation Study and Endpoint Tests

    As described earlier, the Tier 2 health effects testing program 
includes a 90-day subchronic inhalation study and ancillary assays/
tests for the examination of specific health effects endpoints. 
Specific assays or analyses for carcinogenicity, mutagenicity, 
teratogenicity, reproductive toxicity, and neurotoxicity may be 
conducted at the end of the 90-day study, coordinated with the 90-day 
exposure, or conducted separately.
    In the NPRM, EPA proposed a separate acute pulmonary test (i.e., 
lung lavage assay) for the evaluation of pulmonary toxicity. However, 
the Tier 2 testing framework of this final rule does not require a 
separate test for the assessment of pulmonary effects because the 90-
day subchronic study includes gross pathology and histopathology of the 
lungs and respiratory tract. Pulmonary effects in this final rule will 
be examined as part of the standard 90-day inhalation study. Positive 
results at the end of the exposure period for pulmonary toxicity will 
be indicated by abnormal gross or histopathological findings relative 
to appropriate control animals.
1. Subchronic Inhalation Study
    The subchronic inhalation study is designed to determine a 
concentration-response relationship for potential toxic effects in 
rodents, resulting from exposure to vehicle/engine emissions over a 
period of 90 days. This test will provide valuable information on 
general systemic and target organ toxicity, including pulmonary 
effects. This information is considered an essential component for the 
assessment of potential health hazards resulting from the exposure to 
F/FA emissions. The exposure period of the subchronic study (i.e., 90 
days) covers approximately one tenth of the life span for the 
recommended test animal species (i.e., rats). Although life-shortening 
or tumors are not likely to be observed within the 90 days of exposure, 
the subchronic study should be able to identify a wide variety of 
adverse effects.
    The specific guidelines for the subchronic inhalation study are 
included in Sec. 79.62 of this final rule. The subchronic study 
requires that animals be observed and weighed during the exposure 
period. Ophthalmological examination, blood chemistry analysis, organ 
examination, and histopathology are basic requirements of the 90-day 
protocol. Hematology and clinical biochemistry determinations are 
required to be carried out after 30 days of exposure and just prior to 
termination. Hematology analyses include: hematocrit, hemoglobin 
concentration, erythrocyte count, total and differential leukocyte 
count, and a measure of clotting potential (e.g., clotting time, 
prothrombin time, thromboplastin time, or platelet count). Clinical 
biochemical testing includes assessment of electrolyte balance, 
carbohydrate metabolism, and liver and kidney function. Other specific 
biochemical tests are described in the regulatory text of the final 
rule.
    At the end of the exposure period, tissues and/or organs from a 
subgroup of the test animals are specially preserved according to the 
requirements of the neurological, pulmonary, and reproductive organ 
examinations. Tissues/organs from the main test population are 
preserved using standard techniques for the general toxicity 
evaluation. Test animals will be subjected to a full gross necropsy 
which includes examination of the external surface of the body, all 
orifices, and the cranial, thoracic, and abdominal cavities and their 
contents. All major organs must be weighed. Gross pathology must be 
performed on the following target organs and tissues: liver, kidneys, 
lungs, adrenals, brain, and gonads. Histopathology must be performed on 
all gross lesions and specific organs/tissues, as follows: respiratory 
tract (i.e., lungs, nasopharyngeal tissues, trachea), brain, heart, 
sternum with bone marrow, salivary glands, liver, spleen, kidneys, 
adrenals, pancreas, reproductive organs (i.e., uterus, cervix, ovaries, 
testes, epididymides), aorta, gall bladder, esophagus, stomach, 
intestinal tract, urinary bladder, representative lymph node, and 
peripheral nerve/tissue. Other organs and tissues must be preserved in 
a suitable medium for possible future histopathological examination, as 
described in the regulatory text of the final rule.
    As described earlier, the 90-day subchronic inhalation study will 
serve as a basic framework for the Tier 2 testing program. The 
following sections provide brief descriptions of the ancillary assays 
and other additional test and/or measures performed under Tier 2.
2. Carcinogenicity and Mutagenicity Assays
    For the evaluation of carcinogenicity and mutagenicity, Tier 2 
includes a battery of three genotoxic assays: Salmonella, micronucleus 
(MN) and sister chromatid exchange (SCE). These assays are specific for 
mutagenic/carcinogenic outcomes at a cellular level, but the tests may 
not be indicative of non-mutagenic or initiation/promotion cancer 
mechanisms.
    In general, the Tier 2 genotoxic assays are considered cost-
effective indicators of mutagenicity and, by implication, predictors of 
suspect carcinogens. The rationale for using these tests for the 
assessment of potential mutagenic and carcinogenic effects is based on 
the general assumption that cancer is a multi-stage process involving a 
variety of events that can include genotoxic steps. The general 
consensus among scientists is that many of the cancers may be 
attributable to adverse genetic changes. Because genotoxic steps are 
generally implicated early in the process of cancer development, their 
detection has assumed the status of presumptive carcinogen 
identification. EPA recognizes that this working assumption has some 
limitations because there is always a possibility of having false 
positive (mutagenic noncarcinogens) or false negatives (nonmutagenic 
carcinogens) when evaluating the carcinogenic potential of the test 
substance. However, these limitations do not preclude the usefulness 
and the effectiveness of the genotoxic assays (i.e., Salmonella, MN, 
and SCE) as cost-effective predictors of potential mutagenic and 
carcinogenic effects in the context of the Tier 2 testing program.
    A description of the Tier 2 assays required for the evaluation of 
carcinogenicity and mutagenicity is provided in the following sections. 
The two in vivo assays (MN and SCE) can be conducted concurrently with 
the 90-day subchronic inhalation study (i.e., same animal population). 
The Salmonella assay is run separately, because it does not require the 
exposure of live animals to emissions.
    a. Salmonella Assay. The Salmonella assay is an in vitro test for 
mutagenicity and, by implication, for carcinogenicity. The assay makes 
use of five mutant strains of the bacterium Salmonella typhimurium 
which cannot grow in a medium deficient in histidine due to an 
inherited inability to produce this amino acid. Exposure to mutagenic 
or carcinogenic substances can elicit reverse mutations, such that the 
bacteria regain their ability to grow in a histidine-deficient medium. 
In this test, bacteria will be exposed to the semi-volatile and 
particulate extracts of combustion emissions (see Section VI.B.2 for 
information on sampling of combustion emissions for testing). Test 
procedures for this assay have been described in previous 
literature.\57\ After exposure, the cells will be plated on histidine-
deficient media (both with and without metabolic activation) and 
incubated for a designated period of time. The number of emissions-
induced mutant colonies (revertants) growing on the plates will then be 
compared to the number of spontaneous revertants in control cultures. 
The testing guidelines for the Salmonella assay are included in 
Sec. 79.68 of this final rule.
---------------------------------------------------------------------------

    \57\Huisingh, J.L., et al, ``Mutagenic and Carcinogenic Potency 
of Extracts of Diesel and Related Environmental Emissions: Study 
Design, Sample Generation, Collection, and Preparation,'' In: Health 
Effects of Diesel Engine Emissions, Vol. II, W.E. Pepelko, et al. 
(Eds.), US EPA, Cincinnati, EPA-600/9-80-057b, pp. 788-800, 1980; 
available in Docket A-90-07, Item No. II-J-13.
---------------------------------------------------------------------------

    A positive result for the Salmonella assay occurs when there is a 
statistically significant concentration-related increase in the number 
of revertants or a reproducible and statistically significant positive 
response for at least one of the test concentrations.
    b. In Vivo Micronucleus (MN) Assay. Micronuclei are sub-cellular 
structures containing chromosomes and chromosome fragments not 
incorporated into the main nucleus during cell division. While 
micronuclei do form under natural conditions, exposure to potentially 
mutagenic or carcinogenic agents can cause an increase in 
micronucleated cells. In this assay, live rodents will be exposed by 
inhalation to the emissions of the particular fuel or additive/base 
fuel mixture (this assay is applicable to the evaluation of both 
combustion and evaporative emissions). Subsequently, erythrocytes in 
the bone marrow will be sampled, stained, and viewed under a light 
microscope. The number of erythrocytes containing micronuclei will then 
be counted and compared with erythrocytes from untreated animals. The 
use of erythrocytes in this procedure facilitates the visualization of 
micronuclei, since their primary nucleus is normally extruded during 
cell development. The testing guidelines for the MN assay are contained 
in Sec. 79.62 and Sec. 79.64 of this final rule.
    A positive result for the MN assay is determined by a statistically 
significant concentration-related increase in the number of 
micronucleated erythrocytes or a reproducible and statistically 
significant positive response for at least one of the test 
concentrations.
    c. In Vivo Sister Chromatid Exchange (SCE). SCEs are believed to be 
caused by chromosome strand breakage resulting in exchanges of genetic 
material between the halves of a chromosome ``pair'' (i.e., the 
chromatids). While some SCEs occur normally, an increase in the 
frequency of such exchanges may be indicative of carcinogenic activity. 
In this assay, animals which have undergone inhalation exposure to the 
emissions will be sacrificed (this assay is applicable to the 
evaluation of both combustion and evaporative emissions). Peripheral 
blood lymphocytes will then be isolated and cultured. The cells will be 
treated with a DNA base analog (bromodeoxyuridine, BrdU) and with a 
spindle inhibitor such as colchicine. After appropriate staining for 
labeled DNA, SCEs will be scored from cells arrested in the second 
mitotic division and the results compared with appropriate controls. 
Details on the SCE testing procedures are included in Sec. 79.62 and 
Sec. 79.65 of this final rule.
    A positive result for the SCE assay is determined by a 
statistically significant concentration-related increase in the number 
of SCE or a reproducible and statistically significant positive 
response for at least one of the test concentrations.
3. Fertility Screen for Reproductive and Teratologic Effects
    The fertility screen involves mating of test animals previously 
exposed (by inhalation) to F/FA emissions to examine the effects of 
such exposure on conception. The females will continue their exposures 
throughout pregnancy and will be assessed for teratologic effects on 
their offspring. This test is applicable to the evaluation of both 
combustion and evaporative emissions. The fertility assessment is 
coordinated with the 90-day subchronic inhalation study and replaces 
the separate reproductive and teratology studies originally proposed in 
the NPRM.
    The reproductive assessment includes vaginal cell smears to track 
effects on the estrous cycle. Commenters discouraged the use of 
frequent vaginal smearing. In response to the comments, vaginal 
cyclicity determinations will be performed on the test subjects for two 
weeks prior to the start of the exposure period (for culling acyclic 
females), and will resume after seven weeks of exposure (for four weeks 
or until the female is confirmed pregnant). The assessment for 
reproductive effects also includes a mating trial and the weighing and 
histopathological examination of male and female reproductive organs 
(i.e., uterus, ovaries, testes, epididymis, and seminal vesicles), all 
of which can be performed within the context of the 90-day subchronic 
inhalation study.
    As compared with appropriate control animals, positive results for 
reproductive effects include: changes in the length or stages of the 
estrous cycle as indicated by the vaginal cytology data, changes in 
reproductive organ weights, and pathological changes found during gross 
or microscopic examination of male or female reproductive organs. 
Changes in fecundity, fertility or litter size (number of normal 
fetuses) will also be regarded as positive results.
    To assess potential teratogenic effects, each dam will be 
sacrificed on the day prior to normal parturition and its uterus 
examined for embryonic or fetal deaths. Viable fetuses will be counted 
and then examined for skeletal and soft tissue anomalies. These results 
will be evaluated relative to the number of spontaneous embryonic or 
fetal deaths and abnormalities in unexposed controls. The required 
analysis covers specific fetal effects outcomes, i.e., in utero death, 
growth alteration, and structural abnormalities.
    Although the NPRM proposed the use of two different species for the 
evaluation of teratogenic effects, today's action requires the use of 
only one species to satisfy the Tier 2 requirements. This modification 
will reduce the number of animals needed for Tier 2 tests, as requested 
by some of the commenters, while maintaining an adequate approach for 
the screening of teratology effects.
    The above described combined protocol will detect fertility 
problems and teratogenic effects. EPA realizes that the protocol will 
not detect adverse effects on reproductive development that might 
appear in the offspring as they grow and mature (since pups are 
examined just before birth). However, EPA judges that the combined 
protocol provides a reasonable screen for both reproductive and 
teratogenic effects. The testing guidelines for the fertility 
assessment are included in Sec. 79.62 and Sec. 79.63 of this final 
rule.
4. Neurotoxicity Screen
    EPA proposed in the NPRM to require the Functional Observational 
Neurotoxicity Battery (FONB) for the evaluation of neurotoxic effects. 
In addition to the FONB, EPA asked for comment on the possible 
inclusion of a biochemical assay to measure the level of glial 
fibrillary acidic protein (GFAP). Comments were received on both 
proposed tests.
    The regulated industry objected to the use of the FONB on the basis 
of their belief that it would be likely to give artifactual test 
results. Commenters also expressed concern about the use of the GFAP 
assay. They considered it to be a sensitive marker for neurotoxicity, 
but discouraged its use because they perceived that adequate historical 
data are not available for this test. While not necessarily agreeing 
with either of these comments, EPA has reconsidered its original 
proposal regarding the specific content of the Tier 2 neurotoxicity 
assessment.
    EPA considers the FONB to be a well-validated standard test and 
recognizes its extensive use in the past. In the context of this F/FA 
emissions testing program, however, EPA believes that, rather than 
requiring the FONB on a standard basis within the Tier 2 testing 
regimen, the FONB is more appropriately reserved for use when a 
neurotoxicity concern has been identified and additional testing is 
needed to clarify the nature and/or significance of the potential 
adverse effects (e.g., within Tier 3).
    In regard to the GFAP assay, EPA judges that concerns about the 
amount of historical data are not sufficient justification to prevent 
its use in the F/FA program, given its high specificity and potential 
applicability as a screening test. This final rule thus includes the 
GFAP assay as an element in the Tier 2 assessment of potential 
neurotoxic effects.
    The GFAP assay is a biochemical assay that measures the level of a 
major intermediate filament protein of astrocytes (cells of the 
supporting structure of the nervous system) from brain tissues. An 
increase in the GFAP level is highly specific for detecting the 
existence and location of chemical-induced injury to the central 
nervous system (CNS) associated with astrocytic hypertrophy. However, 
due to its specificity, the assay does not provide information on other 
potential sites for neurotoxic effects. To provide a more comprehensive 
evaluation, today's rule supplements the GFAP assay with an expanded 
neurohistopathology examination as part of the 90-day subchronic 
inhalation study. The neurohistopathology involves the examination of 
several brain sections, including the cerebrum, cerebellum, medulla, 
cervical bulb of the spinal cord, and peripheral nerves (e.g., tibial 
or sciatic nerve). Testing guidelines for the neurotoxicity evaluation 
are included in Sec. 79.62, Sec. 79.66 (neuropathology assessment), and 
Sec. 79.67 (GFAP) of this final rule. Positive results at the end of 
the exposure period for neurotoxicity will be indicated by an increase 
in the GFAP level and/or abnormal gross or histopathological findings 
relative to appropriate control animals. The neurotoxicity screen is 
applicable to the evaluation of both combustion and evaporative 
emissions.

C. Adequate Endpoint Information in Lieu of Tier 2 Tests

    One of the functions of the data search requirement under Tier 1 
(as discussed in Section VII.A.2) is to enable F/FA manufacturers to 
examine the available literature and determine if adequate data exists 
(for both combustion and evaporative emissions, as applicable) that 
would satisfy the Tier 2 testing requirements, so that duplication of 
effort can be avoided. In addition to existing test data from protocols 
similar to those specified in the Tier 2 testing program, EPA will 
consider results from other test protocols to be adequate in lieu of 
new testing, as long as the alternative methods provide comparable 
information. Table 2 provides criteria for determining what constitutes 
adequate existing data in lieu of the specified Tier 2 tests, and 
includes an example list of comparable tests for each Tier 2 endpoint. 
EPA recognizes that changes and scientific advances in toxicology 
testing may result in the development of additional techniques and 
methods that could be applicable to the Tier 2 testing requirements of 
this program in the future. In deciding if a specific protocol is 
acceptable in lieu of a Tier 2 test, manufacturers must also address 
other specific criteria for Tier 2 requirements, as explained below.

           Table 2.--Criteria for Determining Adequacy of Existing Data in Lieu of Tier 2 (T2) Tests            
----------------------------------------------------------------------------------------------------------------
                                                   Minimum requirements for                                     
                       T2 testing in the absence     existing data to be      Examples of other existing studies
      Endpoint         of adequate existing data     considered adequate           which may be acceptable      
                                                  substitutes for T2 tests                                      
----------------------------------------------------------------------------------------------------------------
Carcinogenicity......  Salmonella assay, in vivo  Salmonella assay plus two  Alternative to all assays: Lifetime
                        Micronucleus assay, and    other assays (at least     cancer study.                     
                        in vivo Sister Chromatid   one of which shall be in  In vivo assays: Sister Chromatid   
                        Exchange assay.            vivo).                     Exchange (SCE), Chromosomal       
                                                                              Aberrations (CA), Micronucleus    
                                                                              (MN), Unscheduled DNA Synthesis   
                                                                              (UDS).                            
                                                                             In vitro or microbial assays: E.   
                                                                              coli Reverse Mutation, DNA Repair,
                                                                              Yeast Mutation, Yeast Mitotic     
                                                                              Recombination, Mouse Lymphoma, CHO/
                                                                              V79 Mutation, UDS, CA, SCE, Cell  
                                                                              Transformation.                   
Mutagenicity.........  Salmonella assay and in    Salmonella assay plus one  MN, Dominant Lethal Assay, CA,     
                        vivo Micronucleus assay.   in vivo assay.             Heritable Translocation Assay,    
                                                                              Specific Locus Assay.             
Teratogenicity.......  T2 fertility/teratology    FDA/Phase II (gd6-15)      If fetal effects analysis is       
                        assessment with 90-day     Study.                     included: Two-generation study,   
                        exposure.                                             Reproductive Assessment by        
                                                                              Continuous Breeding (RACB), One-  
                                                                              generation study.                 
Adult reproductive     T2 fertility/teratology    T2 fertility/teratology    Two-generation study, Reproductive 
 effects.               assessment with 90-day     assessment with 90-day     Assessment by Continuous Breeding 
                        exposure.                  exposure.                  (RACB), One-generation study.     
Neurotoxicity........  GFAP assay and neuro-      GFAP assay and neuro-      Detailed characterization of       
                        histopathology with 90-    histopathology with 90-    neurotoxicity using behavior,     
                        day exposure.              day exposure.              neurophysiological, and/or        
                                                                              neurochemical assessments (e.g.,  
                                                                              EPA Neurotoxicity Assessment).    
Pulmonary effects....  T2 respiratory tract       T2 respiratory tract       Chronic toxicity study, with or    
                        pathology after 90-day     pathology after 90-day     without lifetime cancer study;    
                        exposure.                  exposure.                  subchronic toxicity study.        
----------------------------------------------------------------------------------------------------------------

    In general, for existing information to fulfill the Tier 2 testing 
requirements, it must include the in vivo inhalation exposure to whole 
motor vehicle emissions (combustion or evaporative, as applicable), 
except for the few in vitro studies acceptable for carcinogenicity/
mutagenicity assessment. In order to be acceptable, previous toxicity 
studies must include exposure to non-catalyzed emissions, as required 
in this final rule. EPA is requiring the use of an evaporative emission 
generator for the evaporative emission testing. However, EPA will 
accept previous inhalation exposure studies of whole evaporative 
emissions in which the emissions were generated using standard 
evaporative emission procedures under FTP conditions. Raw product 
tests, using F/FAs in the uncombusted state, are not considered 
adequate replacements for Tier 2 combustion emission testing. In 
addition, studies using whole aerosolized preparations or tests on 
individual emission products of the fuel or additive cannot be used as 
substitutes for whole emission testing for either combustion or 
evaporative emissions. The reason for this requirement is that, as 
explained in Section III.A.2 of this preamble, this rule focuses on the 
effects of whole emissions rather than raw F/FAs or individual emission 
products. Tests performed on the emissions of F/FAs which are 
classified in the same group as the subject fuel or additive are 
considered relevant, but tests on products not conforming to the 
grouping criteria of the subject fuel or additive do not apply.
    Other important parameters to consider in determining if existing 
studies are adequate include: the type and number of test subjects, the 
number and adequacy of dosages, the methodology and duration of 
exposure, and the technical methods used for monitoring the progress of 
the test and for analyzing the results. Generally, 90-days is the 
minimum acceptable length of exposure. However, an existing study 
having a shorter exposure period might be considered adequate if the 
test results are positive, i.e., adverse effects are observed. 
Previously-conducted studies using mammals other than rodents may be 
acceptable in lieu of rodent testing, if the existing studies meet all 
other applicable criteria for adequacy.
    For carcinogenicity and mutagenicity, the Salmonella assay is 
always required because of its broad sensitivity and specificity for 
detecting chemical exposures having mutagenic and/or carcinogenic 
potential. Manufacturers should note (see Table 2) that when a 
registrant needs to conduct new Tier 2 testing, two in vivo assays (MN 
and SCE), in addition to the Salmonella assay, are required to satisfy 
the carcinogenicity endpoint. However, in lieu of new testing, a 
manufacturer may rely on one existing in vivo assay for 
carcinogenicity, in addition to the Salmonella assay and another in 
vitro assay. In other words, two in vivo tests for carcinogenicity are 
required if new testing is performed, but only one in vivo study is 
required if the manufacturer relies on existing carcinogenicity 
information. The reason for this is that the incremental costs of 
performing the two specified in vivo assays (MN and SCE) within the 90-
day inhalation protocol (as required in this final rule) is low in 
comparison with the amount of useful data obtained. This approach is 
cost-effective because the same animals used in the inhalation study 
can also be used for the in vivo carcinogenicity assays. In fact, fewer 
resources will be spent to do both the MN and SCE assays within a 
single group of inhalation-exposed animals than to conduct just one of 
these tests along with a separate in vitro carcinogenicity assay (in 
addition to the Salmonella assay).

D. Alternative Tier 2 Provision

    The Tier 2 tests described above pertain to all designated F/FAs, 
unless mitigated by special provisions or comparable data from 
adequately performed and documented previous studies. In general, EPA 
considers this standard testing program to be necessary for the health 
effects evaluation of F/FA emissions, even if further evaluation may be 
required under the provisions of Tier 3. However, this final rule also 
adopts the special provision discussed in the Reopening Notice, under 
which EPA retains the authority to modify the standard Tier 2 test 
requirements in certain instances.
    The exercise of this authority will be done wholly at EPA's 
initiative and discretion. The alternative Tier 2 provision is intended 
to provide a degree of flexibility to EPA when available information 
indicates that, in a specific case, another testing regimen is 
preferable to the standard set of Tier 2 tests. There are three 
scenarios under which EPA generally anticipates the possible use of 
this provision.
    First, for a particular fuel or fuel additive (or group), 
information may be available (independent of the requirements of this 
rule) which may cause EPA to be concerned about potential health 
effects related to an endpoint not specifically addressed in Tier 2. In 
such an instance, the alternative Tier 2 provision allows EPA to 
require additional studies targeted to the identified area of concern, 
even though these studies are not normally included in Tier 2. While 
the standard structure of this rule also allows EPA to prescribe the 
additional tests under Tier 3, the alternative Tier 2 provision enables 
EPA to prescribe and receive the desired data earlier in the process. 
This flexibility is particularly important given that this final rule 
allows up to six years for Tier 2 submittal. When the additional 
testing can be coordinated with the standard Tier 2 testing program, 
the alternative Tier 2 provision will also save costs relative to 
conducting the additional tests at a separate point in time.
    Second, independent of the information to be submitted under this 
rule for a particular F/FA product (or group), EPA may identify a 
potentially significant public health risk related to a Tier 2 
endpoint, such that EPA knows that more definitive testing will be 
required for this endpoint than is ordinarily required under Tier 2. 
Again, EPA could require such testing under Tier 3 after the evaluation 
of Tier 2. However, the alternative Tier 2 provision can facilitate 
earlier and potentially more efficient acquisition of the required 
data. If appropriate to the case at hand, EPA would substitute the more 
definitive endpoint test for the standard Tier 2 test (with appropriate 
deadline adjustment). In such a case, EPA's authority to waive the 
requirement to provide the respective Tier 2 test derives from its 
authority in section 211(e) to provide exemptions from testing when 
such testing would be duplicative. In this example, because the 
substituted test would address the endpoint more rigorously than the 
standard Tier 2 assessment, it would be duplicative to require both 
evaluations.
    Third, EPA may identify concerns about the effects of F/FA 
emissions involving different engine and/or emission control 
technologies than those ordinarily required for generating the 
emissions tested in Tier 2. For example, biological testing using 
catalyzed instead of non-catalyzed emissions might be required if 
emission species of concern are present in the catalyzed exhaust of a 
fuel or additive that are not represented in the untreated exhaust.\58\ 
In this case, EPA could prescribe a Tier 2 program using catalyzed 
instead of non-catalyzed emissions under the alternative Tier 2 
provision. Otherwise, the manufacturer of this product would likely be 
required to conduct a second series of biological tests with catalyzed 
emissions, under Tier 3.
---------------------------------------------------------------------------

    \58\As described in Section VI.A, the use of non-catalyzed 
emissions are normally required in the Tier 2 toxicology tests. 
Under Tier 1, however both catalyzed and non-catalyzed emissions are 
required to be characterized.
---------------------------------------------------------------------------

    In summary, the alternative Tier 2 provision will give EPA the 
flexibility, when indicated, to prescribe additional tests to be 
performed along with the standard Tier 2 program, to substitute 
different tests, and/or to modify the underlying vehicle/engine 
specifications for Tier 2. When EPA exercises its authority under this 
special provision, it will allow an appropriate time for completion of 
the prescribed alternative tests. EPA may also use the alternative Tier 
2 authority to waive certain Tier 2 endpoint evaluations (generally on 
occasions when additional and/or more rigorous tests are being required 
for other Tier 2 endpoints). However, Tier 2 endpoint tests will not be 
waived in the absence of adequate information or requirements for more 
rigorous testing of the endpoint(s).
    EPA intends to exercise this special authority only in exceptional 
cases. When EPA decides to use the alternative Tier 2 provision, EPA 
will notify the responsible manufacturer (or group) by certified mail 
letter of the specific modifications in lieu of the standard Tier 2 
program, along with a schedule for compliance and submittal of test 
results. The manufacturer (or group of manufacturers) will have 60 days 
to comment on the prescribed alternative Tier 2 testing program and 
timing requirements. If the responsible manufacturer does not provide 
any comments, EPA will assume that the manufacturer has consented in 
full with the prescribed testing regimen. EPA will publish a notice in 
the Federal Register to inform the public of its intent to require 
alternative testing for a particular F/FA manufacturer and that a copy 
of the letter to the manufacturer is available in the public record of 
this rule for review and comment. Additional correspondence between EPA 
and the responsible manufacturer regarding alternative testing 
requirements will also be placed in the public record. After receipt 
and review of all comments received (or, if no comments are received), 
EPA will publish a notice of final action on the proposed alternative 
Tier 2 requirements in the Federal Register.
    In the Reopening Notice, EPA proposed to notify manufacturers of 
proposed alternative Tier 2 requirements within 18 months of 
promulgation of the final rule (for registered F/FAs) or within 18 
months of EPA's receipt of intent to register (for currently 
unregistered F/FAs). Comments received from the industry indicated that 
this notification period was too long relative to the proposed three 
year deadline for submittal of Tier 2 results. However, EPA believes 
the proposed notification period is reasonable, given that this final 
rule allows manufacturers up to six years for submittal of Tier 2 
results. Furthermore, EPA believes that restricting the time period for 
exercising the alternative Tier 2 authority is unnecessary and 
disadvantageous to the public interest as well as the regulated 
industry, since this provision establishes a mechanism to provide 
needed data on a timely basis and to eliminate unnecessary screening 
studies when substitute tests will be required. Therefore, while EPA 
will endeavor to notify manufacturers of proposed alternative Tier 2 
requirements within 18 months, this final rule permits EPA to notify 
manufacturers of proposed alternative Tier 2 requirements at any time 
prior to EPA's receipt of Tier 2 data. If a manufacturer receiving such 
notification has already begun the standard Tier 2 toxicology testing, 
then EPA will refrain from requiring the testing; however, in such 
cases, the manufacturer is required to submit the results of the 
standard Tier 2 tests within one year of the date when testing began. 
In other cases, EPA will consider the potential costs, burdens, and 
timing factors in making its final decisions on alternative Tier 2 
requirements.

IX. Tier 3 Requirements

A. Scope

    On the basis of the submitted Tier 1 and/or Tier 2 data, or any 
other available information, EPA will determine whether further testing 
and/or analysis for the subject fuel or fuel additive is needed under 
the provisions of Tier 3. Given the variety of evaluations included in 
Tiers 1 and 2 and the wide range of possible interrelated outcomes 
which could be obtained, EPA proposed to use its discretion in 
determining the need for Tier 3 testing on a case-by-case basis. 
Decisions on the need for follow-up testing within Tier 3 would depend 
on expert scientific judgment as to the availability of adequate data 
to enable a health risk evaluation and the need for more definitive 
information for developing regulatory decisions.
    EPA requested comments on the proposed discretionary nature of Tier 
3 determinations and on a possible alternative approach involving the 
establishment of ``automatic triggers'' for Tier 3 decisions, i.e., 
specific outcomes of Tiers 1 and 2 which would make Tier 3 testing 
mandatory. Responses included both support and opposition to the 
proposed Tier 3 discretionary approach. EPA evaluated all comments and 
determined that decisions on Tier 3 requirements should remain at the 
discretion of EPA. In order to accomplish the goals of the program, it 
is essential for EPA to be able to examine the Tier 1 and Tier 2 data 
prior to prescribing additional tests. The need for and content of Tier 
3 testing will most often be dependent on the results of the earlier 
tiers. In these cases, decisions on Tier 3 can only be reasonably 
specified after EPA's review of the applicant's initial submittal. The 
purpose of Tier 3 is not to fill all data gaps, but to establish a 
program that provides the Administrator with the necessary and 
reasonable information to make regulatory decisions. Based on this 
rationale, EPA judges that it is neither practical nor desirable to 
specify criteria which will automatically force F/FA manufacturers to 
perform additional testing under Tier 3. Thus, EPA will use its 
discretion to determine Tier 3 requirements on a case-by-case basis, 
allowing EPA to target specific regulatory needs. Although EPA can use 
the Tier 3 data to support regulatory actions, referral to Tier 3 level 
is not mandatory before beginning actions under CAA section 211(c).
    The need for Tier 3 testing and/or analysis will depend, in part, 
on whether Tier 1 and/or Tier 2 data provide sufficient toxicity and 
exposure information to determine the potential health risks associated 
with a particular fuel or fuel additive. The endpoints to be addressed 
and the nature of the studies to be performed under Tier 3 are to be 
determined on a case-by-case basis. Tier 3 studies will most often be 
required to further explicate the results of the tests/analyses 
performed under Tiers 1 and 2 or to address other areas of concern 
highlighted by the literature search. If additional toxicity testing is 
required under Tier 3, the test might entail whole emissions (as in 
Tier 2), or the testing of one or more individual emission species 
identified to be of particular concern. EPA could also use other 
information (available outside this program) to require testing under 
Tier 3.
    While the specific objectives and scope of Tier 3 testing will vary 
depending on the concerns identified in the earlier tiers or any other 
information available to EPA, examples of possible areas for further 
testing are: chronic/lifetime studies, chemical disposition/metabolism 
studies, exposure studies, dosimetry analyses, additional emission 
characterization/speciation, additional modeling analysis, 
environmental toxicity tests, testing using different emission 
generation procedures or emission control systems, or any other 
additional evaluation approach EPA deems necessary to assess the health 
and/or welfare effects of a particular fuel or fuel additive. The 
previous examples and the discussion below in no way limit the scope of 
Tier 3 or EPA's authority to require further testing under this 
program.
    Today's rule specifically includes under Tier 3 any health effects 
testing to be performed on aerosol additives (see Section X.C for 
details on the special provision for aerosol products). Due to the 
special nature and use of this type of product, EPA judges that any 
testing needs will require non-standard test procedures that can be 
better addressed under Tier 3. EPA will review the composition 
information and literature data on the specific aerosol product and 
will determine if health effects testing is needed on a case-by-case 
basis. For example, if available literature, submitted under Tier 1, 
indicates that a component of an aerosol product is highly toxic, and 
the product is widely produced, then Tier 3 testing on the aerosol 
product would likely be indicated.
    Modeling of potential exposures, atmospheric reactivity, and/or 
environmental fate/partitioning may also be required under Tier 3, as 
explained in Section III.C. For example, if a fuel or additive is 
widely distributed and Tier 2 data indicate serious concern for health 
effects, EPA might require more extensive exposure analysis and/or 
modeling under Tier 3 to better define potential risks.
    Although the grouping mechanism included in this rule allows F/FA 
manufacturers to submit Tier 1 and Tier 2 data on the representative of 
a designated group for purposes of registration, EPA retains the 
authority under Tier 3 to require testing on any member product of a 
group. Thus, when follow-up testing is required under Tier 3 authority, 
the specified test(s) could be required to be conducted on the selected 
representative or on any other member of an existing group. Testing on 
additional F/FA products could be required if EPA identifies a concern 
for any member of the group other than the group representative. For 
example, the testing representative for the M85 group (consisting of 
methanol formulations containing 50-95 percent methanol) is an M85 base 
fuel containing 85 percent methanol by volume. In the future, other 
methanol fuels could enter the market and be used extensively, 
triggering a concern for exposure and potential health risks. In this 
case, EPA could require Tier 3 testing for another methanol formulation 
within the M85 group.
    Another possible Tier 3 effort could involve the testing of 
combinations of oxygenates. Although groups for non-baseline gasolines 
are defined on the basis of individual oxygenates, EPA could request 
additional testing to address mixtures of oxygenates under Tier 3 
authority. For example, if a registered non-baseline gasoline 
formulation containing a mixture of oxygenates is widely produced and 
used, resulting in high exposures and potential health risks, EPA could 
require testing of this formulation under Tier 3. Tier 3 tests for this 
formulation could involve standard Tier 1 or Tier 2 tests, testing for 
other endpoints not addressed under Tier 2, or testing for other areas 
of concern (e.g., exposure analysis, environmental testing, etc.).
    EPA also retains the authority to require additional testing using 
different vehicle/engine technologies and/or emission generation 
specifications than those prescribed for Tier 1 and Tier 2 analyses. To 
reduce the costs of the program, today's action requires the use of a 
single vehicle model for emission generation. However, if EPA 
determines that emissions from other applicable vehicle/engine 
technologies might differ significantly for a given fuel or additive/
base fuel mixture, then EPA may require additional emission 
characterization and/or toxicological testing under Tier 3 using 
different vehicle/engine technology. Also, although standard Tier 2 
tests are to be conducted using non-catalyzed emissions (as discussed 
in Section VI.A), EPA could require the use of catalyzed emissions for 
testing under Tier 3.
    When a determination has been made that Tier 3 testing is required, 
EPA will inform the responsible manufacturer by certified mail of the 
purpose and nature of the testing to be performed along with a schedule 
for compliance and submittal of the Tier 3 report to EPA. EPA will also 
publish a notice in the Federal Register, notifying the public that the 
letter to the manufacturer is available in the public record for review 
and comment. The affected manufacturer's comments and EPA's response to 
these comments will be placed in the public record, as well. After 
receipt and review of all comments received (or, if no comments are 
received), EPA will publish a notice of final action on the proposed 
Tier 3 requirements in the Federal Register.
    EPA proposed in the NPRM to provide the responsible manufacturer or 
group a 30-day comment period to respond to EPA's requirements under 
Tier 3. In their commentary, the regulated industry asked EPA to extend 
this comment period to 60 days to allow for appropriate time for 
review, analysis, and preparation of a written response to EPA 
regarding the designated protocol(s) to be used for Tier 3 tests. EPA 
judged that this request was reasonable, thus today's rule gives F/FA 
manufacturers 60 days to comment on the EPA-prescribed Tier 3 
requirements.
    The responsible manufacturer is expected to submit detailed 
protocols for review and approval by EPA prior to beginning Tier 3 
testing. Tier 3 tests must comply with the pre-approved specifications 
given by EPA. If manufacturers experience unforeseen difficulties while 
conducting the prescribed Tier 3 tests approved by EPA (e.g., excess 
mortality observed half-way through a chronic bioassay), they will be 
allowed to request a modification of the requirements. This mechanism 
would apply to unusual circumstances that are outside the control of 
the manufacturer. If testing problems are identified, EPA must be 
notified as soon as possible so that requirements can be modified.

B. Criteria for Referral to Tier 3

    This section presents some of the guidelines and considerations 
which EPA will use in determining the necessity for additional testing 
under a discretionary Tier 3 testing approach. Consistent with the 
discretionary decision-making process for Tier 3, this discussion is 
not intended to provide an exhaustive, limiting, or definitive listing 
of relevant criteria.
    The decision to require manufacturers to submit additional testing 
on the health, environmental, or welfare effects of F/FA emissions will 
take into account the cumulative information provided by Tiers 1 and 2, 
including previous scientific data, emissions characterization data, 
biological test results, and any ancillary information which may be 
available to EPA. Thus, decisions to require Tier 3 level testing will 
be made only after all the requirements of Tiers 1 and 2 have been 
adequately satisfied (with the exception of special cases as discussed 
in Section VIII.D). Adherence to this principle will prevent 
unnecessarily costly or poorly targeted decisions based on piecemeal, 
out-of-context information, and will promote more precise 
identification and evaluation of data gaps, and more cost-efficient 
coordination of potential test requirements.
    Ultimately, EPA must be able to decide whether or not the use of a 
fuel or fuel additive is likely to create unacceptable health or 
welfare risks. If a risk decision is made possible by the information 
from Tiers 1 and 2, then Tier 3 will not be required. However, if such 
a risk decision cannot be made on the basis of the Tiers 1 and 2 data, 
then Tier 3 testing will be mandated. Therefore, to make a 
determination on the need for Tier 3 testing, EPA scientists will 
evaluate the extent to which the results of Tiers 1 and 2 are adequate 
for such decisions, guided by the basic principles of risk assessment.
    A risk assessment requires the merging of a health effects 
assessment (including hazard identification and concentration-response 
relationship) and an exposure assessment. Such an assessment can range 
from a qualitative to a highly quantitative analysis, depending upon 
the extent of the available data. EPA recognizes that a quantitative 
assessment might not be possible at the end of Tier 2.\59\ However, 
Tiers 1 and 2 might indicate that little hazard is present and that 
exposures may be quite low and limited geographically. In such a case, 
there may be no reason to pursue further testing at the Tier 3 level to 
improve risk assessment information. On the other hand, Tiers 1 and 2 
might suggest that a hazard is likely and that exposures could be 
significant because of the production volume and ubiquitous use of a 
product, but the data may still be inadequate for a quantitative risk 
assessment. In this case, Tier 3 testing could be indicated to provide 
the needed information.
---------------------------------------------------------------------------

    \59\However, if adequate information exists, EPA does not rule 
out the possibility of conducting such risk assessment.
---------------------------------------------------------------------------

    In general, the principles and critical data elements of the risk 
assessment process will provide a useful guide for identifying whether 
meaningful information gaps remain and for determining the specific 
objectives of potential Tier 3 testing. However, EPA does not intend to 
conduct a formal risk assessment as part of its decision on whether to 
promote a fuel or fuel additive (or group) to Tier 3. Rather, EPA will 
evaluate the quality and certainty of the toxicity and concentration-
response data and consider qualitatively whether such data weighs in 
favor of or against further testing. A formal risk assessment will be 
more likely to be developed at such a time that there is a need for 
action to control or prohibit a product under the regulatory authority 
of CAA section 211(c).
    The following sections discuss key factors which EPA will consider 
in identifying the need for and content of Tier 3 testing.
1. Statistical Issues
    As previously mentioned, scientific judgment will be exercised in 
determining whether Tier 3 testing is indicated. An important factor in 
such judgments will be the interpretation of and significance ascribed 
to ``negative'' results obtained in Tiers 1 and 2. To address this 
issue, EPA will consider statistical information such as the 
probability of Type I and Type II errors.
    A Type I error occurs when a false positive conclusion is made, 
while a Type II error is a false negative conclusion. The acceptability 
of a specific Type II error is related to the acceptability of false 
negatives in the particular study being performed. For example, from a 
toxicological perspective, screening assays often have a relatively 
high probability of producing false negative (Type II) outcomes, since 
some major aspects of organ or tissue toxicity are not being examined. 
Thus, an acceptable Type II error for screening assays will typically 
be high. However, the level of Type II error considered acceptable 
should be tempered by the goal of the study. A higher false negative 
conclusion (e.g., Type II error of 0.2) will generally be acceptable if 
it refers to an effect of minimal severity at a high-exposure test 
level relative to ambient concentration and if few people are likely to 
be exposed. The converse will also hold true.
    Scientifically sound statistical analyses are a crucial part of any 
reliable study and will provide key information for EPA to make 
judgments on whether or not Tier 3 testing is needed. While it is not 
feasible to list all possible scenarios and results for each Tier 2 
endpoint, the above discussion describes how some of the statistical 
factors will be incorporated into EPA's decisions.
2. Exposure Assessment
    The Tier 1 requirements will provide EPA with information on the 
composition of F/FA emissions and potential exposures to F/FA products. 
This information includes: (1) Types and emission rates of speciated 
emission components, (2) possible literature search findings on 
ambient, occupational, or epidemiological exposures, (3) literature 
information on the potential fate and environmental effects, and (4) a 
qualitative exposure analysis (submitted by the manufacturer or group) 
based on the production and use of the F/FA product(s). As mentioned 
above, this information will be considered qualitatively by EPA in 
determining the extent of potential exposures and whether additional 
testing or analysis is needed to assess risks quantitatively. As 
discussed earlier, EPA has the authority to require exposure modeling 
or analysis under Tier 3.
    Significant public health concerns might sometimes be revealed by 
the submitted information on product composition, total annual 
production volume and market distribution data, and emissions data. 
This might be the case, for example, if there was a significant annual 
release of emission compounds with known toxicities, or if the 
anticipated exposures approached or exceeded current estimates of 
apparently safe levels of known toxicants. In the case of high-volume 
fuels and their associated bulk additives, EPA will generally assume 
that human and environmental exposures will be of sufficient level and 
extent that significant observed adverse effects could indicate a need 
for follow-up in Tier 3. This exposure assumption reflects the high 
production and consumption of these products, either at the present 
time or as anticipated in the future. Thus, decisions to refer these 
products to Tier 3 will be based on the degree to which additional 
testing is needed to clarify the results and potential health effect 
and environmental implications identified in the previous tiers. On the 
other hand, it cannot be assumed that fuel additives used in relatively 
low concentrations or produced in relatively low volumes will 
automatically be excused from Tier 3. For these products, test results 
indicative of severe health effects and/or high exposure levels (e.g., 
during consumer use) might be cause for escalation to Tier 3.
3. Health Assessment
    General criteria for evaluating the potential public health effects 
associated with fuel and additive emissions will include: (1) The 
number of positive and negative outcomes related to each endpoint, (2) 
the identification of a concentration-effect relationship, (3) the 
statistical sensitivity and significance of such studies, (4) the 
severity of the observed effects (e.g., whether the effects will likely 
lead to incapacitating or irreversible conditions), and (5) the 
consistency and clarity of apparent mechanisms, target organs, and 
outcomes. Additional parameters which will influence the decision on 
whether to require Tier 3 will include: (1) The nature and amount of 
known toxic agents in the emissions stream and (2) the observation of 
lesions which specifically implicate inhalation as an important 
exposure route for inducing adverse health effects.
    These criteria will be evaluated in conjunction with the Tier 1 and 
Tier 2 results to determine whether or not higher level testing is 
needed. In this decision, both the biological and statistical 
significance of the Tiers 1 and 2 results will be taken into account. 
Generally, escalation to Tier 3 may be judged necessary when remaining 
uncertainties about the significance of observed outcomes and/or 
potential exposures interfere with EPA's ability to make reasonable 
estimates of potential health risks. On the other hand, if no 
statistically significant effects are obtained at any exposure level in 
a scientifically sound Tier 2 study (or existing test submitted in lieu 
of Tier 2 testing and not contradicted by other published reports of 
equal or greater reliability), and if other major sources of concern do 
not arise (e.g., toxic effects of structurally related compounds), then 
Tier 3 testing is not likely to be required for the endpoint in 
question.
    This discussion assumes that relevant, high-quality statistical 
analyses have been done to permit the negative test results to be 
properly evaluated and interpreted. The statistical analyses 
recommended for Tier 2, including determination of Type I and II error 
(as discussed in Section IX.B.1), should enable reasonable conclusions 
to be drawn as to the significance of negative findings. Factors to be 
taken into account include the toxicological nature of the findings and 
the exposure levels used in the test. For example, if the statistical 
analyses are applied to a ``severe'' endpoint (e.g., major fetal 
abnormalities, major lung pathology, etc.) and the exposure in question 
is moderate, then a relatively low Type II error level may be 
appropriate. In contrast, if a high concentration limit test causes a 
relatively minor effect (e.g., a small change in estrous cyclicity), a 
higher Type II error may be allowed, effectively increasing the chance 
of false negative conclusions.
4. Evaluation of Tier 2 Results
    The specific outcomes which will be considered positive and 
negative results for each Tier 2 test were mentioned briefly in the 
previous descriptions of Tier 2 requirements (see Section VIII.B) and 
are defined and interpreted more precisely in the regulatory text of 
this final rule. For example, three primary assays (i.e., Salmonella, 
MN, and SCE) are included in Tier 2 for the screening of 
carcinogenicity and mutagenicity. As compared with appropriate 
controls, a statistically significant concentration-related positive 
response in any one of these assays could be cause for concern, as will 
be positive outcomes for at least one concentration in two or more of 
these tests. Such outcomes will be indicative of mutagenic and/or 
carcinogenic risk. Positive results will also indicate that the 
emissions could initiate some of the mechanisms involved in 
carcinogenesis. However, these results will generally not in themselves 
be sufficient to determine whether the emissions were in fact 
carcinogenic because the development of cancer is a multi-step process. 
Depending on the internal and historical consistency of the results and 
their relationship to projected exposures, further testing might be 
required to determine the significance of the mutagenic and/or 
carcinogenic activity/risks in human populations exposed by inhalation. 
In contrast, if no statistically significant results are obtained in 
the three assays and no conflicting results are found in the literature 
or in any other Tier 2 tests, then Tier 3 follow-up of potential 
carcinogenic/mutagenic effects is not likely to be required.
    To take another example, determination of the need to investigate 
further reproductive or teratogenic risks will take into account the 
outcome of the results of the estrous cyclicity measurements, the 
histopathological evaluation of reproductive organs, the outcome of the 
fertility screen, and the outcome of the teratogenicity evaluation. If 
negative results are obtained in the above evaluations (according to 
statistically sound principles), and if these results are not refuted 
by the existing literature, then additional testing is not likely to be 
required at the Tier 3 level for reproductive or teratogenic effects. 
Positive results for the teratogenicity study will include a decrease 
in neonatal viability relative to that in control studies, a 
significant change in the proportion of viable male versus female 
fetuses or offspring, the presence of soft tissue or skeletal 
abnormalities, and an increased rate of embryonic or fetal resorption. 
Other positive outcomes related to reproductive effects, such as 
decreased fertility, decreased litter size, abnormal changes in vaginal 
cytology or reproductive organ histopathology, will be indicative of 
hazards to the adult reproductive systems. The need for additional 
evaluation under Tier 3 will depend on the specificity, severity, and 
consistency of results, the presence or absence of a concentration-
effect relationship, and the significance of these outcomes in view of 
projected exposures. The greater the remaining uncertainty regarding 
the risk of reproductive or teratogenic effects after analysis of such 
factors, the higher will be the likelihood that Tier 3 will be 
required.
    Similarly, consistent negative results (according to statistically 
sound principles) obtained in other Tier 2 tests, in the absence of 
significant related concerns raised in the literature, will (in all 
likelihood) make Tier 3 unnecessary. If adverse effects are found at 
Tier 2 and/or reported in the literature, EPA will determine if Tier 3 
follow-up is required by attempting to evaluate the nature, severity, 
and significance of the findings in light of the likely exposures. If 
EPA determines that Tier 3 testing is required to resolve the remaining 
uncertainties, the Tier 3 requirements will reflect both positive and 
negative results. For example, if the results of Tier 2 were positive 
for pulmonary effects but negative for neurotoxicity (according to 
criteria discussed earlier), and if these results were consistent with 
the literature, only pulmonary toxicity would be a likely candidate for 
Tier 3 follow-up testing.

C. Potential Tier 3 Tests

    To be most cost-effective, Tier 3 testing will be designed to 
address specific data gaps regarding health effects endpoints of 
concern or health and environment-related issues requiring further 
analysis. For instance, Tier 3 requirements could potentially include 
further emission characterization procedures, perhaps involving 
additional vehicles and/or more rigorous pre-conditioning methods, to 
identify and quantify harmful emission products with greater precision. 
Higher-order modeling calculations or exposure field studies could be 
required to resolve uncertainties in the Tier 1 emissions exposure 
information. Health or welfare effects testing requirements will be 
aimed at providing sufficient information to make sound conclusions 
about the degree of health or welfare risk. If more than one endpoint 
is of concern, EPA will attempt to reduce testing costs by permitting 
combined protocols insofar as possible.
    Tier 3 tests for specific endpoints could require the determination 
of a NOAEL. Depending on the endpoint under evaluation, consideration 
will be given to including a mid-duration examination in the case of 
chronic inhalation tests. A mid-duration evaluation will be useful for 
affirming the adequacy of exposure levels and, in some cases, might 
enable interim risk conclusions to be drawn which will avoid the need 
for further examination. Inhalation studies will generally make use of 
rodent species, but higher order mammals could be required.
    While Tier 3 testing requirements will be targeted to critical 
areas of concern, EPA will also exercise its judgment to avoid the 
false economy of establishing overly narrow requirements. Just as 
requirements for too many assays would be wasteful of resources, 
requirements for too few assays might result in inconclusive findings, 
creating needs for still further testing at greater total expense than 
would have been necessary at the start. Similarly, EPA will consider 
the value of including secondary evaluations as useful and low-cost 
adjuncts to tests already required. For example, if the histopathology 
of a specified target organ was the primary examination required at the 
conclusion of an inhalation exposure, other organs could be weighed and 
saved in storage for a limited time period, at low incremental expense. 
If indicated, these other organs would then be available for subsequent 
examination, avoiding the possible need to repeat the chronic 
inhalation procedures to assess the effects on other organs.
    Because the specific health testing requirements which will be 
imposed in Tier 3 will be tailored to individual circumstances, precise 
test guidelines cannot be provided in advance. However, some examples 
of testing scenarios which might be required under Tier 3 are cited in 
Sec. 79.54 in the accompanying regulatory text. Where possible, 
existing standard guidelines for these tests are referenced. It should 
be recognized, however, that such guidelines might need to be revised 
to accommodate emission inhalation requirements and/or to evaluate 
certain structures or functions which the current guidelines do not 
adequately address. Study parameters which might require modification 
include exposure routes and concentrations, species selection, number 
of animal subjects, examination procedures and frequencies, and 
analytic requirements. Furthermore, interim advances in the underlying 
science and testing technology may provide superior approaches which 
could be available for use by the time Tier 3 requirements are 
implemented.

X. Special Provisions

    The following sections describe special provisions included in the 
F/FA registration program to avoid duplication of effort, to alleviate 
the financial impact on small businesses, and to ease the burdens of 
the program on the regulated industry in general.

A. Experimental F/FAs

    EPA requested comments on the possibility of providing a temporary 
program exemption or deferment for experimental F/FAs. Eligibility for 
this special provision was to be limited to unregistered products 
(i.e., F/FAs which are not registered as of the effective date of this 
final rule) or registered products that had not been placed into 
wholesale or retail commerce prior to promulgation of this rule.
    The regulated industry provided comments questioning the need for a 
special provision for experimental F/FAs under this rulemaking because 
a provision for experimental products already exists under existing 
registration rules. EPA agrees with the comments received and today's 
rule relies on existing regulations under 40 CFR 79.4(a)(3) and 
79.4(b)(2) for the exemption of experimental F/FAs. 40 CFR section 
79.4(a)(3) exempts fuels used for research, development or testing, and 
40 CFR section 79.4(b)(2) similarly exempts fuel additives. Based on 
these existing provisions, any designated F/FA product sold to 
automobile, engine, or component manufacturers for research, 
development or test purposes, or sold to automobile manufacturers for 
factory fill, and not in any case offered for commercial sale to the 
public, is exempted from the registration requirements of today's rule.

B. Relabeled Products

    A company's product is registered as ``relabeled'' if it is simply 
a repackaged and rebranded version of a formulation which is already 
registered by another manufacturer and is procured from that 
manufacturer for sale or use. Requiring companies which sell relabeled 
products to conduct the testing program in today's rulemaking would 
clearly duplicate the efforts of the original manufacturer. Thus, under 
the authority of CAA section 211(e)(3)(C), which provides that the 
Administrator may ``exempt any person from such regulations with 
respect to a particular fuel or fuel additive upon a finding that any 
additional testing of such fuel or fuel additive would be duplicative 
of adequate existing testing,'' today's rule includes a special 
provision exempting relabeled products from the evaluation and testing 
requirements. For relabeled products, only basic registration 
information will be required, as described below in Section XII.A.
    About half of the 4,800 fuel additives registered (as of March 
1994) are relabeled products. Manufacturers of these relabeled products 
will therefore not be required to comply with the health and welfare 
effects assessment provisions of the F/FA registration program.

C. Aerosols

    Several commenters requested an exemption for aftermarket aerosol 
additives because of the nature of their mode of application and their 
low frequency of use. EPA examined available data on aerosol products 
and concluded that the required testing procedures of this rule are not 
well suited to this type of product. The evaluation of currently 
registered additives indicates that aerosols include carburetor 
cleaners and engine starters that are sprayed into the air intake valve 
of the engine and are used only intermittently and, even then, for only 
a very brief period of time (i.e., a few seconds). Because of their 
intermittent use and method of use, it is unlikely that their 
recommended application would affect the overall characteristics of 
vehicle emissions. Direct exposure to the aerosol product itself is 
likely to be a more important source of potential hazards than is 
exposure to its combustion or evaporative emission products. Thus, the 
evaluation of potential health and welfare effects of aerosols would 
require a different testing regimen tailored to the specific nature of 
these products.
    Today's rule therefore establishes a special provision for 
aerosols. Manufacturers of aftermarket aerosol additives are required 
to provide only the basic registration data required for all F/FA 
manufacturers (see Section XII.A), plus a literature survey of existing 
information on their products and a discussion on the potential 
exposures. Thus, the standard emission characterization and Tier 2 
tests are not required for aerosol products. The literature search (as 
described in Section VII.A) must include existing data on potential 
health and welfare effects on the uncombusted aerosol product as a 
whole and on the individual components of the product. The analysis of 
potential exposures should be based on the total annual production 
volume data and the market distribution of the product, as explained in 
Section VII.C. EPA will review the submitted information and will 
determine if there is a need to require testing on a case-by-case 
basis. Thus, if testing of an aerosol is prescribed by EPA, the testing 
is to be performed under Tier 3. Using this approach, EPA will be able 
to tailor specific tests (as needed) addressing the particular problems 
related to aerosol product exposure.

D. Small Business Provisions

    In the NPRM, EPA requested comments on the possibility of 
establishing special provisions for small businesses. The regulated 
industry expressed concerns regarding the costs of the program and the 
potential impact of the F/FA regulations on the financial status of 
small companies. EPA understands that small businesses might be 
particularly affected by the F/FA regulations due to their smaller 
resource base, generally lower rate of representation in trade 
organizations and, consequently, their potentially limited opportunity 
to participate in grouping and cost-sharing arrangements. In view of 
these circumstances, EPA is including in today's rule special 
provisions to alleviate the economic impact of this rule on small 
manufacturers of F/FAs, taking into consideration the comments 
received.
    EPA examined the distribution of currently registered F/FA 
companies across various sales ranges and identified a sales level 
which will define a ``small business'' for the purposes of this rule. A 
comparison between registered fuel manufacturers and registered 
additive manufacturers indicates that the distribution across sales 
ranges is similar for both industries. Each industry appears to be made 
up of many small companies and relatively few larger companies. 
Approximately a fifth of registered fuel manufacturers and a third of 
registered additive manufacturers have sales above $100 million. For 
both industries, most of the companies with sales under $100 million 
tend to cluster under $50 million. Thus, within each of the industries, 
companies tend to cluster above $100 million and below $50 million in 
sales. Furthermore, companies with sales below $50 million also tend to 
have sales below $10 million, especially within the additive industry.
    Based on the analysis of F/FA manufacturers' sales data, EPA 
identified $50 million as a reasonable sales level for differentiating 
between small and large companies within each industry.\60\ As a 
result, this final rule defines a small business as any motor vehicle 
fuel or fuel additive manufacturer with total annual sales of less than 
$50 million.\61\ The small business provisions established for this 
rule are defined below.
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    \60\In cases where subsidiary, divisional, or other complex 
business arrangements exist, the business entity to which this sales 
level pertains is the parent company with ultimate ownership. The 
``ultimate'' parent is defined as the uppermost headquarters or 
topmost company encompassing all related parents, subsidiaries, 
divisions, branches, or other operating units. This definition 
follows that used by the Small Business Administration. It also 
helps to ensure that companies will not subdivide merely to become 
eligible for the small business provisions of this program.
    \61\``Total annual sales'' means the average of the 
manufacturer's sales revenue in each of the previous three years 
(i.e., the three years prior to the submittal of the supplemental 
notification form required under this rule).
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    Special provisions for small
F/FA companies were developed based on assumptions regarding the degree 
to which manufacturers will have the opportunity to group with other 
manufacturers to share costs, and the degree to which information on 
similar products can be expected to be submitted by larger companies. A 
review of EPA's registration data base shows that nearly every 
registered F/FA manufacturer produces at least one baseline or non-
baseline product. Hence, even if generous special provisions are 
provided for small manufacturers of these products, EPA is assured that 
test data on such products will still be submitted by other, larger 
companies which do not qualify for the special provisions. Therefore, 
in regard to registration of products in the baseline and non-baseline 
categories, this final rule requires small businesses (i.e., companies 
with annual sales less than $50 million) to submit only the basic 
registration data for their baseline and/or non-baseline products. Such 
companies are not required to meet the Tier 1 and Tier 2 requirements 
in order to register their baseline and non-baseline F/FAs. Since the 
larger companies will still be submitting the data for these products, 
this provision does not compromise EPA's ability to get the necessary 
information to evaluate the potential health and welfare effects of 
baseline and non-baseline products.
    The above assumption, however, is not valid in the case of atypical 
F/FAs. Analysis of current registrations shows that there are fewer 
atypical products than baseline products and fewer large companies that 
produce atypical F/FAs. Therefore, if the same small business 
provisions described above for baseline/non-baseline F/FAs were also 
applied to atypical F/FAs, then there would be no data submitted for 
many atypical F/FAs. EPA would then be unable to evaluate the health 
and welfare effects for these atypical products. Nevertheless, EPA 
realizes that the testing program will be particularly burdensome for 
very small manufacturers of atypical F/FAs. In part, this is due to the 
added compliance requirements for atypical products (e.g., more 
rigorous mileage accumulation requirements prior to emission generation 
(see Section VI.E) and more extensive emission characterization 
requirements (see Section VII.B). An even greater impact results from 
the fact that grouping opportunities for these manufacturers might be 
quite limited. Not only do atypical products tend to be unique, but 
also, the smallest manufacturers rarely are members of the dominant 
trade associations which are most likely to take the lead in organizing 
and administering F/FA group functions. Thus, high program costs could 
fall on manufacturers with low financial resources and perhaps few 
opportunities to share the costs.
    Based on the above reasons, this final rule includes additional 
special provisions applicable to the atypical F/FAs of small 
businesses. However, these provisions apply to fewer manufacturers and 
are less liberal than those established for baseline and non- baseline 
products. Specifically, for manufacturers of atypical products with 
less than $10 million in annual sales (rather than the $50 million 
sales level applicable to manufacturers of baseline/non-baseline F/
FAs), the minimal requirements of the program include only basic 
registration and Tier 1 data. These requirements fall between those 
applicable to small manufacturers of baseline/non-baseline F/FAs and 
the general requirements of the program for larger manufacturers.
    In summary, manufacturers qualifying for small business special 
provisions for their baseline and non-baseline products (having less 
than $50 million annual sales) are excused from both Tier 1 and Tier 2 
requirements, while small manufacturers qualifying for special 
provisions for their atypical products (having less than $10 million 
annual sales) are excused only from Tier 2 requirements for these 
products. Since small business provisions are based on both the annual 
sales and the product category, the possibility exists for a 
manufacturer to have some products excused from program requirements 
while having to comply with testing requirements for others. For 
example, an additive manufacturer having both baseline and atypical 
products and annual sales of $30 million can use the special small 
business provision for the baseline additives, but not for the atypical 
products.
    Pursuant to CAA section 211(b), F/FAs which are excused from any 
program requirements under these special provisions may still be 
subject to testing under Tier 3 at EPA's discretion (on a case-by-case 
basis). The Tier 3 testing might include (but would not be necessarily 
limited to) information which would otherwise have been required under 
the provisions of Tier 1 and/or Tier 2.

XI. Timing and Compliance Requirements

    The timing and compliance requirements for the F/FA registration 
program are dependent on the type of product and the registration 
status of the product. As discussed in Section III.A of this preamble, 
both Tier 1 and Tier 2 are mandatory requirements for all F/FAs (or 
groups), except as may be modified by any applicable special 
provisions. Special provisions affecting the content and/or timing of 
these requirements are discussed in Sections VII.A.3, VIII.D, and X of 
this preamble. For F/FAs registered as of the effective date of this 
rule, Tier 1 data and evidence of a suitable contractual arrangement 
for satisfactory completion of Tier 2 requirements must be submitted to 
EPA within three years of the effective date. The results of Tier 2 
must be submitted to EPA no later than six years from the effective 
date. The schedule for completion of any Tier 3 requirements which EPA 
may prescribe will be determined based on the nature of the particular 
requirements. The general reporting format for submittal of all of 
these requirements is described in Section XII.
    For registrable F/FAs, i.e., F/FAs not registered as of the 
effective date of this rule but meeting the criteria for grouping with 
a currently registered fuel or bulk additive in the same fuel family, 
the content and timing of requirements is essentially the same as for 
the currently registered F/FAs. Thus, manufacturers of these products 
will be granted registration and be permitted to market registrable F/
FAs upon EPA's receipt of basic registration data for such products. In 
contrast, for manufacturers of new F/FAs (i.e., F/FAs that are not 
currently registered and do not meet the registrable criteria), all 
testing requirements must be completed prior to registration and 
introduction into commerce, including Tier 3 when prescribed by EPA.
    After receipt of Tier 1 and/or Tier 2 data, EPA will determine 
whether the submitted information is in compliance with the specified 
guidelines and whether further testing of a particular fuel or fuel 
additive is required under the provisions of Tier 3. For registered F/
FAs, EPA intends to determine the adequacy of the submitted data within 
two years after receipt. However, if EPA is unable to inform the 
registered manufacturer of the adequacy of the Tier 1 and/or Tier 2 
data within two years after submittal, EPA retains the authority to 
require that satisfactory data be submitted if, upon subsequent review, 
EPA finds that the original submittal was inadequate for compliance. In 
such a case, EPA will not hold the manufacturer liable for penalties 
for violating this rule from the time period between the date on which 
the data were due and the date on which EPA informs the responsible 
manufacturer of a violation. Regarding new F/FAs, EPA will send a 
notification of compliance within six months after submission of Tier 1 
and Tier 2 data. If the manufacturer of the new F/FA product does not 
receive a notification of compliance with Tier 1 and Tier 2 within this 
time frame, then the manufacturer should assume that the Tier 1 and 
Tier 2 requirements have been satisfactorily met. EPA's determination 
of the need for Tier 3 testing for new F/FAs will occur within six 
months after EPA notifies the manufacturer of satisfactory compliance 
with Tier 1 and Tier 2 requirements or within twelve months of the Tier 
1 and Tier 2 submittal, whichever occurs first.
    If Tier 3 testing is deemed necessary, EPA will notify the 
responsible manufacturer (or group) by certified letter of the specific 
Tier 3 requirement(s) along with a schedule for compliance and a 
deadline for submittal of the Tier 3 report to EPA (see Section IX.A). 
This final rule gives the responsible manufacturer (or group) 60 days 
to comment on the prescribed Tier 3 requirements, compliance schedule, 
and submission deadline. In the event that EPA receives no comment 
within the given period, the manufacturer will be assumed to have 
consented in full to the prescribed Tier 3 requirements. Compliance 
with Tier 3 requirements is not optional.
    Registered (and registrable) F/FAs required to undergo Tier 3 
testing will retain their registration for that time determined to be 
necessary for the completion of Tier 3 tests. This registration will be 
contingent on the satisfactory compliance with the Tier 3 requirements 
according to a timetable determined by EPA to be appropriate to those 
requirements. When Tier 3 is prescribed for new F/FAs (i.e., those not 
meeting the registrable criteria), EPA may withhold registration until 
completion of all testing requirements. For new F/FAs, EPA will 
determine whether the Tier 3 requirements have been met within one year 
of receiving the Tier 3 submittal. If Tier 3 requirements are 
satisfied, then EPA will send a notification to the manufacturer 
granting registration to the new F/FA product. Registration of new F/
FAs will not occur until that time when EPA determines that all Tier 3 
requirements have been satisfactorily met.
    As described above, EPA's review times for data on new F/FAs are 
shorter than those for registered F/FAs. The reason for this 
discrepancy is that manufacturers of new F/FAs are barred from 
marketing such products until EPA approves their compliance with all 
testing requirements and grants them registration. On the other hand, 
manufacturers of registered products can maintain their registration, 
and thus their ability to sell their products, while EPA is reviewing 
their submitted data. Thus, to ensure that undue hindrance is not 
created for manufacturers of new products wanting to enter the 
marketplace, EPA has abbreviated the review times for new F/FA 
products.
    Notwithstanding the granting of a registration (or continued 
registration for registered F/FAs), if EPA determines that a fuel or 
fuel additive causes or contributes to air pollution that may 
reasonably be anticipated to endanger the public health or welfare, 
then EPA could invoke available regulatory authority under CAA section 
211(c). Referral to Tier 3 is not required for EPA to begin a 
regulatory action under 211(c).
    If additional testing is needed to make up for deficiencies in 
information content or testing technique/procedures related to Tier 1, 
Tier 2, and/or Tier 3, then the original compliance deadlines will 
still be in force. Manufacturers of existing products who fail to 
submit data in the prescribed time frames or who submit data from tests 
that do not comply with the specified guidelines will be in violation 
of this rule and will be subject to the penalties specified in CAA 
section 211(d). According to CAA section 211(d), persons who fail to 
submit any information or conduct any tests required by the 
Administrator under CAA section 211(b) shall be liable to the United 
States for a civil penalty of not more than $25,000 for every day of 
such violation plus the amount of economic benefit or savings resulting 
from the violation. Each day after the due date for submission of data 
will constitute a separate day of violation. Civil penalties will be 
assessed in accordance with CAA sections 205(b) and (c), which permit 
EPA to proceed either in court or in an administrative action. If a 
group of manufacturers commits to performing joint testing, each 
manufacturer would separately be in violation of the rule. However, the 
Administrator would retain the authority to remit or mitigate any 
penalty under CAA section 211(d).
    In addition to the above penalties, the district courts of the 
United States have jurisdiction to compel the furnishing of information 
and/or the conduct of tests required under CAA section 211(b). This 
means that, in addition to the financial penalties, persons failing to 
submit data or comply with the specified guidelines would still need to 
submit the data originally required. Furthermore, if EPA determines 
that the data requirements of the rule were not met, EPA could revoke 
the registration of the fuel(s) or additive(s) in question.
    Because EPA recognizes that unusual circumstances, outside the 
control of the manufacturer, may occasionally interfere with the 
manufacturer's ability to comply with the provisions of the rule, 
today's rule contains a mechanism to allow manufacturers to request 
modification of the requirements under some specific circumstances. 
This special mechanism allows persons who experience unforeseen 
problems or accidents in conducting the EPA-prescribed tests to request 
modification of the requirements in order to avoid being in violation 
of the rule. This mechanism would apply to unusual mechanical problems 
or other unavoidable problems that could arise during the performance 
of the required tests. The modification requests must be submitted as 
soon as the manufacturer is aware of the difficulty, but not later than 
thirty days following the event precipitating the request. Additional 
details on this special mechanism for modification of requirements is 
included in the regulatory text of this rule.

XII. Reporting Requirements

    The materials to be submitted to EPA include the basic registration 
data, a summary report with Tier 1 and Tier 2 results, and associated 
appendices. If the results of Tiers 1 and 2 are submitted at the same 
time, then the summary report must include both Tier 1 and Tier 2 
information and associated appendices, as described below. If Tier 1 
and Tier 2 results are submitted separately, then two individual 
reports must be provided to EPA, i.e., Tier 1 report and Tier 2 report. 
In such a case, each individual report must include the summary 
information applicable to the respective tier (including a cover page, 
executive summary, test substance information, a summary of tier 
results, conclusions, and associated appendices). If the Tier 2 report 
for registered F/FAs is not submitted within three years after the 
effective date of the final rule, then evidence of a suitable 
arrangement for completion of Tier 2 (e.g., a copy of a signed contract 
with a qualified laboratory to conduct the required Tier 2 tests) must 
be submitted to EPA prior to that date. F/FA manufacturers who must 
conduct additional testing under Tier 3 are required to submit a Tier 3 
report when the designated Tier 3 testing is complete. The nature of 
the information to be included in the basic registration data, reports, 
and associated appendices is described below.

A. Basic Registration Data

    The basic information already required for F/FA registration 
includes product and manufacturer identification, concentration and 
purpose-in-use, and specific compositional data. Today's rule adds the 
following items to the basic registration data requirements: total 
annual production volume data, marketing distribution data, 
notification about group participation, and notification on the use of 
special provisions (i.e., relabeled products, aerosols, and small 
business, as discussed in Section X). Manufacturers of F/FAs registered 
as of the effective date of this rule must submit the additional basic 
registration data items to EPA within six months of that date. Other 
manufacturers are strongly encouraged to submit the basic registration 
data prior to starting the evaluation tiers (i.e., Tier 1, Tier 2, and/
or Tier 3).
    The production volume information is to be reported in units of 
gallons per year for F/FA products that are generally sold in liquid 
form and kilograms per year for F/FA products that are generally sold 
in solid form. For F/FAs already in production, the submitted figure 
must reflect the most recent annual period as well as the volume 
projected to be produced in the third subsequent year. For products not 
yet in production, the best estimate of expected total production 
volume during the third year of production must be provided. Market 
distribution data for each product must also be provided. For fuels and 
bulk additives, the distribution data must be reported as the percent 
of total annual sales volume marketed in each Petroleum Administration 
for Defense District (PADD), as defined in Sec. 79.59(b)(3) of this 
rule. For aftermarket additives, the distribution data must be reported 
as the percent of total annual sales volume marketed in each state. For 
products not yet in production, the manufacturer must report projected 
distribution data by PADD or state, as applicable.
    Manufacturers of F/FAs registered as of the effective date of this 
rule who intend to comply with registration requirements as part of a 
group must identify the person or entity which is organizing the 
testing for the applicable group. Similarly, if an applicant is relying 
on another manufacturer's (or group's) previous registration materials 
in compliance with the testing requirements for an unregistered 
product, then the other manufacturer or group must be identified. In 
addition, the manufacturer of the unregistered product must provide 
evidence that the original submitter has been notified and that 
reimbursement will occur.
    The basic registration data must be submitted (or resubmitted) 
individually for each product being registered, using EPA forms which 
are in effect at the time of the submittal. This requirement pertains 
to all F/FA products registered as of the effective date of this rule, 
including relabeled products, as well as those for which first-time 
registration is sought after promulgation of this rule. If the basic 
registration data previously submitted for a currently registered fuel 
or additive is accurate and complete, then a statement asserting that 
this is so will suffice in lieu of the submittal of duplicate 
information. A finding by EPA that this information is not, in fact, 
accurate and complete as claimed will result in the report being 
considered inadequate.
    A fuel manufacturer may at any time modify an existing fuel 
registration by submitting a request to EPA to add or delete a bulk 
additive to the existing registration information for such fuel 
product, provided that any additional additive must be registered by 
EPA for use in the specific fuel family to which the fuel product 
belongs. The addition or deletion of a bulk additive to a fuel 
registration does not necessarily cause the fuel to be considered 
``new''. However, if the change affects the grouping of such registered 
fuel, it may affect the testing responsibilities of the fuel 
manufacturer.

B. Summary Report

    This report will provide a summary of the evaluation procedures, 
results, and conclusions, pertaining to Tier 1 and/or Tier 2 
requirements.\62\ References used to support Tier 1 and/or Tier 2 
conclusions must be cited in the report. A cover page must be included, 
identifying the test substance, the manufacturer's name and address, a 
designated contact person and phone number, and grouping information 
(if applicable). The grouping information must identify the group name 
or grouping criteria, all products and manufacturers to which the 
report pertains, and the name and address of the responsible 
organization or entity reporting for the group. The body of the summary 
report must be divided into the following sections.
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    \62\These reporting requirements may pertain to separate 
submittals for Tier 1 and Tier 2 or a single submittal for both 
tiers, depending on the relative timing of these compliance 
activities.
---------------------------------------------------------------------------

1. Executive Summary
    This section must include a brief description of the general 
results and conclusions for the tier(s) included in the report (i.e., 
Tier 1 and/or Tier 2), emphasizing information and test data which 
provide evidence for potential adverse health and/or welfare effects.
2. Test Substance Information
    This section must include a detailed test substance description, 
including (as applicable) base fuel parameter values or test fuel 
composition (if other than base fuel), and test additive composition. 
The base fuel description must include the types and concentrations of 
base fuel additive components and values for each of the parameters 
specified in the base fuel definition for the applicable fuel family. 
Similar parameter values must be identified for test fuels other than 
base fuels.
3. Tier 1 Summary
    This section is intended to provide an overview of Tier 1 analyses. 
Detailed procedural descriptions, tables, and other outputs are to be 
included in the appendices.
    a. Literature Search. The search methods must be described, 
including the identity of data bases and time periods accessed. Any in-
house and/or other unpublished studies included in the literature 
search must also be described briefly. The results and conclusions of 
the literature search with respect to potential health and welfare 
effects of the subject fuel or fuel additive must be summarized. If 
test documentation provided by the literature search is used to satisfy 
some or all of the other program requirements, the relevant studies 
must be discussed and their adequacy to fulfill the specific purposes 
of the associated program requirements must be justified. Finally, the 
person(s) or contractors conducting the search are to be identified.
    b. Emission Generation and Characterization. This section of the 
summary report must identify the vehicle selected and describe the 
procedures followed in vehicle/engine preparation and maintenance and 
in the generation, storage, and processing of emissions for testing. 
For group submissions, the report must include a complete description 
of the group representative used in the generation of emissions. A 
description of the analytic methods used to characterize the F/FA 
emission products must also be provided. Problems encountered in 
generating and/or characterizing the emissions must be discussed, 
including attempts to resolve the problems and their potential effects 
on testing outcomes. The laboratories performing these procedures must 
be identified.
    c. Exposure Analysis. This section must include a qualitative 
discussion on the potential exposures to the general, area-specific, 
and/or special at-risk population groups based on the production and 
use of the particular fuel or additive in question. For group 
submissions, the analysis must consider potential exposures due to all 
members of the group. When available, EPA recommends the use of 
existing modeling data to support the exposure analysis.
4. Tier 2 Summary
    For each study, the objectives, principles, and general procedures 
must be outlined and the findings and conclusions summarized. 
Discussion must be included regarding problems encountered during the 
performance of the tests and the methods used to resolve them. This 
discussion must include the impact which such problems may have had on 
the study outcomes.
5. Conclusions
    Further testing needs must be identified or else a discussion must 
be provided explaining why the results of Tiers 1 and/or Tier 2 should 
not trigger Tier 3 testing requirements.

C. Appendices

    Detailed information in support of the general discussions 
contained in the summary report are to be submitted as appendices to 
the report. In regard to the literature search, the appendices must 
contain (1) summary tables of existing studies regarding health and 
environmental effects, including such information as the type of study, 
species/strain used, exposure concentration(s), duration of study, 
endpoints evaluated, results (incidence and statistical significance), 
and references, (2) a complete copy of reference lists and associated 
abstracts obtained from data base searches (in printed form or on 3\1/
2\ inch (IBM compatible) computer diskettes), (3) complete 
documentation of in-house studies and other unpublished information 
sources, and (4) complete documentation (e.g., copies of journal 
articles) of previous studies which are being cited in satisfaction of 
Tier 1 and/or Tier 2 test requirements. Appendices to the emission 
characterization section must contain detailed protocols, copies of all 
relevant laboratory reports, a list of all speciated emission products 
and their emission rates, and documentation and results of calibration/
verification procedures. For the section that discusses potential 
exposures, an appendix must be provided for detailed background 
information on the production volume and market distribution data used 
in the exposure analysis. If exposure models are used, background 
calculations and/or model data must also be included in an appendix.
    An appendix is also required for each of the tests conducted in 
compliance with Tier 2 requirements. These appendices must contain the 
full detailed study protocol, complete laboratory report, statistical 
analysis of the findings, and scientific conclusions. These materials 
must conform to the reporting requirements of the individual study 
guidelines as well as the general standards for recordkeeping and 
reporting specified in the GLP standards of this final rule (see 
Sec. 79.60). A final appendix must be provided, containing laboratory 
certifications and associated personnel credentials.

D. Tier 3 Report

    Reports for additional tests required under the Tier 3 provisions 
must include a cover page with identifying information as described 
above for the Tier 1 and 2 summary report(s). The report must begin 
with a discussion of the concerns arising under the previous tiers 
which led to the Tier 3 requirements, the specific objectives of the 
additional studies, and a summary of pertinent results and conclusions. 
References used in support of Tier 3 conclusions must be cited in the 
report. The Tier 3 summary discussion must be supported with appendices 
containing the kinds of documentation discussed above with respect to 
Tier 2. The laboratory conducting the required tests must be 
identified, and relevant certifications and personnel credentials 
provided.

E. Confidential Business Information

    CAA section 211(b)(2)(B) states that the results of tests 
``conducted in conformity with test procedures and protocols 
established by the Administrator,'' pursuant to CAA section 
211(b)(2)(A), shall not be considered confidential. Thus, health and 
welfare information supplied to EPA in compliance with Tier 1, Tier 2, 
and Tier 3 testing requirements will be made available to the public.
    Manufacturers (or groups) claiming business confidentiality on any 
information submitted under the F/FA testing program must make a claim 
of confidentiality in writing at the time of submittal of the reporting 
requirements. To assert a business confidentiality claim the submitter 
must clearly mark the confidential information and must submit a 
separate document setting forth the claim and listing each location at 
which the confidential information appears in the submitted data. If 
any person subsequently requests access to the test data submitted 
under the F/FA testing program (other than health and welfare effects 
information) and such information is subject to a claim of business 
confidentiality, the request and any subsequent disclosure will be 
governed by the provisions of 40 CFR part 2.

XIII. Administrative Requirements

A. Administrative Designation and Regulatory Analysis

    Under Executive Order 12866 (58 FR 51735), EPA must determine 
whether the regulatory action is ``significant'' and therefore subject 
to the Office of Management and Budget (OMB) review and the 
requirements of the Executive Order. The Order defines ``economically 
significant regulatory action'' as one that is likely to result in a 
rule that may: (1) Have an annual effect on the economy of $100 million 
or more or adversely affect in a material way the economy, a sector of 
the economy, productivity, competition, jobs, the environment, public 
health or safety, or State, local, or tribal governments or 
communities; (2) create a serious inconsistency or otherwise interfere 
with an action taken or planned by another agency; (3) materially alter 
the budgetary impact of entitlements, grants, user fees, or loan 
programs or the rights and obligations of recipients thereof; or (4) 
raise novel legal or policy issues arising out of legal mandates, the 
President's priorities, or the principles set forth in the Executive 
Order.
    A regulatory support document which presents EPA's analysis of the 
cost impacts of this final rule is available for review in the public 
docket (A-90-07). EPA estimates that the costs to industry for 
submittal of the requisite data for Tiers 1 and 2 would total 
approximately $66 million, assumed to be incurred over the first three-
year period after promulgation of this final rule. Thus, the average 
annual cost during this period would be about $22 million. In the 
subsequent three years, Tier 3 requirements might cost an additional $1 
million annually, per product or group. If ten products or groups were 
required to conduct Tier 3 testing in the three-year period following 
the initial compliance period, the cost would be $10 million per year. 
These projected overall costs are far less than the $100 million annual 
cost criterion which is a major determinant in defining an 
``economically significant regulatory action.'' In addition, this final 
rule is not expected to adversely affect in a material way the economy, 
a sector of the economy, productivity, competition, jobs, the 
environment, public health or safety, or State, local, or tribal 
governments or communities.
    Although not ``economically significant'' based on the above 
criteria, this final rule is still considered a ``significant 
regulatory action'' pursuant to the terms of Executive Order 12866 and 
was thus submitted to OMB for review. Any written comments from OMB and 
any EPA response to OMB's comments are available in the public docket 
for this rule.

B. Regulatory Flexibility Act

    Under section 605 of the Regulatory Flexibility Act, 5 U.S.C. 601 
et seq., the Administrator is required to assess the economic impact of 
regulatory actions on small businesses. Accordingly, a Regulatory 
Flexibility Analysis (RFA) has been prepared for this rule and is 
presented as part of the Regulatory Support Document (available in the 
public docket). The RFA compares the financial impacts of this rule on 
small F/FA manufacturers to the impacts on large F/FA manufacturers. 
The analysis explains the small business definition specifically 
developed in this rule to provide special provisions for small F/FA 
manufacturers (see Section X.D) and assesses the effectiveness of these 
provisions.
    This final rule defines a small business as any motor vehicle fuel 
or fuel additive manufacturer with total annual sales of less than $50 
million. A comparison of companies classified as small under this 
definition to those classified as small by the Small Business 
Administration (SBA) reveals that the F/FA program definition 
classifies a larger number of F/FA manufacturers as small, and more 
closely groups companies with similar financial characteristics. Based 
on this analysis, EPA determined that the small business definition 
established in this rule is reasonable and applicable to the F/FA 
industry.
    Impacts of this rule on F/FA manufacturers were determined by 
projecting the effects of the estimated compliance costs on each 
company's return on assets (ROA). In general, a reduction in ROA (after 
compliance costs) to less than 2.5 percent is indicative of financial 
distress. A ROA less than -4 percent indicates that a company is in 
severe financial distress, and a ROA less than -30 percent generally 
indicates closure. According to these ROA thresholds, results of the 
RFA show that approximately 68 unique F/FA manufacturers (43 additive 
manufacturers, 23 fuel manufacturers, and 2 that produce both fuels and 
additives) could potentially be pushed into some level of financial 
distress. All of these companies are classified as small (i.e., have 
sales less than $50 million). The majority of these companies would 
fall into the first level of financial distress; however, less than 
one-fifth of them (12 companies) would potentially be in danger of 
closure. In relation to the total population of F/FA manufacturers, the 
companies potentially falling into some level of financial distress 
account to about six percent, with about 1.2 percent potentially in 
danger of closure.
    As previously described, this final rule includes two types of 
special provisions for small businesses. With respect to baseline and 
non-baseline F/FAs, all small manufacturers (i.e., annual sales under 
$50 million) are excused from all Tier 1 and Tier 2 testing 
requirements. A segment of these small manufacturers, i.e., those 
having sales under $10 million, are also excused from Tier 2 testing 
responsibility for their atypical F/FAs. Nevertheless, all twelve of 
the manufacturers who are projected to be in danger of closure as a 
result of this rule are very small companies with one or more atypical 
F/FAs. The RFA analysis shows that the special provision for small 
manufacturers with atypical products lowers overall compliance costs 
for 60 such manufacturers by roughly $20 million (as compared with 
costs that would otherwise occur in the absence of this special 
provision). Without this provision, an additional 15 manufacturers of 
atypical products would potentially be pushed into closure.
    It should be noted that the RFA has assumed no changes in prices, 
sales, product mix, or financial strategies. In many cases, a portion 
of regulatory costs can be actually passed on to consumers or back to 
suppliers. Manufacturers also have the option of reformulating a 
product to a ``baseline'' standard which has much lower compliance 
costs, or even of dropping products with the greatest cost and smallest 
profit potential. A more detailed discussion on circumstances which 
could mitigate compliance cost impacts is provided in the RFA.

C. Recordkeeping Requirements

    The information collection requirements in this rule have been 
submitted for approval to OMB under the Paperwork Reduction Act, 44 
U.S.C. 3501 et seq. An Information Collection Request document has been 
prepared by EPA (ICR #1696.01) and a copy may be obtained from Sandy 
Farmer, Information Policy Branch, EPA, 401 M Street SW., Washington, 
DC 20460 or by calling 202-260-2740. These requirements are not 
effective until OMB approves them and a technical amendment to that 
effect is published in the Federal Register.
    This collection of information has an estimated annual reporting 
and recordkeeping burden averaging 43 hours per response. These 
estimates include time for reviewing instructions, searching existing 
data sources, gathering and maintaining the data needed, and completing 
and reviewing the collection of information.
    Send comments regarding the burden estimate or any other aspect of 
this collection of information, including suggestions for reducing this 
burden to Chief, Information Policy Branch; EPA; 401 M Street, SW., 
(Mail Code 2136); Washington, DC 20460; and to the Office of 
Information and Regulatory Affairs, Office of Management and Budget, 
Washington, DC 20503, marked ``Attention: Desk Officer for EPA.''

XIV. Electronic Availability of Rulemaking Documents

    Electronic copies of the preamble, the Regulatory Support Document 
and Regulatory Flexibility Analysis, the Summary and Analysis of 
Comments, and the regulations for the Fuels and Fuel Additives 
Registration rulemaking are available on the Office of Air Quality 
Planning and Standards (OAQPS) Technology Transfer Network Bulletin 
Board System (TTNBBS). Instructions for accessing TTNBBS and 
downloading F/FA files are described below.
    TTNBBS can be accessed using a dial-in telephone line (919-541-
5742) and a 1200, 2400, or 9600 bps modem (equipment up to 14.4 Kbps 
can be accommodated). The parity of the modem should be set to N or 
none, the data bits to 8, and the stop bits to 1. When first signing on 
to the bulletin board, the user will be required to answer some basic 
informational questions to register into the system. After registering, 
proceed through the following options from a series of menus:

(M) OMS
(K) Rulemaking and Reporting
(3) Fuels
(4) Fuels/Fuel Additives

    A list of ``.ZIP'' files will be displayed, all of which relate to 
the Fuels and Fuel Additives Registration rulemaking. The four 
documents listed will be in the form of ``.ZIP'' files and are 
identified by the following titles:

``FFA--PRE.ZIP'' (Preamble)
``FFA--RSD.ZIP'' (Regulatory Support Document and Regulatory 
Flexibility Analysis)
``FFA--COM.ZIP'' (Summary and Analysis of Comments)
``FFA--REG.ZIP'' (Regulations)

    File information can be obtained from the ``READ.ME'' file. Choose 
from the following options when prompted:

ownload, rotocol, xamine, ew, ist, elp or  to 
exit.

    To download a file, e.g.,  filename.ZIP, the user needs to 
choose a file transfer protocol appropriate for the user's computer 
from the options listed on the terminal. The user's computer is then 
ready to receive the file by invoking the user's resident file transfer 
software. Programs and instructions for de-archiving compressed files 
can be found under ystems Utilities from the top menu, under 
rchivers/de-archivers.
    TTNBBS is available 24 hours a day, 7 days a week except Monday 
morning from 8-12 EST, when the system is down for maintenance and 
backup. For help in accessing the system, call the systems operator at 
919-541-5384 in Research Triangle Park, North Carolina, during normal 
business hours EST.

List of Subjects in 40 CFR Part 79

    Environmental protection, Fuel additives, Fuels, Gasoline, 
Incorporation by reference, Motor vehicle pollution, Penalties, 
Reporting and recordkeeping requirements.

    Dated: May 27, 1994.
Carol M. Browner,
Administrator.

    Part 79 of title 40 of the Code of Federal Regulations is amended 
as follows:
    1. The authority citation for part 79 is revised to read as 
follows:

    Authority: 42 U.S.C. 7414, 7524, 7545 and 7601.

    2. Section 79.2 is amended by revising paragraphs (d), (e), and (f) 
to read as follows:


Sec. 79.2  Definitions.

* * * * *
    (d) Fuel manufacturer means any person who, for sale or 
introduction into commerce, produces, manufactures, or imports a fuel 
or causes or directs the alteration of the chemical composition of, or 
the mixture of chemical compounds in, a bulk fuel by adding to it an 
additive.
    (e) Additive means any substance that is intentionally added to a 
fuel named in the designation (including any added to a motor vehicle's 
fuel system) and that is not intentionally removed prior to sale or 
use.
    (f) Additive manufacturer means any person who produces, 
manufactures, or imports an additive for use as an additive and/or 
sells or imports for sale such additive under the person's own name.
* * * * *
    3. Section 79.3 is revised to read as follows:


Sec. 79.3  Availability of information.

    The availability to the public of information provided to, or 
otherwise obtained by, the Administrator under this part shall be 
governed by part 2 of this chapter except as expressly noted in subpart 
F of this part.
    4. Section 79.4 is amended by revising paragraph (b)(1) to read as 
follows:


Sec. 79.4  Requirement of registration.

* * * * *
    (b) Additives. (1) No manufacturer of any fuel additive designated 
under this part shall, after the date by which the additive must be 
registered under this part, sell, offer for sale, or introduce into 
commerce such additive for use in any type of fuel designated under 
this part unless the Administrator has registered that additive for use 
in that type of fuel.
* * * * *
    5. Section 79.6 is revised to read as follows:


Sec. 79.6  Requirement for testing.

    Provisions regarding testing that is required for registration of a 
designated fuel or fuel additive are contained in subpart F of this 
part.
    6. Section 79.10 is revised to read as follows:


Sec. 79.10  Application for registration by fuel manufacturer.

    Any manufacturer of a designated fuel who wishes to register that 
fuel shall submit an application for registration including all of the 
information set forth in Sec. 79.11. If the manufacturer produces more 
than one grade or brand of a designated fuel, a manufacturer may 
include more than one grade or brand in a single application, provided 
that the application includes all information required for registration 
of each such grade or brand by this part. Each application shall be 
signed by the fuel manufacturer and shall be submitted on such forms as 
the Administrator will supply on request.
    7. Section 79.11 is amended by revising the introductory text of 
the section, removing the period in paragraph (h) and adding a 
semicolon and paragraphs (i) and (j) to read as follows:


Sec. 79.11  Information and assurances to be provided by the fuel 
manufacturer.

    Each application for registration submitted by the manufacturer of 
a designated fuel shall include the following:
* * * * *
    (i) The manufacturer of any fuel which will be sold, offered for 
sale, or introduced into commerce for use in motor vehicles 
manufactured after model year 1974 shall demonstrate that the fuel is 
substantially similar to any fuel utilized in the certification of any 
1975 or subsequent model year vehicle or engine, or that the 
manufacturer has obtained a waiver under 42 U.S.C. 7545(f)(4); and
    (j) The manufacturer shall submit, or shall reference prior 
submissions, including all of the test data and other information 
required prior to registration of the fuel by the provisions of subpart 
F of this part.
    8. Section 79.12 is revised to read as follows:


Sec. 79.12  Determination of noncompliance.

    If the Administrator determines that an applicant for registration 
of a designated fuel has failed to submit all of the information 
required by Sec. 79.11, or determines within the applicable period 
provided for Agency review that the applicant has not satisfactorily 
completed any testing which is required prior to registration of the 
fuel by any provision of subpart F of this part, he shall return the 
application to the manufacturer, along with an explanation of all 
deficiencies in the required information.
    9. Section 79.13 is amended by revising paragraph (a) to read as 
follows:


Sec. 79.13  Registration.

    (a) If the Administrator determines that a manufacturer has 
submitted an application for registration of a designated fuel which 
includes all of the information and assurances required by Sec. 79.11 
and has satisfactorily completed all of the testing required by subpart 
F of this part, the Administrator shall promptly register the fuel and 
notify the fuel manufacturer of such registration.
* * * * *
    10. Section 79.20 is revised to read as follows:


Sec. 79.20  Application for registration by additive manufacturer.

    Any manufacturer of a designated fuel additive who wishes to 
register that additive shall submit an application for registration 
including all of the information set forth in Sec. 79.21. Each 
application shall be signed by the fuel additive manufacturer and shall 
be submitted on such forms as the Administrator will supply on request.
    11. Section 79.21 is amended by revising the introductory text of 
the section and paragraph (d) and adding paragraphs (h) and (i) to read 
as follows:


Sec. 79.21  Information and assurances to be provided by the additive 
manufacturer.

    Each application for registration submitted by the manufacturer of 
a designated fuel additive shall include the following:
* * * * *
    (d) The specific types of fuels designated under Sec. 79.32 for 
which the fuel additive will be sold, offered for sale, or introduced 
into commerce, and the fuel additive manufacturer's recommended range 
of concentration and purpose-in-use for each such type of fuel.
* * * * *
    (h) The manufacturer of any fuel additive which will be sold, 
offered for sale, or introduced into commerce for use in any type of 
fuel intended for use in motor vehicles manufactured after model year 
1974 shall demonstrate that the fuel additive, when used at the 
recommended range of concentration, is substantially similar to any 
fuel additive included in a fuel utilized in the certification of any 
1975 or subsequent model year vehicle or engine, or that the 
manufacturer has obtained a waiver under 42 U.S.C. 7545(f)(4).
    (i) The manufacturer shall submit, or shall reference prior 
submissions, including all of the test data and other information 
required prior to registration of the fuel additive by the provisions 
of subpart F of this part.
    12. Section 79.22 is revised to read as follows:


Sec. 79.22  Determination of noncompliance.

    If the Administrator determines that an applicant for registration 
of a designated fuel additive has failed to submit all of the 
information required by Sec. 79.21, or determines within the applicable 
period provided for Agency review that the applicant has not 
satisfactorily completed any testing which is required prior to 
registration of the fuel additive by any provision of subpart F of this 
part, he shall return the application to the manufacturer, along with 
an explanation of all deficiencies in the required information.
    13. Section 79.23 is amended by removing paragraph (b), by 
redesignating paragraph (c) as paragraph (b), and by revising paragraph 
(a) to read as follows:


Sec. 79.23  Registration.

    (a) If the Administrator determines that a manufacturer has 
submitted an application for registration of a designated fuel additive 
which includes all of the information and assurances required by 
Sec. 79.21 and has satisfactorily completed all of the testing required 
by subpart F of this part, the Administrator shall promptly register 
the fuel additive and notify the fuel manufacturer of such 
registration.
* * * * *
    14. Section 79.31 is amended by revising paragraph (b) to read as 
follows:


Sec. 79.31  Additives.

* * * * *
    (b) All designated additives must be registered by July 7, 1976.
* * * * *
    15. A new subpart F, consisting of Secs. 79.50-79.68, is added to 
part 79 to read as follows:

Subpart F--Testing Requirements for Registration

Sec.
79.50  Definitions.
79.51  General requirements and provisions.
79.52  Tier 1.
79.53  Tier 2.
79.54  Tier 3.
79.55  Base fuel specifications.
79.56  Fuel and fuel additive grouping system.
79.57  Emission generation.
79.58  Special provisions.
79.59  Reporting requirements.
79.60  Good laboratory practice (GLP) standards for inhalation 
exposure health effects testing.
79.61  Vehicle emissions inhalation exposure guideline.
79.62  Subchronic toxicity study with specific health effect 
assessments.
79.63  Fertility assessment/teratology.
79.64  In vivo micronucleus assay.
79.65  In vivo sister chromatid exchange assay.
79.66  Neuropathology assessment.
79.67  Glial fibrillary acidic protein assay.
79.68  Salmonella typhimurium reverse mutation assay.

Subpart F--Testing Requirements for Registration


Sec. 79.50  Definitions.

    The definitions listed in this section apply only to subpart F of 
this part.
    Additive/base fuel mixture means the mixture resulting when a fuel 
additive is added in specified proportion to the base fuel of the fuel 
family to which the additive belongs.
    Aerosol additive means a chemical mixture in aerosol form generally 
used as a motor vehicle engine starting aid or carburetor cleaner and 
not recommended to be placed in the fuel tank.
    Aftermarket fuel additive means a product which is added by the 
end-user directly to fuel in a motor vehicle or engine to modify the 
performance or other characteristics of the fuel, the engine, or its 
emissions.
    Atypical element means any chemical element found in a fuel or 
additive product which is not allowed in the baseline category of the 
associated fuel family, and an ``atypical fuel or fuel additive'' is a 
product which contains such an atypical element.
    Base fuel means a generic fuel formulated from a set of 
specifications to be representative of a particular fuel family.
    Basic emissions means the total hydrocarbons, carbon monoxide, 
oxides of nitrogen, and particulates occurring in motor vehicle or 
engine emissions.
    Bulk fuel additive means a product which is added to fuel at the 
refinery as part of the original blending stream or after the fuel is 
transported from the refinery but before the fuel is purchased for 
introduction into the fuel tank of a motor vehicle.
    Emission characterization means the determination of the chemical 
composition of emissions.
    Emission generation means the operation of a vehicle or engine or 
the vaporization of a fuel or additive/fuel mixture under controlled 
conditions for the purpose of creating emissions to be used for testing 
purposes.
    Emission sampling means the removal of a fraction of collected 
emissions for testing purposes.
    Emission speciation means the analysis of vehicle or engine 
emissions to determine the individual chemical compounds which comprise 
those emissions.
    Engine Dynamometer Schedule (EDS) means the transient engine speed 
versus torque time sequence commonly used in heavy-duty engine 
evaluation. The EDS for heavy-duty diesel engines is specified in 40 
CFR part 86, appendix I(f)(2).
    Evaporative Emission Generator (EEG) means a fuel tank or vessel to 
which heat is applied to cause a portion of the fuel to evaporate at a 
desired rate.
    Evaporative emissions means chemical compounds emitted into the 
atmosphere by vaporization of contents of a fuel or additive/fuel 
mixture.
    Evaporative fuel means a fuel which has a Reid Vapor Pressure (RVP, 
pursuant to 40 CFR part 80, appendix ``E'') of 2.0 pounds per square 
inch or greater and is not supplied to motor vehicle engines by way of 
sealed containment and delivery systems.
    Evaporative fuel additive means a fuel additive which, when mixed 
with its specified base fuel, causes an increase in the RVP of the base 
fuel by 0.4 psi or more relative to the RVP of the base fuel alone and 
results in an additive/base fuel mixture whose RVP is 2.0 psi, or 
greater. Excluded from this definition are fuel additives used with 
fuels which are supplied to motor vehicle engines by way of sealed 
containment and delivery systems.
    Federal Test Procedure (FTP) means the body of exhaust and 
evaporative emissions test procedures described in 40 CFR 86 for the 
certification of new motor vehicles to Federal motor vehicle emissions 
standards.
    Fuel family means a set of fuels and fuel additives which share 
basic chemical and physical formulation characteristics and can be used 
in the same engine or vehicle.
    Manufacturer means a person who is a fuel manufacturer or additive 
manufacturer as defined in Sec. 79.2 (d) and (f).
    Nitrated polycyclic aromatic hydrocarbons (NPAH) means the class of 
compounds whose molecular structure includes two or more aromatic rings 
and contains one or more nitrogen substitutions.
    Non-catalyzed emissions means exhaust emissions not subject to an 
effective aftertreatment device such as a functional catalyst or 
particulate trap.
    Oxygenate compound means an oxygen-containing, ashless organic 
compound, such as an alcohol or ether, which may be used as a fuel or 
fuel additive.
    Polycyclic aromatic hydrocarbons (PAH) means the class of 
hydrocarbon compounds whose molecular structure includes two or more 
aromatic rings.
    Relabeled additive means a fuel additive which is registered by its 
original manufacturer with EPA and is also registered and sold, 
unchanged in composition, under a different label and/or by a different 
entity.
    Semi-volatile organic compounds means that fraction of gaseous 
combustion emissions which consists of compounds with greater than 
twelve carbon atoms and can be trapped in sorbent polymer resins.
    Urban Dynamometer Driving Schedule (UDDS) means the 1372 second 
transient speed driving sequence used by EPA to simulate typical urban 
driving. The UDDS for light-duty vehicles is described in 40 CFR part 
86, appendix I(a).
    Vapor phase means the gaseous fraction of combustion emissions.
    Vehicle classes/subclasses means the divisions of vehicle groups 
within a vehicle type, including light-duty vehicles, light-duty 
trucks, and heavy-duty vehicles as specified in 40 CFR part 86.
    Vehicle type means the divisions of motor vehicles according to 
combustion cycle and intended fuel class, including, but not 
necessarily limited to, Otto cycle gasoline-fueled vehicles, Otto cycle 
methanol-fueled vehicles, diesel cycle diesel-fueled vehicles, and 
diesel cycle methanol-fueled vehicles.
    Whole emissions means all components of unfiltered combustion 
emissions or evaporative emissions.


Sec. 79.51  General requirements and provisions.

    (a) Overview of requirements. (1) All manufacturers of fuels and 
fuel additives that are designated for registration under this part are 
required to comply with the requirements of subpart F of this part 
either on an individual basis or as a participant in a group of 
manufacturers of the same or similar fuels and fuel additives, as 
defined in Sec. 79.56. If manufacturers elect to comply by 
participation in a group, each manufacturer continues to be 
individually subject to the requirements of subpart F of this part, and 
responsible for testing under this subpart. Each manufacturer, subject 
to the provisions for group applications in Sec. 79.51(b) and the 
special provisions in Sec. 79.58, shall submit all Tier 1 and Tier 2 
information required by Secs. 79.52, 79.53 and 79.59 for each fuel or 
additive, except that the Tier 1 emission characterization requirements 
in Sec. 79.52(b) and/or the Tier 2 testing requirements in Sec. 79.53 
may be satisfied by adequate existing information pursuant to the Tier 
1 literature search requirements in Sec. 79.52(d). The adequacy of 
existing information to serve in compliance with specific Tier 1 and/or 
Tier 2 requirements shall be determined according to the criteria and 
procedures specified in Secs. 79.52(b) and 79.53 (c) and (d). In all 
cases, EPA reserves the right to require, based upon the information 
contained in the application or any other information available to the 
Agency, that manufacturers conduct additional testing of any fuel or 
additive (or fuel/additive group) if EPA determines that there is 
inadequate information upon which to base regulatory decisions for such 
product(s). In any case where EPA determines that the requirements of 
Tiers 1 and 2 have been satisfied but that further testing is required, 
the provisions of Tier 3 (Sec. 79.54) shall apply.
    (2) Laboratory facilities shall perform testing in compliance with 
Good Laboratory Practice (GLP) requirements as those requirements apply 
to inhalation toxicology studies. All studies shall be monitored by the 
facilities' Quality Assurance units (as specified in Sec. 79.60).
    (b) Group Applications. Subject to the provisions of Sec. 79.56 (a) 
through (c), EPA will consider any testing requirements of this subpart 
to have been met for any fuel or fuel additive when a fuel or fuel 
additive which meets the criteria for inclusion in the same group as 
the subject fuel or fuel additive has met that testing requirement, 
provided that all fuels and additives must be individually registered 
as described in Sec. 79.59(b). For purposes of this subpart, a 
determination of which group contains a particular fuel or additive 
will be made pursuant to the provisions of Sec. 79.56 (d) and (e). 
Nothing in this subsection (b) shall be deemed to require a 
manufacturer to rely on another manufacturer's testing.
    (c) Application Procedures and Dates. Each application submitted in 
compliance with this subpart shall be signed by the manufacturer of the 
designated fuel or additive, or by the manufacturer's agent, and shall 
be submitted to the address and in the format prescribed in Sec. 79.59. 
A manufacturer who chooses to comply as part of a group pursuant to 
Sec. 79.56 shall be covered by the group's joint application. Subject 
to any modifications pursuant to the special provisions in 
Secs. 79.51(f) or 79.58, the schedule for compliance with the 
requirements of this subpart is as follows:
    (1) Fuels and fuel additives with existing registrations. (i) The 
manufacturer of a fuel or fuel additive product which, pursuant to 
subpart B or C of this part, is registered as of May 27, 1994 must 
submit the additional basic registration data specified in 
Sec. 79.59(b) before November 28, 1994.
    (ii) For these products, the manufacturer must also satisfy the 
requirements and time schedules in either of the following paragraphs 
(c)(1)(ii) (A) or (B) of this section:
    (A) Within May 27, 1997, all applicable Tier 1 and Tier 2 
requirements must be submitted to EPA, pursuant to Secs. 79.52, 79.53, 
and 79.59; or
    (B) Within May 27, 1997, all applicable Tier 1 requirements 
(pursuant to Secs. 79.52 and 79.59), plus evidence of a contract with a 
qualified laboratory (or other suitable arrangement) for completion of 
all applicable Tier 2 requirements, must be submitted to EPA. For this 
purpose, a qualified laboratory is one which can demonstrate the 
capabilities and credentials specified in Sec. 79.53(c)(1). In 
addition, within May 26, 2000, all applicable Tier 2 requirements 
(pursuant to Secs. 79.53 and 79.59) must be submitted to EPA.
    (iii) In the case of such fuels and fuel additives which, pursuant 
to applicable special provisions in Sec. 79.58, are not subject to Tier 
2 requirements, all other requirements (except Tier 3) must be 
submitted to EPA before May 27, 1997.
    (iv) In the event that Tier 3 testing is also required (under 
Sec. 79.54), EPA shall determine an appropriate timeline for completion 
of the additional requirements and shall communicate this schedule to 
the manufacturer according to the provisions of Sec. 79.54(b).
    (v) The manufacturer may at any time modify an existing fuel 
registration by submitting a request to EPA to add or delete a bulk 
additive to the existing registration information for such fuel 
product, provided that any additional additive must be registered by 
EPA for use in the specific fuel family to which the fuel product 
belongs. However, the addition or deletion of a bulk additive to a fuel 
registration may effect the grouping of such registered fuel under the 
criteria of Sec. 79.56, and thus may effect the testing 
responsibilities of the fuel manufacturer under this subpart.
    (2) Registrable fuels and fuel additives. (i) A fuel product which 
is not registered pursuant to subpart B of this part as of May 27, 1994 
shall be considered registrable if, under the criteria established by 
Sec. 79.56, the fuel can be enrolled in the same fuel/additive group 
with one or more currently registered fuels. A fuel additive product 
which is not registered for a specific type of fuel pursuant to subpart 
C of this part as of May 27, 1994 shall be considered registrable for 
that type of fuel if, under the criteria established by Sec. 79.56, the 
fuel/additive mixture resulting from use of the additive product in the 
specific type of fuel can be enrolled in the same fuel/additive group 
with one or more currently registered fuels or bulk fuel additives. For 
the purpose of this determination, currently registered fuels and bulk 
additives are those with existing registrations as of the date on which 
EPA receives the basic registration data (pursuant to Sec. 79.59(b)) 
for the product in question.
    (ii) A manufacturer seeking to register under subpart B of this 
part a fuel product which is deemed registrable under this section, or 
to register under subpart C of this part a fuel additive product for a 
specific type of fuel for which it is deemed registrable under this 
section, shall submit the basic registration data (pursuant to 
Sec. 79.59(b)) for that product as part of the application for 
registration. If the Administrator determines that the product is 
registrable under this section, then the Administrator shall promptly 
register the product, provided that the applicant has satisfied all of 
the other requirements for registration under subpart B or subpart C of 
this part, and contingent upon satisfactory submission of required 
information under paragraph (c)(2)(iii) of this section.
    (iii) Registration of a registrable fuel or additive shall be 
subject to the same requirements and compliance schedule as specified 
in paragraph (c)(1) of this section for existing fuels and fuel 
additives. Accordingly, manufacturers of registrable fuels or additives 
may be granted and may retain registration for such products only if 
any applicable and due Tier 1, 2, and 3 requirements have also been 
satisfied by either the manufacturer of the product or the fuel/
additive group to which the product belongs.
    (3) New fuels and fuel additives. A fuel product shall be 
considered new if it is not registered pursuant to subpart B of this 
part as of May 27, 1994 and if, under the criteria established by 
Sec. 79.56, it cannot be enrolled in the same fuel/additive group with 
one or more currently registered fuels. A fuel additive product shall 
be considered new with respect to a specific type of fuel if it is not 
expressly registered for that type of fuel pursuant to subpart C of 
this part as of May 27, 1994 and if, under the criteria established by 
Sec. 79.56, the fuel/additive mixture resulting from use of the 
additive product in the specific type of fuel cannot be enrolled in the 
same fuel/additive group with one or more currently registered fuels or 
bulk fuel additives. For the purpose of this determination, currently 
registered fuels and bulk additives are those with existing 
registrations as of the date on which EPA receives the basic 
registration data (pursuant to Sec. 79.59(b)) for the product in 
question. For such new product, the manufacturer must satisfactorily 
complete all applicable Tier 1 and Tier 2 requirements, followed by any 
Tier 3 testing which the Administrator may require, before registration 
will be granted.
    (d) Notifications. Upon receipt of a manufacturer's (or group's) 
submittal in compliance with the requirements of this subpart, EPA will 
notify such manufacturer (or group) that the application has been 
received and what, if any, information, testing, or retesting is 
necessary to bring the application into compliance with the 
requirements of this subpart. EPA intends to provide such notification 
of receipt in a timely manner for each such application.
    (1) Registered fuel and fuel additive notification. (i) The 
manufacturer of a registered fuel or fuel additive product who is 
notified that the submittal for such product contains adequate 
information pursuant to the Tier 1 and Tier 2 testing and reporting 
requirements (Secs. 79.52, 79.53, and 79.59 (a) through (c)) may 
continue to sell, offer for sale, or introduce into commerce the 
registered product as permitted by the existing registration for the 
product under Sec. 79.4.
    (ii) If the manufacturer of a registered fuel or fuel additive 
product is notified that testing or retesting is necessary to bring the 
Tier 1 and/or Tier 2 submittal into compliance, the continued sale or 
importation of the product shall be conditional upon satisfactorily 
completing the requirements within the time frame specified in 
paragraph (c)(1) of this section.
    (iii) EPA intends to notify the manufacturer of the adequacy of the 
submitted data within two years of EPA's receipt of such data. However, 
EPA retains the right to require that adequate data be submitted to EPA 
if, upon subsequent review, EPA finds that the original Tier 1 and/or 
Tier 2 submittal is not consistent with the requirements of this 
subpart. If EPA does not notify the manufacturer of the adequacy of the 
Tier 1 and/or Tier 2 data within two years, EPA will not hold the 
manufacturer liable for penalties for violating this rule for the 
period beginning when the data was due until the time EPA notifies the 
manufacturer of the violation.
    (iv) If the manufacturer of a registered fuel or fuel additive 
product is notified (pursuant to Sec. 79.54(b)) that Tier 3 testing is 
required for its product, then the manufacturer may continue to sell, 
offer for sale, introduce into commerce the registered product as 
permitted by the existing registration for the product under Sec. 79.4. 
However, if the manufacturer fails to complete the specified Tier 3 
requirements within the specified time, the registration of the product 
will be subject to cancellation under Sec. 79.51(f)(6).
    (v) EPA retains the right to require additional Tier 3 testing 
pursuant to the procedures in Sec. 79.54.
    (2) New fuel and fuel additive notification. (i) Within six months 
following its receipt of the Tier 1 and Tier 2 submittal for a new 
product (as defined in paragraph (c)(3) of this section), EPA shall 
notify the manufacturer of the adequacy of such submittal in compliance 
with the requirements of Secs. 79.52, 79.53, and 79.59 (a) through (c).
    (A) If EPA notifies the manufacturer that testing, retesting, or 
additional information is necessary to bring the Tier 1 and Tier 2 
submittal into compliance, the manufacturer shall remedy all 
inadequacies and provide Tier 3 data, if required, before EPA shall 
consider the requirements for registration to have been met for the 
product in question.
    (B) If EPA does not notify the manufacturer of the adequacy of the 
Tier 1 and Tier 2 submittal within six months following the submittal, 
the manufacturer shall be deemed to have satisfactorily completed Tiers 
1 and 2.
    (ii) Within six months of the date on which EPA notifies the 
manufacturer of satisfactory completion of Tiers 1 and 2 for a new 
product, or within one year of the submittal of the Tier 1 and Tier 2 
data (whichever is earlier), EPA shall determine whether additional 
testing is currently needed under the provisions of Tier 3 and, 
pursuant to Sec. 79.54(b), shall notify the manufacturer of its 
determination.
    (A) If the manufacturer of a new fuel or fuel additive product is 
notified that Tier 3 testing is required for such product, then EPA 
shall have the authority to withhold registration until the specified 
Tier 3 requirements have been satisfactorily completed. EPA shall 
determine whether the Tier 3 requirements have been met, and shall 
notify the manufacturer of this determination, within one year of 
receiving the manufacturer's Tier 3 submittal.
    (B) If EPA does not notify the manufacturer of potential Tier 3 
requirements within the prescribed timeframe, then additional testing 
at the Tier 3 level is deemed currently unnecessary and the 
manufacturer shall be considered to have complied with all current 
registration requirements for the new fuel or additive product.
    (iii) Upon completion of all current Tier 1, Tier 2, and Tier 3 
requirements, and submission of an application for registration which 
includes all of the information and assurances required by Sec. 79.11 
or Sec. 79.21, the registration of the new fuel or additive shall be 
granted, and the registrant may then sell, offer for sale, or introduce 
into commerce the registered product as permitted by Sec. 79.4.
    (iv) Once the new product becomes registered, EPA reserves the 
right to require additional Tier 3 testing pursuant to the procedures 
specified in Sec. 79.54.
    (e) Inspection of a testing facility. (1) A testing facility, 
emissions analysis or health and/or welfare effects, shall permit an 
authorized employee or duly designated representative of EPA, at 
reasonable times and in a reasonable manner, to inspect the facility 
and to inspect (and in the case of records also to copy) all records 
and specimens required to be maintained regarding studies to which this 
rule applies. The records inspection and copying requirements shall not 
apply to quality assurance unit records of findings and problems, or to 
actions recommended and taken, except the EPA may seek production of 
these records in litigation or informal hearings.
    (2) EPA will not consider reliable for purposes of showing that a 
test substance does or does not present a risk of injury to health or 
the environment any data developed by a testing facility or sponsor 
that refuses to permit inspection in accordance with this section. The 
determination that a study will not be considered reliable does not, 
however, relieve the sponsor of a required test of any obligation under 
any applicable statute or regulation to submit the results of the study 
to EPA.
    (3) Effects of non-compliance. Pursuant to sections 114, 208, and 
211(d) of the CAA, it shall be a violation of this section and a 
violation of 40 CFR part 79, subpart F to deny entry to an authorized 
employee or duly designated representative of EPA for the purpose of 
auditing a testing facility or test data.
    (f) Penalties and Injunctive Relief. (1) Any person who violates 
these regulations shall be subject to a civil penalty of up to $25,000 
for each and every day of the continuance of the violation and the 
economic benefit or savings resulting from the violation. Action to 
collect such civil penalties shall be commenced in accordance with 
paragraph (b) of section 205 of the Clean Air Act or assessed in 
accordance with paragraph (c) of section 205 of the Clean Air Act, 42 
U.S.C. 7524 (b) and (c).
    (2) Under section 205(b) of the CAA, the Administrator may commence 
a civil action for violation of this subpart in the district court of 
the United States for the district in which the violation is alleged to 
have occurred or in which the defendant resides or has a principal 
place of business.
    (3) Under section 205(c) of the CAA, the Administrator may assess a 
civil penalty of $25,000 for each and every day of the continuance of 
the violation and the economic benefit or savings resulting from the 
violation, except that the maximum penalty assessment shall not exceed 
$200,000, unless the Administrator and the Attorney General jointly 
determine that a matter involving a larger penalty amount is 
appropriate for administrative penalty assessment. Any such 
determination by the Administrator and the Attorney General shall not 
be subject to judicial review.
    (4) The Administrator may, upon application by the person against 
whom any such penalty has been assessed, remit or mitigate, with or 
without conditions, any such penalty.
    (5) The district courts of the United States shall have 
jurisdiction to compel the furnishing of information and the conduct of 
tests required by the Administrator under these regulations and to 
award other appropriate relief. Actions to compel such actions shall be 
brought by and in the name of the United States. In any such action, 
subpoenas for witnesses who are required to attend a district court in 
any district may run into any other district.
    (6) Cancellation.
    (i) The Administrator of EPA may issue a notice of intent to cancel 
a fuel or fuel additive registration if the Administrator determines 
that the registrant has failed to submit in a timely manner any data 
required to maintain registration under this part or under section 
211(b) or 211(e) of the Clean Air Act.
    (ii) Upon issuance of a notice of intent to cancel, EPA will 
forward a copy of the notice to the registrant by certified mail, 
return receipt requested, at the address of record given in the 
registration, along with an explanation of the reasons for the proposed 
cancellation.
    (iii) The registrant will be afforded 60 days from the date of 
receipt of the notice of intent to cancel to submit written comments 
concerning the notice, and to demonstrate or achieve compliance with 
the specific data requirements which provide the basis for the proposed 
cancellation. If the registrant does not respond in writing within 60 
days from the date of receipt of the notice of intent to cancel, the 
cancellation of the registration shall become final by operation of law 
and the Administrator shall notify the registrant of such cancellation. 
If the registrant responds in writing within 60 days from the date of 
receipt of the notice of intent to cancel, the Administrator shall 
review and consider all comments submitted by the registrant before 
taking final action concerning the proposed cancellation. The 
registrants' communications should be sent to the following address: 
Director, Field Operations and Support Division, 6406J--Fuel/Additives 
Registration, U.S. Environmental Protection Agency, 401 M Street SW., 
Washington, DC 20460.
    (iv) As part of a written response to a notice of intent to cancel, 
a registrant may request an informal hearing concerning the notice. Any 
such request shall state with specificity the information the 
registrant wishes to present at such a hearing. If an informal hearing 
is requested, EPA shall schedule such a hearing within 60 days from the 
date of receipt of the request. If an informal hearing is held, the 
subject matter of the hearing shall be confined solely to whether or 
not the registrant has complied with the specific data requirements 
which provide the basis for the proposed cancellation. If an informal 
hearing is held, the designated presiding officer may be any EPA 
employee, the hearing procedures shall be informal, and the hearing 
shall not be subject to or governed by 40 CFR part 22 or by 5 U.S.C. 
554, 556, or 557. A verbatim transcript of each informal hearing shall 
be kept and the Administrator shall consider all relevant evidence and 
arguments presented at the hearing in making a final decision 
concerning a proposed cancellation.
    (v) If a registrant who has received a notice of intent to cancel 
submits a timely written response, and the Administrator decides after 
reviewing the response and the transcript of any informal hearing to 
cancel the registration, the Administrator shall issue a final 
cancellation order, forward a copy of the cancellation order to the 
registrant by certified mail, and promptly publish the cancellation 
order in the Federal Register. Any cancellation order issued after 
receipt of a timely written response by the registrant shall become 
legally effective five days after it is published in the Federal 
Register.
    (g) Modification of Regulation. (1) In special circumstances, a 
manufacturer subject to the registration requirements of this rule may 
petition the Administrator to modify the mandatory testing requirements 
in the test standard for any test required by this rule by application 
to Director, Field Operations and Support Division, at the address in 
paragraph (f)(6)(iii) of this section.
    (i) Such request shall be made as soon as the test sponsor is aware 
that the modification is necessary, but in no event shall the request 
be made after 30 days following the event which precipitated the 
request.
    (ii) Upon such request, the Administrator may, in circumstances 
which are outside the control of the manufacturer(s) or his/their agent 
and which could not have been reasonably foreseen or avoided, modify 
the mandatory testing requirements in the rule if such requirements are 
infeasible.
    (iii) If the Administrator determines that such modifications would 
not significantly alter the scope of the test, EPA will not ask for 
public comment before approving the modification. The Administrator 
will notify the test sponsor by certified mail of the response to the 
request. EPA will place copies of each application and EPA response in 
the public docket. EPA will publish a notice in the Federal Register 
annually describing such changes which have occurred during the 
previous year. Until such Federal Register notice is published, any 
modification approved by EPA shall apply only to the person or group 
who requested the modification; EPA shall state the applicability of 
each modification in such notice.
    (iv) Where, in EPA's judgment, the requested modification of a test 
standard would significantly change the scope of the test, EPA will 
publish a notice in the Federal Register requesting comment on the 
request and proposed modification. However, EPA may approve a requested 
modification of a test standard without first seeking public comment if 
necessary to preserve the validity of an ongoing test undertaken in 
good faith.
    (2) [Reserved]
    (h) Special Requirements for Additives. An additive which is a 
direct test subject, either because it is the chosen representative of 
a group or because it is not a member of a group, is subject to the 
following rules:
    (1) All required emission characterization and health effects 
testing procedures shall be performed on the mixture which results when 
the additive is combined with the base fuel for the appropriate fuel 
family (as specified in Sec. 79.55) at the maximum concentration 
recommended by the additive manufacturer pursuant to Sec. 79.21(d). 
This combination shall be known as the additive/base fuel mixture.
    (i) The appropriate fuel family to be utilized for the additive/
base fuel mixture is the fuel family which contains the specific 
type(s) of fuel for which the additive is presently registered or for 
which the manufacturer of the additive is seeking registration.
    (ii) Fuels and additives belonging to more than one fuel family.
    (A) If a fuel or additive product is registered in two or more fuel 
families as of May 27, 1994, then the manufacturer of that product is 
responsible for testing (or participating in group testing of) each 
formulation in compliance with the requirements of this subpart for 
each fuel family in which the manufacturer wishes to maintain a product 
registration for its fuel or additive.
    (B) If a fuel or additive manufacturer is seeking to register such 
product in two or more fuel families, then the product shall be 
considered, for testing and registration purposes, to be a member of 
each fuel family in which the manufacturer is seeking registration. The 
manufacturer is responsible for testing (or participating in group 
testing of) each formulation in compliance with the requirements of 
this subpart for each fuel family in which the manufacturer wishes to 
obtain a product registration for its fuel or additive.
    (iii) In the case of the methanol fuel family, which contains two 
base fuels (M100 and M85 base fuels, pursuant to Sec. 79.55(d)), the 
applicable base fuel is the one which represents the fuel/additive 
group (specified in Sec. 79.56(e)(4)(i)(C)) containing fuels of which 
the most gallons are sold annually.
    (iv) Aftermarket additives which are intended by the manufacturer 
to be added to the fuel tank only at infrequent intervals shall be 
applied according to the manufacturer's specifications during mileage 
accumulation, pursuant to Sec. 79.57(c). However, during emission 
generation and testing, each tankful of fuel used must contain the fuel 
additive at its maximum recommended level. If the additive manufacturer 
believes that this maximum treatment rate will cause adverse effects to 
the test engine and/or that the engine's emissions may be subject to 
artifacts due to overuse of the additive, then the manufacturer may 
submit a request to EPA for modification of this requirement and 
related test procedures. Such request must include objective evidence 
that the modification(s) are needed, along with data demonstrating the 
maximum concentration of the additive which may actually reach the fuel 
tanks of vehicles in use.
    (v) Additives produced exclusively for use in #1 diesel fuel shall 
be tested in the diesel base fuel specified in Sec. 79.55(c), even 
though that base fuel is formulated with #2 diesel fuel. If a 
manufacturer is concerned that emissions generated from this 
combination of fuel and additive are subject to artifacts due to this 
blending, then that manufacturer may submit a request for a 
modification in test procedure requirements to the EPA. Any such 
request must include supporting test results and suggested test 
modifications.
    (vi) Bulk additives which are used intermittently for the direct 
purpose of conditioning or treating a fuel during storage or transport, 
or for treating or maintaining the storage, pipeline, and/or other 
components of the fuel distribution system itself and not the vehicle/
engine for which the fuel is ultimately intended, shall, for purposes 
of this program, be added to the base fuel at the maximum concentration 
recommended by the additive manufacturer for treatment of the fuel or 
distribution system component. However, if the additive manufacturer 
believes that this treatment rate will cause adverse effects to the 
test engine and/or that the engine's emissions may be subject to 
artifacts due to overuse of the additive, then the manufacturer may 
submit a request to EPA for modification of this requirement and 
related test procedures. Such request must include objective evidence 
that the modification(s) are needed, along with data demonstrating the 
maximum concentration of the additive which may actually reach the fuel 
tanks of vehicles in use.
    (2) EPA shall use emissions speciation and health effects data 
generated in the analysis of the applicable base fuel as control data 
for comparison with data generated for the additive/base fuel mixture.
    (i) The base fuel control data may be:
    (A) Generated internally as an experimental control in conjunction 
with testing done in compliance with registration requirements for a 
specific additive; or
    (B) Generated externally in the course of testing different 
additive(s) belonging to the same fuel family, or in the testing of a 
base fuel serving as representative of the baseline group for the 
respective fuel family pursuant to Sec. 79.56(e)(4)(i).
    (ii) Control data generated using test equipment (including vehicle 
model and/or engine, or Evaporative Emissions Generator specifications, 
as appropriate) and protocols identical or nearly identical to those 
used in emissions and health effects testing of the subject additive/
base fuel mixture would be most relevant for comparison purposes.
    (iii) If an additive manufacturer chooses the same vehicle/engine 
to independently test the base fuel as an experimental control prior to 
testing the additive/base fuel mixture, then the test vehicle/engine 
shall undergo two mileage accumulation periods, pursuant to 
Sec. 79.57(c). The initial mileage accumulation period shall be 
performed using the base fuel alone. After base fuel testing, and prior 
to testing of the additive/base fuel mixture, a second mileage 
accumulation period shall be performed using the additive/base fuel 
mixture. The procedures outlined in this paragraph shall not preclude a 
manufacturer from testing a base fuel and the manufacturer's additive/
base fuel mixture separately in identical, or nearly identical, 
vehicles/engines.
    (i) Multiple Test Potential for Non-Baseline Products. (1) When the 
composition information reported in the registration application or 
basic registration data for a gasoline or diesel product meets criteria 
for classification as a non-baseline product (pursuant to 
Sec. 79.56(e)(3)(i)(B) or Sec. 79.56(e)(3)(ii)(B)), then the 
manufacturer is responsible for testing (or participating in group 
testing) of a separate formulation for each reported oxygenating 
compound, specified class of oxygenating compounds, or other substance 
which defines a separate non-baseline fuel/additive group pursuant to 
Sec. 79.56(e)(4)(ii)(A) or (B). For each such substance, testing shall 
be performed on a mixture of the relevant substance in the appropriate 
base fuel, formulated according to the specifications for the 
corresponding group representatives in Sec. 79.56(e)(4)(ii).
    (2) When the composition information reported in the registration 
application or basic registration data for a non- baseline gasoline 
product contains a range of total oxygenate concentration-in-use which 
encompasses gasoline formulations with less than 1.5 weight percent 
oxygen as well as gasoline formulations with 1.5 weight percent oxygen 
or more, then the manufacturer is required to test (or participate in 
applicable group testing of) a baseline gasoline formulation as well as 
one or more non-baseline gasoline formulations as described in 
paragraph (h)(1) of this section.
    (3) When the composition information reported in the registration 
application or basic registration data for a non- baseline diesel 
product contains a range of total oxygenate concentration-in-use which 
encompasses diesel formulations with less than 1.0 weight percent 
oxygen as well as diesel formulations with 1.0 weight percent oxygen or 
more, then the manufacturer is required to test (or participate in 
applicable group testing) of a baseline diesel formulation as well as 
one or more non-baseline diesel formulations as described in paragraph 
(h)(1) of this section.
    (j) Multiple Test Potential for Atypical Fuel Formulations. When 
the composition information reported in the registration application or 
basic registration data for a fuel product includes more than one 
atypical bulk additive product (pursuant to Sec. 79.56(e)(2)(iii)), and 
when these additives belong to different fuel/additive groups (pursuant 
to Sec. 79.56(e)(4)(iii)), then:
    (1) When such disparate additive products are for the same purpose-
in-use and are not ordinarily used in the fuel simultaneously, the fuel 
manufacturer shall be responsible for testing (or participating in the 
group testing of) a separate formulation for each such additive 
product. Testing related to each additive product shall be performed on 
a mixture of the additive in the applicable base fuel, as described in 
paragraph (g)(1) of this section, or by participation in the costs of 
testing the designated representative of the fuel/additive group to 
which each separate atypical additive product belongs.
    (2) When the disparate additive products are not for the same 
purpose-in-use, the fuel manufacturer shall nevertheless be responsible 
for testing a separate formulation for each such additive product, as 
described in paragraph (g)(1) of this section, if these additives are 
not ordinarily blended together in the same commercial formulation of 
the fuel.
    (3) When the disparate additive products are ordinarily blended 
together in the same commercial formulation of the fuel, then the fuel 
manufacturer shall be responsible for the testing of a single test 
formulation containing all such simultaneously used atypical additive 
products. Alternatively, this responsibility can be satisfied by 
enrolling such fuel product in a group which includes other fuel or 
additive products with the same total combination of atypical elements 
as that occurring in the fuel product in question. If the basic 
registration data for the subject fuel includes any alternative 
additives which contain atypical elements not represented in the test 
formulation, then the fuel manufacturer is also responsible for testing 
a separate formulation for each such additional disparate additive 
product.
    (k) Emission Control System Testing. If any information submitted 
in accordance with this subpart or any other information available to 
EPA shows that a fuel or fuel additive may have a deleterious effect on 
the performance of any emission control system or device currently in 
use or which has been developed to a point where in a reasonable time 
it would be in general use were such effect avoided, EPA may, in its 
judgment, require testing to determine whether such effects in fact 
exist. Such testing will be required in accordance with such protocols 
and schedules as the Administrator shall reasonably require and shall 
be paid for by the fuel or fuel additive manufacturer.


Sec. 79.52  Tier 1.

    (a) General Specifications. Tier 1 requires manufacturers of 
designated fuels or fuel additives (or groups of manufacturers pursuant 
to Sec. 79.56) to supply to the Administrator: the identity and 
concentration of certain emission products of such fuels or additives; 
an analysis of potential emissions exposures; and any available 
information regarding the health and welfare effects of the whole and 
speciated emissions. In addition to any information required under 
Sec. 79.59 and in conformance with the reporting requirements thereof, 
manufacturers shall provide, pursuant to the timing provisions of 
Sec. 79.51(c), the following information.
    (b) Emissions Characterization. Manufacturers must provide a 
characterization of the emission products which are generated by 
evaporation (if required pursuant to Sec. 79.58(b)) and by combustion 
of the fuel or additive/base fuel mixture in a motor vehicle. For this 
purpose, manufacturers may perform the characterization procedures 
described in this section or may rely on existing emission 
characterization data. To be considered adequate in lieu of performing 
new emission characterization procedures, the data must be the result 
of tests using the product in question or using a fuel or additive/base 
fuel mixture meeting the same grouping criteria as the product in 
question. In addition, the emissions must be generated in a manner 
reasonably similar to those described in Sec. 79.57, and the 
characterization procedures must be adequately performed and documented 
and must give results reasonably comparable to those which would be 
obtained by performing the procedures described herein. Reports of 
previous tests must be sufficiently detailed to allow EPA to judge the 
adequacy of protocols, techniques, and conclusions. After the 
manufacturer's submittal of such data, if EPA finds that the 
manufacturer has relied upon inadequate test data, then the 
manufacturer will not be considered to be in compliance until the 
corresponding tests have been conducted and the results submitted to 
EPA.
    (1) General Provisions.
    (i) The emissions to be characterized shall be generated, 
collected, and stored according to the processes described in 
Sec. 79.57. Characterization of combustion and evaporative emissions 
shall be performed separately on each emission sample collected during 
the applicable emission generation procedure.
    (ii) As provided in Sec. 79.57(d), if the emission generation 
vehicle/engine is ordinarily equipped with an emission aftertreatment 
device, then all requirements in this section for the characterization 
of combustion emissions must be completed both with and without the 
aftertreatment device in a functional state. The emissions shall be 
generated three times (on three different days) without a functional 
aftertreatment device and, if applicable, three times (on three 
different days) with a functional aftertreatment device, and each such 
time shall be analyzed according to the remaining provisions in this 
paragraph (b) of this section.
    (iii) Measurement of background emissions. It is required that 
ambient/dilution air be analyzed for levels of background chemical 
species present at the time of emission sampling (for both combustion 
and evaporative emissions) and that background chemical species 
profiles be reported with emissions speciation data. Background 
chemical species measurement/analysis during the FTP is specified in 
Secs. 86.109-94(c)(5) and 86.135-94 of this chapter.
    (iv) Concentrations of emission products shall be reported in units 
of grams (g) per mile and in units of weight percent of measured total 
hydrocarbons.
    (v) Laboratory practice must be of high quality and must be 
consistent with state-of-the-art methods as presented in current 
environmental and analytical chemistry literature. Examples of 
analytical procedures which may be used in conducting the emission 
characterization/speciation requirements of this section can be found 
among the references in paragraph (b)(5) of this section.
    (2) Characterization of the combustion emissions shall include, for 
products in all fuel families (except when expressly noted in this 
section):
    (i) Determination of the concentration of the basic emissions as 
follows: total hydrocarbons, carbon monoxide, oxides of nitrogen, and 
particulates. Manufacturers are referred to the vehicle certification 
procedures in 40 CFR part 86, subparts B and D (Secs. 86.101 through 
86.145 and Secs. 86.301 through 86.348) for guidance on the measurement 
of the basic emissions of interest to this subpart.
    (ii) Characterization of the vapor phase of combustion emissions, 
as follows:
    (A) Determination of the identity and concentration of individual 
species of hydrocarbon compounds containing 12 or fewer carbon atoms. 
Such characterization shall begin within 30 minutes after emission 
collection is completed.
    (B) Determination of the identity and concentration of individual 
species of aldehyde and ketone compounds containing eight or fewer 
carbon atoms. Characterization of these emissions captured in 
cartridges shall be performed within two weeks if the cartridge is 
stored at room temperature, and one month if the cartridge is stored at 
0  deg.C or less. If the emissions are sampled using the impinger 
method, the sample must be stored in a capped sample vial at 0  deg.C 
or less and characterized within one week.
    (C) Determination of the identity and concentration of individual 
species of alcohol and ether compounds containing six or fewer carbon 
atoms, for those fuels and additive/base fuel mixtures which contain 
alcohol and/or ether compounds containing from one to six carbon atoms 
in the uncombusted state. For fuel and additive formulations containing 
alcohols or ethers with more than six carbon atoms in the uncombusted 
state, alcohol and ether species with that higher number of carbon 
atoms or less must be identified and measured in the emissions. Such 
characterization shall begin within four hours after emission 
collection is completed.
    (iii) Characterization of the semi-volatile and particulate phases 
of combustion emissions to identify and measure polycyclic aromatic 
compounds, as follows:
    (A) Analysis for polycyclic aromatic compounds shall not be 
conducted at or soon after the start of a recommended engine lubricant 
change interval.
    (B) Analysis for polycyclic aromatic hydrocarbons (PAHs) and 
nitrated polycyclic aromatic hydrocarbons (NPAHs), specified in 
paragraph (b)(2)(iii)(D) of this section, need not be done for any 
fuels and additives in the methane or propane fuel families, nor for 
fuels and additives in the atypical categories of any other fuel 
families, pursuant to the definitions of such families and categories 
in Sec. 79.56.
    (C) Analysis for poly-chlorinated dibenzodioxins and dibenzofurans 
(PCDD/PCDFs), specified in paragraph (b)(2)(iii)(E) of this section, is 
required only for fuels and additives which contain chlorine as an 
atypical element, pursuant to paragraph (b)(2)(iv) of this section, 
which requires all individual emission products containing atypical 
elements to be determined for atypical fuels and additives. However, 
manufacturers of baseline and nonbaseline fuels and fuel additives in 
all fuel families, except those in the methane and propane fuel 
families, are strongly encouraged to conduct these analyses on a 
voluntary basis.
    (D) The analytical method used to measure species of PAHs and NPAHs 
should be capable of detecting at least 1 ppm (equivalent to 0.001 
microgram (g) of compound per milligram of organic extract) of 
these compounds in the extractable organic matter. The concentration of 
each individual PAH or NPAH compound identified shall be reported in 
units of microgram per mile. Each compound which is present at 0.001 
g per mile or more must be identified, measured, and reported. 
The following individual species shall be measured:
    (1) PAHs:
    (i) Benzo(a)anthracene;
    (ii) Benzo[b]fluoranthene;
    (iii) Benzo[k]fluoranthene;
    (iv) Benzo(a)pyrene;
    (v) Chrysene;
    (vi) Dibenzo[a,h]anthracene; and
    (vii) Indeno[1,2,3-c,d]pyrene.
    (2) NPAHs:
    (i) 7-Nitrobenzo[a]anthracene;
    (ii) 6-Nitrobenzo[a]pyrene;
    (iii) 6-Nitrochrysene;
    (iv) 2-Nitrofluorene; and
    (v) 1-Nitropyrene.
    (E) The analytical method used to measure species and classes of 
PCDD/PCDFs should be capable of detecting at least 1 part per trillion 
(ppt) (equivalent to 0.001 picogram (pg) of compound per milligram of 
organic extract) of these compounds in the extractable organic matter. 
The concentration of each individual PCDD/PCDF compound identified 
shall be reported in units of picograms (pg) per mile. Each compound 
which is present at 0.5 pg per mile or more must be identified, 
measured, and reported.
    (1) With respect to measurement of PCDD/PCDFs only, the liquid 
extracts from the particulate and semi-volatile emissions fractions may 
be combined into one sample for analysis.
    (2) The manufacturer is referred to 40 CFR part 60, appendix A, 
Method 23 for a protocol which may be used to identify and measure any 
potential PCDD/PCDFs which might be present in exhaust emissions from a 
fuel or additive/base fuel mixture.
    (3) The following individual compounds and classes of compounds of 
PCDD/PCDFs shall be identified and measured:
    (i) Individual tetra-chloro-substituted dibenzodioxins (tetra-
CDDs);
    (ii) Individual tetra-chloro-substituted dibenzofurans (tetra-
CDFs);
    (iii) Penta-CDDs and penta-CDFs, as one class;
    (iv) Hexa-CDDs and hexa-CDFs, as one class;
    (v) Hepta-CDDs and hepta-CDFs as one class; and
    (vi) Octo-CDDs and octo-CDFs as one class.
    (iv) With respect to all phases (vapor, semi-volatile, and 
particulate) of combustion emissions generated from those fuels and 
additive/base fuel mixtures classified in the atypical categories 
(pursuant to Sec. 79.56), the identity and concentration of individual 
emission products containing such atypical elements shall also be 
determined.
    (3) For evaporative fuels and evaporative fuel additives, 
characterization of the evaporative emissions shall include:
    (i) Determination of the concentration of total hydrocarbons for 
the applicable vehicle type and class in 40 CFR part 86, subpart B 
(Secs. 86.101 through 86.145).
    (ii) Determination of the identity and concentration of individual 
species of hydrocarbon compounds containing 12 or fewer carbon atoms. 
Such characterization shall begin within 30 minutes after emission 
collection is completed.
    (iii) In the case of those fuels and additive/base fuel mixtures 
which contain alcohol and/or ether compounds in the uncombusted state, 
determination of the identity and concentration of individual species 
of alcohol and ether compounds containing six or fewer carbon atoms. 
For fuel and additive formulations containing alcohols or ethers with 
more than six carbon atoms in the uncombusted state, alcohol and ether 
species with that higher number of carbon atoms or less must be 
identified and measured in the emissions. Such characterization shall 
begin within four hours after emission collection is completed.
    (iv) In the case of those fuels and additive/base fuel mixtures 
which contain atypical elements, determination of the identity and 
concentration of individual emission products containing such atypical 
elements.
    (4) Laboratory quality control. (i) At a minimum, laboratories 
performing the procedures specified in this section shall conduct 
calibration testing of their emissions characterization equipment 
before each new fuel/additive product test start-up. Known samples 
representative of the compounds potentially to be found in emissions 
from the product to be characterized shall be used to calibrate such 
equipment.
    (ii) Laboratories performing the procedures specified in this 
section shall agree to permit quality control inspections by EPA, and 
for this purpose shall admit any EPA Enforcement Officer, upon proper 
presentation of credentials, to any facility where vehicles are 
conditioned or where emissions are generated, collected, stored, 
sampled, or characterized in meeting the requirements of this section. 
Such laboratory audits may include EPA distribution of ``blind'' 
samples for analysis by participating laboratories.
    (5) References. For additional background information on the 
emission characterization procedures outlined in this paragraph, the 
following references may be consulted:
    (i) ``Advanced Emission Speciation Methodologies for the Auto/Oil 
Air Quality Improvement Program--I. Hydrocarbons and Ethers,'' Auto Oil 
Air Quality Improvement Research Program, SP-920, 920320, SAE, February 
1992.
    (ii) ``Advanced Speciation Methodologies for the Auto/Oil Air 
Quality Improvement Research Program--II. Aldehydes, Ketones, and 
Alcohols,'' Auto Oil Air Quality Improvement Research Program, SP-920, 
920321, SAE, February 1992.
    (iii) ASTM D 5197-91, ``Standard Test Method for Determination of 
Formaldehyde and Other Carbonyl Compounds in Air (Active Sampler 
Methodology).''
    (iv) Johnson J. H., Bagley, S. T., Gratz, L. D., and Leddy, D. G., 
``A Review of Diesel Particulate Control Technology and Emissions 
Effects--1992 Horning Memorial Award Lecture,'' SAE Technical Paper 
Series, SAE 940233, 1994.
    (v) Keith et al., ACS Committee on Environmental Improvement, 
``Principles of Environmental Analysis,'' The Journal of Analytical 
Chemistry, Volume 55, pp. 2210-2218, 1983.
    (vi) Perez, J.M., Jabs, R.E., Leddy, D.G., eds. ``Chemical Methods 
for the Measurement of Unregulated Diesel Emissions (CRC-APRAC Project 
No. CAPI-1-64), Coordinating Research Council, CRC Report No. 551, 
August, 1987.
    (vii) Schuetzle, D., ``Analysis of Nitrated Polycyclic Aromatic 
Hydrocarbons in Diesel Particulates,'' Analytical Chemistry, Volume 54, 
pp. 265-271, 1982.
    (viii) Siegl, W.O., et al., ``Improved Emissions Speciation 
Methodology for Phase II of the Auto/Oil Air Quality Improvement 
Research Program--Hydrocarbons and Oxygenates'', SAE Technical Paper 
Series, SAE 930142, 1993.
    (ix) Tejada, S. B. et al., ``Analysis of Nitroaromatics in Diesel 
and Gasoline Car Emissions,'' SAE Paper No. 820775, 1982.
    (x) Tejada, S. B. et al., ``Fluorescence Detection and 
Identification of Nitro Derivatives of Polynuclear Aromatic 
Hydrocarbons by On-Column Catalytic Reduction to Aromatic Amines,'' 
Analytical Chemistry, Volume 58, pp. 1827-1834, July 1986.
    (xi) ``Test Method for Determination of C1-C4 Alcohols and MTBE in 
Gasoline by Gas Chromatography,'' 40 CFR part 80, appendix F.
    (c) Exposure Analysis. Using annual and projected production 
volume, marketing, and distribution data submitted as part of the basic 
registration data, specified in Sec. 79.59(b), manufacturers shall 
provide a qualitative discussion of the potential public health 
exposure(s) of the general population and any special at-risk 
populations to the emission products of their fuel or additive 
product(s). The analysis accompanying a group submission shall address 
the characteristics of the cumulative exposure resulting from the use 
of all fuel or additive products in the group. Modeling and other 
quantitative approaches to the analysis are encouraged when the 
appropriate data is available.
    (d) Literature Search. (1) Manufacturers of fuels and fuel 
additives shall conduct a literature search and compilation of 
information on the potential toxicologic, environmental, and other 
public welfare effects of the emissions of such fuels and additives. 
The literature search shall include all available relevant information 
from in-house, industry, government, and public sources pertaining to 
the emissions of the subject fuel or fuel additive or the emissions of 
similar fuels or additives, with such similarity determined according 
to the provisions of Sec. 79.56.
    (2) The literature search shall address the potential adverse 
effects of whole combustion emissions, evaporative emissions, relevant 
emission fractions, and individual emission products of the subject 
fuel or fuel additive except as specified in the following paragraph. 
The individual emission products to be included are those identified 
pursuant to the emission characterization procedures specified in 
paragraph (b) of this section, other than carbon monoxide, carbon 
dioxide, nitrogen oxides, benzene, 1,3-butadiene, acetaldehyde, and 
formaldehyde.
    (3) In the case of the individual emission products of non-baseline 
or atypical fuels and additives (pursuant to Sec. 79.56(e)(2)), the 
literature data need not be submitted for those emission products which 
are the same as the combustion emission products of the respective base 
fuel for the product's fuel family (pursuant to Sec. 79.55). For this 
purpose, data on the base fuel emission products for the product's fuel 
family:
    (i) May be found in the literature of previously-conducted, 
adequate emission speciation studies for the base fuel, or for a fuel 
or additive/fuel mixture capable of grouping with the base fuel (see, 
for example, the references in paragraph (b)(5) of this section).
    (ii) May be compiled while gathering internal control data during 
emissions characterization studies on the manufacturer's non-baseline 
or atypical product; or
    (iii) May be obtained from various manufacturers in the course of 
their testing different additive(s) belonging to the same fuel family, 
or in the testing of a base fuel serving as representative of the 
baseline group for the respective fuel family.
    (e) Data bases. The literature search must include the results of 
searching appropriate commercially available chemical, toxicologic, and 
environmental databases. The databases shall be searched using, at a 
minimum, CAS numbers (when applicable), chemical names, and common 
synonyms.
    (f) Search period. The literature search shall cover a time period 
beginning at least thirty years prior to the date of submission of the 
reports specified in Secs. 79.59(b) through (c) and ending no earlier 
than six months prior to the date on which testing is commenced or 
reports are submitted in compliance with this subpart.
    (g) References. Information on base fuel emission inventories may 
be found in references in paragraphs (b)(5)(i) through (xi) of this 
section, as well as in the following:
    (1) Auto/Oil Air Quality Improvement Research Program, Technical 
Bulletin #1, December 1990.
    (2) Keith et al., ACS Committee on Environmental Improvement, 
``Principles of Environmental Analysis,'' The Journal of Analytical 
Chemistry, Volume 55, pp. 2210-2218, 1983.
    (3) ``The Composition of Gasoline Engine Hydrocarbon Emissions--An 
Evaluation of Catalyst and Fuel Effects''--SAE 902074 and ``Speciated 
Hydrocarbon Emissions from Aromatic, Olefin, and Paraffinic Model 
Fuels''--SAE 930373.


Sec. 79.53  Tier 2.

    (a) Generally. Subject to the provisions of Sec. 79.53(b) through 
(d), the combustion emissions of each fuel or fuel additive subject to 
testing under this subpart must be tested in accordance with each of 
the testing guidelines in Secs. 79.60 through 79.68, except that fuels 
and additives in the methane and propane fuel families (pursuant to 
Sec. 79.56(e)(1)(v) and (vi)) need not undergo the Salmonella 
mutagenicity assay in Sec. 79.68). Similarly, subject to the provisions 
of Sec. 79.53(b) through (d), the evaporative emissions of each 
designated evaporative fuel and each designated evaporative fuel 
additive subject to testing under this subpart must be tested according 
to each of the testing guidelines in Secs. 79.60 through 79.67 
(excluding Sec. 79.68, Salmonella typhimurium Reverse Mutation Assay).
    (b) Manufacturer Determination. Manufacturers shall determine 
whether the information gathered pursuant to the literature search in 
Sec. 79.52(d) contains the results of adequately performed and 
adequately documented previous testing which provides information 
reasonably comparable to that supplied by the health tests described in 
Secs. 79.62 through 79.68 regarding the carcinogenicity, mutagenicity, 
neurotoxicity, teratogenicity, reproductive/fertility measures, and 
general toxicity effects of the emissions of the fuel or additive. When 
manufacturers make an affirmative determination, they need submit only 
the information gathered pursuant to Sec. 79.52(d) for such tests. EPA 
maintains final authority in judging whether the information is an 
adequate substitution in lieu of conducting the associated tests. EPA's 
determination of the adequacy of existing information shall be guided 
by the considerations described in paragraph (d) of this section. If 
EPA finds that the manufacturer has relied upon inadequate test data, 
then the manufacturer will not be considered to be in compliance until 
the corresponding tests have been conducted and the results submitted 
to EPA.
    (c) Testing. (1) All testing required pursuant to this section must 
be done in accordance with the procedures, equipment, and facility 
requirements described in Secs. 79.57, 79.60, and 79.61 regarding 
emissions generation, good laboratory practices, and inhalation 
exposure testing, respectively, as well as any other requirements 
described in this subpart. The laboratory conducting the animal studies 
shall be registered and in good standing with the United States 
Department of Agriculture and regularly inspected by United States 
Department of Agriculture veterinarians. In addition, the facility must 
be accredited by a generally recognized independent organization which 
sets laboratory animal care standards. Use of inadequate test protocols 
or substandard laboratory techniques in performing any testing required 
by this subpart may result in cancellation of all affected 
registrations.
    (2) Carcinogenic or mutagenic effects in animals from emissions 
exposures shall be determined pursuant to Sec. 79.64 In vivo 
Micronucleus Assay, Sec. 79.65 In vivo Sister Chromatid Exchange Assay, 
and Sec. 79.68 Salmonella typhimurium Reverse Mutation Assay. 
Teratogenic effects and reproductive toxicity shall be examined 
pursuant to Sec. 79.63 Fertility Assessment/Teratology. General 
toxicity and pulmonary effects shall be determined pursuant to 
Sec. 79.62 Subchronic Toxicity Study with Specific Health Effect 
Assessments. Neurotoxic effects shall be determined pursuant to 
Sec. 79.66 Neuropathology Assessment and Sec. 79.67 Glial Fibrillary 
Acidic Protein Assay.
    (d) EPA Determination. (1) After submission of all information and 
testing, EPA in its judgment shall determine whether previously 
conducted tests relied upon in the registration submission are 
adequately performed and documented and provide information reasonably 
comparable to that which would be provided by the tests described 
herein. Manufacturers' submissions shall be sufficiently detailed to 
allow EPA to judge the adequacy of protocols, techniques, experimental 
design, statistical analyses, and conclusions. Studies shall be 
performed using generally accepted scientific principles, good 
laboratory techniques, and the testing guidelines specified in these 
regulations.
    (2) EPA shall give appropriate weight when making this 
determination to the following factors:
    (i) The age of the data;
    (ii) The adequacy of documentation of procedures, findings, and 
conclusions;
    (iii) The extent to which the testing conforms to generally 
accepted scientific principles and practices;
    (iv) The type and number of test subjects;
    (v) The number and adequacy of exposure concentrations, i.e., 
emission dilutions;
    (vi) The degree to which the tested emissions were generated by 
procedures and under conditions reasonably comparable to those set 
forth in Sec. 79.57; and
    (vii) The degree to which the test procedures conform to the 
testing guidelines set forth in Secs. 79.60 through 79.68 and/or 
furnish information comparable to that provided by such testing.
    (3) The test animals shall be rodents, preferably a strain of rat, 
and testing shall include all of the endpoints covered in Secs. 79.62 
through 79.68. All studies shall be properly executed, with appropriate 
documentation, and in accord with the individual health testing 
guidelines (Secs. 79.60 through 79.68) of this part, e.g., 90-day, 6-
hour per day exposure, minimum.
    (4) In general, the data in a manufacturer's registration submittal 
shall be adequate if the duration of a test's exposure period is at 
least as long, in days and hours, as the inhalation exposure specified 
in the related health test guideline(s). Data from tests with shorter 
exposure durations than those specified in the guidelines may be 
acceptable if the test results are positive (i.e., exhibit adverse 
effects) and/or include a demonstrable concentration-response 
relationship.
    (5) Data in support of a manufacturer's registration submittal 
shall directly address the effects of inhalation exposure to the whole 
evaporative and exhaust emissions of the respective fuel or additive or 
to the whole evaporative and exhaust emissions of other fuels or 
additives which satisfy the criteria in Sec. 79.56 for classification 
into the same group as the subject fuel or fuel additive. Data obtained 
in the testing of a raw liquid fuel or additive/base fuel mixture or a 
raw, aerosolized fuel or additive/base fuel mixture shall not be 
adequate to support a manufacturer's registration submittal. Data from 
testing of evaporative emissions cannot substitute for test data on 
combustion emissions. Data from testing of combustion emissions cannot 
substitute for test data on evaporative emissions.


Sec. 79.54  Tier 3.

    (a) General Criteria for Requiring Tier 3 Testing. (1) Tier 3 
testing shall be required of a manufacturer or group of manufacturers 
at EPA's discretion when remaining uncertainties as to the significance 
of observed health effects, welfare effects, and/or emissions exposures 
from a fuel or fuel/additive mixture interfere with EPA's ability to 
make reasonable estimates of the potential risks posed by emissions 
from the fuel or additive products. Tier 3 testing may be conducted 
either on an individual basis or a group basis. If performed on a group 
basis, EPA may require either the same representative to be used in 
Tier 3 testing as was used in Tier 2 testing or may select a different 
member or members of the group to represent the group in the Tier 3 
tests.
    (2) In addition to the criteria specific to particular tests as 
summarized and detailed in the testing guidelines (Secs. 79.62 through 
79.68), EPA may consider a number of factors (including, but not 
limited to):
    (i) The number of positive and negative outcomes related to each 
endpoint;
    (ii) The identification of concentration-effect relationships;
    (iii) The statistical sensitivity and significance of such studies;
    (iv) The severity of the observed effects (e.g., whether the 
effects would be likely to lead to incapacitating or irreversible 
conditions);
    (v) The type and number of species included in the reported tests;
    (vi) The consistency and clarity of apparent mechanisms, target 
organs, and outcomes;
    (vii) The presence or absence of effective health test control data 
for base-fuel-only versus additive/base fuel mixture comparisons;
    (viii) The nature and amount of known toxic agents in the emissions 
stream; and
    (ix) The observation of lesions which specifically implicate 
inhalation as an important exposure route.
    (3) Consideration of exposure. EPA retains discretion to consider, 
in addition to available toxicity data, any Tier 1 data on potential 
exposures to emissions from a particular fuel or fuel additive (or 
group of fuels and/or fuel additives) in determining whether to require 
Tier 3 testing. EPA may consider, but is not limited to, the following 
factors:
    (i) Types and emission rates of speciated emission components;
    (ii) Types and emission rates of combinations of compounds or 
elements of concern;
    (iii) Historical and/or projected production volumes and market 
distributions; and
    (iv) Estimated population and/or environmental exposures obtained 
through extrapolation, modeling, or literature search findings on 
ambient, occupational, or epidemiological exposures.
    (b) Notice. (1) EPA will determine whether Tier 3 testing is 
necessary upon receipt of a manufacturer's (or group's) submittal as 
prescribed under Sec. 79.51(d). If EPA determines on the basis of the 
Tier 1 and 2 data submission and any other available information that 
further testing is necessary, EPA will require the responsible 
manufacturer(s) to conduct testing as described elsewhere in this 
section. EPA will notify the manufacturer (or group) by certified 
letter of the purpose and nature of any proposed testing and of the 
proposed deadline for completing the testing. A copy of the letter will 
be placed in the public record. EPA will provide the manufacturer a 60-
day comment period after the manufacturer's receipt of such notice. EPA 
may extend the comment period if it appears from the nature of the 
issues raised that further discussion is warranted. In the event that 
no comment is received by EPA from the manufacturer (or group) within 
the comment period, the manufacturer (or group) shall be deemed to have 
consented to the adoption by EPA of the proposed Tier 3 requirements.
    (2) EPA will issue a notice in the Federal Register of its intent 
to require testing under Tier 3 for a particular fuel or additive 
manufacturer and that a copy of the letter to the manufacturer 
outlining the Tier 3 testing for that manufacturer is available in the 
public record for review and comment. The public shall have a minimum 
of thirty (30) days after the publication of this notice to comment on 
the proposed Tier 3 testing.
    (3) EPA will include in the public record a copy of any timely 
comments concerning the proposed Tier 3 testing requirements received 
from the affected manufacturer or group or from the public, and the 
responses of EPA to such comments. After reviewing all such comments 
received, EPA will adopt final Tier 3 requirements by sending a 
certified letter describing such final requirements to the manufacturer 
or group. EPA will also issue a notice in the Federal Register 
announcing that it has adopted such final Tier 3 requirements and that 
a copy of the letter adopting the requirements has been included in the 
public record.
    (4) Prior to beginning any required Tier 3 testing, the 
manufacturer shall submit detailed test protocols to EPA for approval. 
Once EPA has determined the Tier 3 testing requirements and approves 
the test protocols, any modification to the requirements shall be 
governed by Sec. 79.51(f).
    (c) Carcinogenicity and Mutagenicity Testing. (1) A potential need 
for Tier 3 carcinogenicity and/or mutagenicity testing may be indicated 
if the results of the In vivo Micronucleus Assay, required under 
Sec. 79.64, the In vivo Sister Chromatid Exchange Assay, required under 
Sec. 79.65, the Salmonella mutagenicity assay required under 
Sec. 79.68, or relevant pathologic findings under Sec. 79.62 
demonstrate a statistically significant dose-related positive response 
as compared with appropriate controls. Alternatively, Tier 3 
carcinogenicity testing and/or mutagenicity testing may be required if 
there are positive outcomes for at least one concentration in two or 
more of the tests required under Secs. 79.64, 79.65, and 79.68.
    (2) The testing for carcinogenicity required under this paragraph 
may, at EPA's discretion, be conducted in accordance with 40 CFR 
798.3300 or 798.3320, or their equivalents (see suggested references 
following each health effects testing guideline). The testing for 
mutagenicity required under this paragraph may likewise be conducted in 
accordance with 40 CFR 798.5195, 798.5500, 798.5955, 798.7100, and/or 
other suitable equivalent testing (see suggested references following 
each health effects testing guideline). EPA may supplement or modify 
guidelines as required to ensure that the prescribed testing addresses 
the identified areas of concern.
    (d) Reproductive and Teratological Effects Testing. (1) A potential 
need for Tier 3 testing may be indicated if the results of the 
Fertility Assessment/Teratology study required under Sec. 79.63 or 
relevant findings under Sec. 79.62 demonstrate, in comparison with 
appropriate controls, a statistically significant dose-related positive 
response in one or more of the possible test outcomes. Similarly, Tier 
3 testing may be indicated if statistically significant positive 
results are confined to either sex, or to the fetus as opposed to the 
pregnant adult.
    (2) The testing for reproductive and teratological effects required 
under this paragraph may, at EPA's discretion, be conducted in 
accordance with 40 CFR 798.4700 and/or by performance of a reproductive 
assay by continuous breeding. These guidelines may be modified or 
supplemented by EPA as required to ensure that the prescribed testing 
addresses the identified areas of concern.
    (e) Neurotoxicity Testing. (1) A potential need for Tier 3 
neurotoxicity testing may be indicated if either the results of the 
Neuropathology Assessment required under Sec. 79.67 shows 
concentration-related effects in exposed animals or the Glial 
Fibrillary Acidic Protein Assay required under Sec. 79.66 demonstrates 
a statistically significant concentration-related positive response as 
compared with appropriate controls. Similarly, Tier 3 neurotoxicity 
testing may be indicated if relevant results under Sec. 79.62 
demonstrate a statistically significant positive response in comparison 
to appropriate controls.
    (2) The testing for neurotoxicity required under this paragraph 
may, at EPA's discretion, be conducted in accordance with 40 CFR 
798.3260 and 40 CFR part 798 subpart G. These guidelines may be 
modified or supplemented by EPA as required to ensure that the 
prescribed testing addresses the identified areas of concern.
    (f) General and Pulmonary Toxicity Testing. (1) A potential need 
for Tier 3 general and/or pulmonary toxicity testing may be indicated 
if, in comparison with appropriate controls, the results of the 
Subchronic Toxicity Study, pursuant to Sec. 79.62, demonstrate abnormal 
gross analysis or histopathological findings (especially as relates to 
lung pathology from whole-body preserved test animals) or persistence 
or delayed occurrence of toxic effects beyond the exposure period.
    (2) A potential need for Tier 3 testing with respect to other organ 
systems or endpoints not addressed by specific Tier 2 tests, e.g., 
hepatic, renal, or endocrine toxicity, may be demonstrated by findings 
in the Tier 2 Subchronic Toxicity Study (pursuant to Sec. 79.62) or by 
findings in the Tier 1 literature search of adverse functional, 
physiologic, metabolic, or histopathologic effects of fuel or additive 
emissions to such other organ systems or any other information 
available to EPA. In addition, findings in the Tier 1 emission 
characterization of significant levels of a known toxicant to such 
other organ systems and endpoints may also indicate a need for relevant 
health effects testing. The testing required under this paragraph may 
include tests conducted in accordance with 40 CFR 798.3260 or 798.3320. 
These guidelines may be modified or supplemented by EPA as necessary to 
ensure that the prescribed testing addresses the identified areas of 
concern.
    (3) The testing for general/pulmonary toxicity required under this 
paragraph may, at EPA's discretion, be conducted in accordance with 40 
CFR 798.2450 or 798.3260. These guidelines may be modified or 
supplemented by EPA as necessary to ensure that the prescribed testing 
addresses the identified areas of concern. Pulmonary function 
measurements, host defense assays, immunotoxicity tests, cell 
morphology/morphometry, and/or enzyme assays of lung lavage cells and 
fluids may be specifically required.
    (g) Other Tier 3 Testing. (1) A manufacturer or group may be 
required to use up-to-date modeling, sampling, monitoring, and/or 
analytic approaches at the Tier 3 level to provide:
    (i) Estimates of exposures to the emission products of a fuel or 
fuel additive or group of products;
    (ii) The expected atmospheric transformation products of such 
emissions; and
    (iii) The environmental partitioning of such emissions to the air, 
soil, water, and biota.
    (2) Additional emission characterization may be required if 
uncertainty over the identity of chemical species or rate of their 
emission interferes with reasonable judgments as to the presence and/or 
concentration of potentially toxic substances in the emissions of a 
fuel or fuel additive. The required tests may include characterization 
of additional classes of emissions, the characterization of emissions 
generated by additional vehicles/engines of various technology mixes 
(e.g., catalyzed versus non-catalyzed emissions), and/or other more 
precise analytic procedures for identification or quantification of 
emissions compounds. Additional emissions testing may also be required 
to evaluate concerns which may arise regarding the potential effects of 
a fuel or fuel additive on the performance of emission control 
equipment.
    (3) A manufacturer or group may be required to conduct biological 
and/or exposure studies at the Tier 3 level to evaluate directly the 
potential public welfare or environmental effects of the emissions of a 
fuel or additive, if significant concerns about such effects arise as a 
result of EPA's review of the literature search or emission 
characterization findings in Tier 1 or the results of the toxicological 
tests in Tier 2.
    (4) With regard to group submittals, Tier 3 studies on a fuel or 
additive product(s) other than the originally specified group 
representative may be required if specific differences in the product's 
composition indicate that its emissions may have different toxicologic 
properties from those of the original group representative.
    (5) Additional emission characterization and/or toxicologic tests 
may be required to evaluate the impact of different vehicle, engine, or 
emission control technologies on the observed composition or health or 
welfare effects of the emissions of a fuel or additive.
    (6) Toxicological tests on individual emission products may be 
required.
    (7) Upon review of information submitted for an aerosol product 
under Sec. 79.58(e), emissions characterization, exposure, and/or 
toxicologic testing at a Tier 3 level may be required.
    (8) A manufacturer which qualifies for and has elected to use the 
special provisions for the products of small businesses (pursuant to 
Sec. 79.58(d)) may be required to conduct emission characterization, 
exposure, and /or toxicologic studies at the Tier 3 level for such 
products, as specified in Sec. 79.58(d)(4).
    (9) The examples of potential Tier 3 tests described in this 
section do not in any way limit EPA's broad discretion and authority 
under Tier 3.


Sec. 79.55  Base fuel specifications.

    (a) General Characteristics. (1) The base fuel(s) in each fuel 
family shall serve as the group representative(s) for the baseline 
group(s) in each fuel family pursuant to Sec. 79.56. Also, as specified 
in Sec. 79.51(h)(1), for fuel additives undergoing testing, the 
designated base fuel for the respective fuel family shall serve as the 
substrate in which the additive shall be mixed prior to the generation 
of emissions.
    (2) Base fuels shall contain a limited complement of the additives 
which are essential for the fuel's production or distribution and/or 
for the successful operation of the test vehicle/engine throughout the 
mileage accumulation and emission generation periods. Such additives 
shall be used at the minimum effective concentration-in-use for the 
base fuel in question.
    (3) Unless otherwise restricted, the presence of trace contaminants 
does not preclude the use of a fuel or fuel additive as a component of 
a base fuel formulation.
    (4) When an additive is the test subject, any additive normally 
contained in the base fuel which serves the same function as the 
subject additive shall be removed from the base fuel formulation. For 
example, if a corrosion inhibitor were the subject of testing and if 
this additive were to be tested in a base fuel which normally contained 
a corrosion inhibitor, this test additive would replace the corrosion 
inhibitor normally included as a component of the base fuel.
    (5) Additive components of the methanol, ethanol, methane, and 
propane base fuels in addition to any such additives included below 
shall be limited to those recommended by the manufacturers of the 
vehicles and/or engines used in testing such fuels. For this purpose, 
EPA will review requests from manufacturers (or their agents) to modify 
the additive specifications for the alternative fuels and, if 
necessary, EPA shall change these specifications based on consistency 
of those changes with the associated vehicle manufacturer's 
recommendations for the operation of the vehicle. EPA shall publish 
notice of any such changes to a base fuel and/or its base additive 
package specifications in the Federal Register.
    (b) Gasoline Base Fuel. (1) The gasoline base fuel is patterned 
after the reformulated gasoline summer baseline fuel as specified in 
CAA section 211(k)(10)(B)(i). The specifications and blending 
tolerances for the gasoline base fuel are listed in Table F94-1. The 
additive types which shall be required and/or permissible in the 
gasoline base fuel are listed in Table 1 as well.

               Table F94-1.--Gasoline Base Fuel Properties              
                                                                        
                                                                        
                                                                        
API Gravity............................  57.40.3            
Sulfur, ppm............................  33925              
Benzene, vol%..........................  1.530.3            
RVP, psi...............................  8.70.3             
Octane, (R+M)/2........................  87.30.5            
Distillation Parameters:                                                
  10%,  deg.F..........................  1285               
  50%,  deg.F..........................  2185               
  90%,  deg.F..........................  3305               
Aromatics, vol%........................  32.02.7            
Olefins, vol%..........................  9.22.5             
Saturates, vol%........................  58.82.0            
Additive Types:                                                         
  Required.............................  Deposit Control                
                                         Corrosion Inhibitor            
                                         Demulsifier                    
                                         Anti-oxidant                   
                                         Metal Deactivator              
  Permissible..........................  Anti-static                    
                                                                        


    (2) The additive components of the gasoline base fuel shall contain 
compounds comprised of no elements other than carbon, hydrogen, oxygen, 
nitrogen, and sulfur. Additives shall be used at the minimum 
concentration needed to perform effectively in the gasoline base fuel. 
In no case shall their concentration in the base fuel exceed the 
maximum concentration recommended by the additive manufacturer. The 
increment of sulfur contributed to the formulation by any additive 
shall not exceed 15 parts per million sulfur by weight and shall not 
cause the gasoline base fuel to exceed the sulfur specifications in 
Table F94-1 of this section.
    (c) Diesel Base Fuel. (1) The diesel base fuel shall be a #2 diesel 
fuel having the properties and blending tolerances shown in Table F94-2 
of this section. The additive types which shall be permissible in 
diesel base fuel are presented in Table F94-2 as well.

                Table F94-2.--Diesel Base Fuel Properties               
                                                                        
                                                                        
                                                                        
API Gravity............................  331                
Sulfur, wt%............................  0.050.0025         
Cetane Number..........................  45.22              
Cetane Index...........................  45.72              
Distillation Parameters:                                                
  10%,  deg.F..........................  4335               
  50%,  deg.F..........................  5165               
  90%,  deg.F..........................  6065               
Aromatics, vol%........................  38.42.7            
Olefins, vol%..........................  1.50.4             
Saturates, vol%........................  60.12.0            
Additive Types:                                                         
  Required.............................  Corrosion Inhibitor            
                                         Demulsifier                    
                                         Anti-oxidant                   
                                         Metal Deactivator              
  Permitted............................  Anti-static                    
                                         Flow Improver                  
  Not Permitted........................  Deposit Control                
                                                                        


    (2) The additive components of the diesel base fuel shall contain 
compounds comprised of no elements other than carbon, hydrogen, oxygen, 
nitrogen, and sulfur. Additives shall be used at the minimum 
concentration needed to perform effectively in the diesel base fuel. In 
no case shall their concentration in the base fuel exceed the maximum 
concentration recommended by the additive manufacturer. The increment 
of sulfur contributed to the base fuel by additives shall not cause the 
diesel base fuel to exceed the sulfur specifications in Table F94-2 of 
this section.
    (d) Methanol Base Fuels. (1) The methanol base fuels shall contain 
no elements other than carbon, hydrogen, oxygen, nitrogen, sulfur, and 
chlorine.
    (2) The M100 base fuel shall consist of 100 percent by volume 
chemical grade methanol.
    (3) The M85 base fuel is to contain 85 percent by volume chemical 
grade methanol, blended with 15 percent by volume gasoline base fuel 
meeting the gasoline base fuel specifications outlined in paragraph 
(b)(1) of this section. Manufacturers shall ensure the methanol 
compatibility of lubricating oils as well as fuel additives used in the 
gasoline portion of the M85 base fuel.
    (4) The methanol base fuels shall meet the specifications listed in 
Table F94-3.

               Table F94-3.--Methanol Base Fuel Properties              
M100:                                                                   
    Chemical Grade MeOH, vol%..................................      100
    Chlorine (as chlorides), wt%, max..........................   0.0001
    Water, wt%, max............................................      0.5
    Sulfur, wt%, max...........................................    0.002
M85                                                                     
    Chemical Grade MeOH, vol%,.................................       85
    Gasoline Base Fuel, vol%...................................       15
    Chlorine (as chlorides), wt%, max..........................   0.0001
    Water, wt%, max............................................      0.5
    Sulfur, wt%, max...........................................    0.004
                                                                        

    (e) Ethanol Base Fuel. (1) The ethanol base fuel, E85, shall 
contain no elements other than carbon, hydrogen, oxygen, nitrogen, 
sulfur, chlorine, and copper.
    (2) The ethanol base fuel shall contain 85 percent by volume 
chemical grade ethanol, blended with 15 percent by volume gasoline base 
fuel that meets the specifications listed in paragraph (b)(1) of this 
section. Additives used in the gasoline component of E85 shall be 
ethanol-compatible.
    (3) The ethanol base fuel shall meet the specifications listed in 
Table F94-4.

               Table F94-4.--Ethanol Base Fuel Properties               
                                                                        
                                                                        
                                                                        
E85:                                                                    
    Chemical Grade EtOH, vol%, min.............................       85
    Gasoline Base Fuel, vol%...................................       15
    Chlorine (as chloride), wt%, max...........................   0.0004
    Copper, mg/L, max..........................................     0.07
    Water, wt%, max............................................      0.5
    Sulfur, wt%, max...........................................    0.004
                                                                        

    (f) Methane Base Fuel. (1) The methane base fuel is a gaseous motor 
vehicle fuel marketed commercially as compressed natural gas (CNG), 
whose primary constituent is methane.
    (2) The methane base fuel shall contain no elements other than 
carbon, hydrogen, oxygen, nitrogen, and sulfur. The fuel shall contain 
an odorant additive for leak detection purposes. The added odorant 
shall be used at a level such that, at ambient conditions, the fuel 
must have a distinctive odor potent enough for its presence to be 
detected down to a concentration in air of not over \1/5\ (one-fifth) 
of the lower limit of flammability. After addition of the odorant, the 
methane base fuel shall contain no more than 16 ppm sulfur by volume.
    (3) The methane base fuel shall meet the specifications listed in 
Table F94-5.

             Table F94-5.--Methane Base Fuel Specifications             
                                                                        
                                                                        
                                                                        
Methane, mole%, min..............................................   89.0
Ethane, mole%, max...............................................    4.5
Propane and higher HC, mole%, max................................    2.3
C6 and higher HC, mole%, max.....................................    0.2
Oxygen, mole%, max...............................................    0.6
Sulfur (including odorant additive) ppmv, max....................     16
Inert gases:                                                            
  Sum of CO2 and N2, mole%, max..................................   4.0 
                                                                        

    (g) Propane Base Fuel. (1) The propane base fuel is a gaseous motor 
vehicle fuel, marketed commercially as liquified petroleum gas (LPG), 
whose primary constituent is propane.
    (2) The propane base fuel may contain no elements other than 
carbon, hydrogen, oxygen, nitrogen, and sulfur. The fuel shall contain 
an odorant additive for leak detection purposes. The added odorant 
shall be used at a level such that at ambient conditions the fuel must 
have a distinctive odor potent enough for its presence to be detected 
down to a concentration in air of not over \1/5\ (one-fifth) of the 
lower limit of flammability. After addition of the odorant, the propane 
base fuel shall contain no more than 120 ppm sulfur by weight.
    (3) The propane base fuel shall meet the specifications listed in 
Table F94-6.

            Table F94--6.--Propane Base Fuel Specifications             
                                                                        
                                                                        
                                                                        
Vapor pressure at 100-F, psig, max...............................    208
Evaporative temperature, 95%,  deg.F, max........................    -37
Propane, vol%, min...............................................   92.5
Propylene, vol%, max.............................................    5.0
Butane and heavier, vol%, max....................................    2.5
Residue-evaporation of 100mL, max, mL............................   0.05
Sulfur (including odorant additive) ppmw, max....................    123
                                                                        

Sec. 79.56  Fuel and fuel additive grouping system.

    (a) Manufacturers of fuels and fuel additives are allowed to 
satisfy the testing requirements in Secs. 79.52, 79.53, and 79.54 and 
the associated reporting requirements in Sec. 79.59 on an individual or 
group basis, provided that such products meet the criteria in this 
section for enrollment in the same fuel/additive group. However, each 
manufacturer of a fuel or fuel additive must individually comply with 
the notification requirements of Sec. 79.59(b). Further, if a 
manufacturer elects to comply by participation in a group, each 
manufacturer continues to be individually subject to the information 
requirements of this subpart.
    (1) The use of the grouping provision to comply with Tier 1 and 
Tier 2 testing requirements is voluntary. No manufacturer is prohibited 
from testing and submitting its own data for its own product 
registration, despite its qualification for membership in a particular 
group.
    (2) The only groups permitted are those established in this 
section.
    (b) Each manufacturer who chooses to enroll a fuel or fuel additive 
in a group of similar fuels and fuel additives as designated in this 
section may satisfy the registration requirements through a group 
submission of jointly-sponsored testing and analysis conducted on a 
product which is representative of all products in that group, provided 
that the group representative is chosen according to the specifications 
in this section.
    (1) The health effects information submitted by a group shall be 
considered applicable to all fuels and fuel additives in the group. A 
fuel or fuel additive manufacturer who has chosen to participate in a 
group may subsequently choose to perform testing of such fuel or fuel 
additive on an individual basis; however, until such independent 
registration information has been received and reviewed by EPA, the 
information initially submitted by the group on behalf of the 
manufacturer's fuel or fuel additive shall be considered applicable and 
valid for that fuel or fuel additive. It could therefore be used to 
support requirements for further testing under the provisions of Tier 3 
or to support regulatory decisions affecting that fuel or fuel 
additive.
    (2) Manufacturers are responsible for determining the appropriate 
groups for their products according to the criteria in this section and 
for enrolling their products into those groups under industry-sponsored 
or other independent brokering arrangements.
    (3) Manufacturers who enroll a fuel or fuel additive into a group 
shall share the applicable costs according to appropriate arrangements 
established by the group. The organization and administration of group 
functions and the development of cost-sharing arrangements are the 
responsibility of the participating manufacturers. If manufacturers are 
unable to agree on fair and equitable cost sharing arrangements and if 
such dispute is referred by one or more manufacturers to EPA for 
resolution, then the provisions in Sec. 79.56(c) (1) and (2) shall 
apply.
    (c) In complying with the registration requirements for a given 
fuel or fuel additive, notwithstanding the enrollment of such fuel or 
additive in a group, a manufacturer may make use of available 
information for any product which conforms to the same grouping 
criteria as the given product. If, for this purpose, a manufacturer 
wishes to rely upon the information previously submitted by another 
manufacturer (or group of manufacturers) for registration of a similar 
product (or group of products), then the previous submitter is entitled 
to reimbursement by the manufacturer for an appropriate portion of the 
applicable costs incurred to obtain and report such information. Such 
entitlement shall remain in effect for a period of fifteen years 
following the date on which the original information was submitted. 
Pursuant to Sec. 79.59(b)(4)(ii), the manufacturer who relies on 
previously-submitted registration data shall certify to EPA that the 
original submitter has been notified and that appropriate reimbursement 
arrangements have been made.
    (1) When private efforts have failed to resolve a dispute about a 
fair amount or method of cost-sharing or reimbursement for testing 
costs incurred under this subpart, then any party involved in that 
dispute may initiate a hearing by filing two signed copies of a request 
for a hearing with a regional office of the American Arbitration 
Association and mailing a copy of the request to EPA. A copy must also 
be sent to each person from whom the filing party seeks reimbursement 
or who seeks reimbursement from that party. The information and fees to 
be included in the request for hearing are specified in 40 CFR 
791.20(b) and (c).
    (2) Additional procedures and requirements governing the hearing 
process are those specified in 40 CFR 791.22 through 791.50, 791.60, 
791.85, and 791.105, excluding 40 CFR 791.39(a)(3) and 791.48(d).
    (d) Basis for Classification. (1) Rather than segregating fuels and 
fuel additives into separate groups, the grouping system applies the 
same grouping criteria and creates a single set of groups applicable 
both to fuels and fuel additives.
    (2) Fuels shall be classified pursuant to Sec. 79.56(e) into 
categories and groups of similar fuels and fuel additives according to 
the components and characteristics of such fuels in their uncombusted 
state. The classification of a fuel product must take into account the 
components of all bulk fuel additives which are listed in the 
registration application or basic registration data submitted for the 
fuel product.
    (3) Fuel additives shall be classified pursuant to Sec. 79.56(e) 
into categories and groups of similar fuels and fuel additives 
according to the components and characteristics of the respective 
uncombusted additive/base fuel mixture pursuant to Sec. 79.51(h)(1).
    (4) In determining the category and group to which a fuel or fuel 
additive belongs, impurities present in trace amounts shall be ignored 
unless otherwise noted. Impurities are those substances which are 
present through contamination or which remain in the fuel or additive 
naturally after processing is completed.
    (5) Reference Standards. (i) American Society for Testing and 
Materials (ASTM) standard D 4814-93a, ``Standard Specification for 
Automotive Spark-Ignition Engine Fuel'', used to define the general 
characteristics of gasoline fuels (paragraph (e)(3)(i)(A)(3) of this 
section) and ASTM standard D 975-93, ``Standard Specification for 
Diesel Fuel Oils'', used to define the general characteristics of 
diesel fuels (paragraph (e)(3)(ii)(A)(3) of this section) have been 
incorporated by reference.
    (ii) This incorporation by reference was approved by the Director 
of the Federal Register in accordance with 5 U.S.C. 552(a) and 1 CFR 
part 51. Copies may be obtained from the American Society for Testing 
and Materials (ASTM), 1916 Race Street, Philadelphia, PA 19103. Copies 
may be inspected at U.S. EPA, OAR, 401 M Street SW., Washington, DC, 
20460 or at the Office of the Federal Register, 800 North
 Capitol Street NW., suite 700, Washington, DC.
    (e) Grouping Criteria. The grouping system is represented by a 
matrix of three fuel/additive categories within six specified fuel 
families (see Table F94-7, Grouping System for Fuels and Fuel 
Additives). Each category may include one or more groups. Within each 
group, a representative may be designated based on the criteria in this 
section and joint registration information may be developed and 
submitted for member fuels and fuel additives.

                                               Table F94-7.--Grouping System for Fuels and Fuel Additives                                               
--------------------------------------------------------------------------------------------------------------------------------------------------------
                              Conventional Fuel Families                                         Alternative Fuel Families                              
                     -----------------------------------------------------------------------------------------------------------------------------------
      Category                                                                                                 Methane (CNG, LNG)                       
                          Gasoline (A)           Diesel (B)           Methanol(C)            Ethanol(D)               (E)            Propane (LPG) (F)  
--------------------------------------------------------------------------------------------------------------------------------------------------------
Baseline............  One group             One group             Two groups: (1) M100  One group (includes   One group (includes   One group           
                       represented by        represented by        group (includes       ethanol-gasoline      both CNG and LNG),    represented by LPG 
                       gasoline base fuel.   diesel base fuel.     methanol-gasoline     formulations with     represented by CNG    base fuel.         
                                                                   formulations with     at least 50%          base fuel.                               
                                                                   at least 96%          ethanol)                                                       
                                                                   methanol)             represented by E85                                             
                                                                   represented by M100   base fuel.                                                     
                                                                   base fuel (2) M85                                                                    
                                                                   (includes methanol-                                                                  
                                                                   gasoline                                                                             
                                                                   formulations with                                                                    
                                                                   50-95% methanol)                                                                     
                                                                   represented by M85                                                                   
                                                                   base fuel.                                                                           
Non-baseline........  One group for each    One group for each    One group for each    One group for each    One group to include  One group to include
                       gasoline-oxygenate    oxygen-contributing   individual non-       individual non-       methane               propane            
                       blend or each         compound or class     methanol, non-        ethanol, non-         formulations          formulations       
                       gasoline-methanol/    of compounds; one     gasoline component    gasoline component    exceeding the         exceeding the      
                       co-solvent blend;     group for each        and one group for     and one group for     specified limit for   specified limit for
                       one group for each    synthetic crude-      each unique           each unique           non-methane           butane and higher  
                       synthetic crude-      derived fuel.         combination of such   combination of such   hydrocarbons.         hydrocarbons.      
                       derived fuel.                               components.           components.                                                    
Atypical............  One group for each    One group for each    One group for each    One group for each    One group for each    One group for each  
                       atypical element/     atypical element/     atypical element/     atypical element/     atypical element/     atypical element/  
                       characteristic, or    characteristic, or    characteristic, or    characteristic, or    characteristic, or    characteristic, or 
                       unique combination    unique combination    unique combination    unique combination    unique combination    unique combination 
                       of atypical           of atypical           of atypical           of atypical           of atypical           of atypical        
                       elements/characteri   elements/characteri   elements/characteri   elements/characteri   elements/characteri   elements/characteri
                       stics.                stics.                stics.                stics.                stics.                stics.             
--------------------------------------------------------------------------------------------------------------------------------------------------------

    (1) Fuel Families. Each of the following six fuel families (Table 
F94-7, columns A-F) includes fuels of the type referenced in the name 
of the family as well as bulk and aftermarket additives which are 
intended for use in those fuels. When applied to fuel additives, the 
criteria in these descriptions refer to the associated additive/base 
fuel mixture, pursuant to Sec. 79.51(h)(1). One or more base fuel 
formulations are specified for each fuel family pursuant to Sec. 79.55.
    (i) The Gasoline Family includes fuels composed of more than 50 
percent gasoline by volume and their associated fuel additives. The 
base fuel for this family is specified in Sec. 79.55(b).
    (ii) The Diesel Family includes fuels composed of more than 50 
percent diesel fuel by volume and their associated fuel additives. The 
Diesel fuel family includes both Diesel #1 and Diesel #2 formulations. 
The base fuel for this family is specified in Sec. 79.55(c).
    (iii) The Methanol Family includes fuels composed of at least 50 
percent methanol by volume and their associated fuel additives. The 
M100 and M85 base fuels are specified in Sec. 79.55(d).
    (iv) The Ethanol Family includes fuels composed of at least 50 
percent ethanol by volume and their associated fuel additives. The base 
fuel for this family is E85 as specified in Sec. 79.55(e).
    (v) The Methane Family includes compressed natural gas (CNG) and 
liquefied natural gas (LNG) fuels containing at least 50 mole percent 
methane and their associated fuel additives. The base fuel for the 
family is a CNG formulation specified in Sec. 79.55(f).
    (vi) The Propane Family includes propane fuels containing at least 
50 percent propane by volume and their associated fuel additives. The 
base fuel for this family is a liquefied petroleum gas (LPG) as 
specified in Sec. 79.55(g).
    (vii) A manufacturer seeking registration for formulation(s) which 
do not fit the criteria for inclusion in any of the fuel families 
described in this section shall contact EPA at the address in 
Sec. 79.59(a)(1) for further guidance in classifying and testing such 
formulation(s).
    (2) Fuel/Additive Categories. Fuel/additive categories (Table F94-
7, rows 1-3) are subdivisions of fuel families which represent the 
degree to which fuels and fuel additives in the family resemble the 
base fuel(s) designated for the family. Three general category types 
are defined in this section. When applied to fuel additives, the 
criteria in these descriptions refer to the associated additive/base 
fuel mixture, pursuant to Sec. 79.51(h)(1).
    (i) Baseline categories consist of fuels and fuel additives which 
contain no elements other than those permitted in the base fuel for the 
respective fuel family and conform to specified limitations on the 
amounts of certain components or characteristics applicable to that 
fuel family.
    (ii) Non-Baseline Categories consist of fuels and fuel additives 
which contain no elements other than those permitted in the base fuel 
for the respective fuel family, but which exceed one or more of the 
limitations for certain specified components or characteristics 
applicable to baseline formulations in that fuel family.
    (iii) Atypical Categories consist of fuels and fuel additives which 
contain elements or classes of compounds other than those permitted in 
the base fuel for the respective fuel family or which otherwise do not 
meet the criteria for either baseline or non-baseline formulations in 
that fuel family. A fuel or fuel additive product having both non-
baseline and atypical characteristics pursuant to Sec. 79.56(e)(3), 
shall be considered to be an atypical product.
    (3) This section defines the specific categories applicable to each 
fuel family. When applied to fuel additives, the criteria in these 
descriptions refer to the associated additive/base fuel mixture, 
pursuant to Sec. 79.51(h)(1).
    (i) Gasoline Categories. (A) The Baseline Gasoline category 
contains gasoline fuels and associated additives which satisfy all of 
the following criteria:
    (1) Contain no elements other than carbon, hydrogen, oxygen, 
nitrogen, and/or sulfur.
    (2) Contain less than 1.5 percent oxygen by weight.
    (3) Sulfur concentration is limited to 1000 ppm per the 
specifications cited in the following paragraph.
    (4) Possess the physical and chemical characteristics of unleaded 
gasoline as specified by ASTM standard D 4814-93a (incorporated by 
reference, pursuant to paragraph (d)(5) of this section), in at least 
one Seasonal and Geographical Volatility Class.
    (5) Derived from conventional petroleum sources only.
    (B) The Non-Baseline Gasoline category is comprised of gasoline 
fuels and associated additives which conform to the specifications in 
paragraph (e)(3)(i)(A) of this section for the Baseline Gasoline 
category except that they contain 1.5 percent or more oxygen by weight 
and/or may be derived from synthetic crudes, such as those prepared 
from coal, shale and tar sands, heavy oil deposits, and other non-
conventional petroleum sources.
    (C) The Atypical Gasoline category is comprised of gasoline fuels 
and associated additives which contain one or more elements other than 
carbon, hydrogen, oxygen, nitrogen, and sulfur.
    (ii) Diesel Categories. (A) The Baseline Diesel category is 
comprised of diesel fuels and associated additives which satisfy all of 
the following criteria:
    (1) Contain no elements other than carbon, hydrogen, oxygen, 
nitrogen, and/or sulfur. Pursuant to 40 CFR 80.29, highway diesel sold 
after October 1, 1993 shall contain 0.05 percent or less sulfur by 
weight;
    (2) Contain less than 1.0 percent oxygen by weight;
    (3) Diesel formulations containing more than 0.05 percent sulfur by 
weight are precluded by 40 CFR 80.29;
    (4) Possess the characteristics of diesel fuel as specified by ASTM 
standard D 975-93 (incorporated by reference, pursuant to paragraph 
(d)(5) of this section); and
    (5) Derived from conventional petroleum sources only.
    (B) The Non-Baseline Diesel category is comprised of diesel fuels 
and associated additives which conform to the specifications in 
paragraph (e)(3)(ii)(A) of this section for the Baseline Diesel 
category except that they contain 1.0 percent or more oxygen by weight 
and/or may be derived from synthetic crudes, such as those prepared 
from coal, shale and tar sands, heavy oil deposits, and other non-
conventional petroleum sources.
    (C) The Atypical Diesel category is comprised of diesel fuels and 
associated additives which contain one or more elements other than 
carbon, hydrogen, oxygen, nitrogen, and sulfur.
    (iii) Methanol Categories. (A) The Baseline Methanol category is 
comprised of methanol fuels and associated additives which contain at 
least 50 percent methanol by volume, no more than 4.0 percent by volume 
of substances other than methanol and gasoline, and no elements other 
than carbon, hydrogen, oxygen, nitrogen, sulfur, and/or chlorine. 
Baseline methanol shall contain no more than 0.004 percent by weight of 
sulfur or 0.0001 percent by weight of chlorine.
    (B) The Non-Baseline Methanol category is comprised of fuel blends 
which contain at least 50 percent methanol by volume, more than 4.0 
percent by volume of a substance(s) other than methanol and gasoline, 
and meet the baseline limitations on elemental composition in paragraph 
(e)(3)(iii)(A) of this section.
    (C) The Atypical Methanol category consists of methanol fuels and 
associated additives which do not meet the criteria for either the 
Baseline or the Non-Baseline Methanol category.
    (iv) Ethanol Categories. (A) The Baseline Ethanol category is 
comprised of ethanol fuels and associated additives which contain at 
least 50 percent ethanol by volume, no more than five (5) percent by 
volume of substances other than ethanol and gasoline, and no elements 
other than carbon, hydrogen, oxygen, nitrogen, sulfur, chlorine, and 
copper. Baseline ethanol formulations shall contain no more than 0.004 
percent by weight of sulfur, 0.0004 percent by weight of chlorine, and/
or 0.07 mg/L of copper.
    (B) The Non-Baseline Ethanol category is comprised of fuel blends 
which contain at least 50 percent ethanol by volume, more than five (5) 
percent by volume of a substance(s) other than ethanol and gasoline, 
and meet the baseline limitations on elemental composition in paragraph 
(e)(3)(iv)(A) of this section.
    (C) The Atypical Ethanol category consists of ethanol fuels and 
associated additives which do not meet the criteria for either the 
Baseline or the Non-Baseline Ethanol categories.
    (v) Methane Categories. (A) The Baseline Methane category is 
comprised of methane fuels and associated additives (including at least 
an odorant additive) which contain no elements other than carbon, 
hydrogen, oxygen, nitrogen, and/or sulfur, and contain no more than 20 
mole percent non-methane hydrocarbons. Baseline methane formulations 
shall not contain more than 16 ppm by volume of sulfur, including any 
sulfur which may be contributed by the odorant additive.
    (B) The Non-Baseline Methane category consists of methane fuels and 
associated additives which conform to the specifications in paragraph 
(e)(3)(v)(A) of this section for the Baseline Methane category except 
that they exceed 20 mole percent non-methane hydrocarbons.
    (C) The Atypical Methane category consists of methane fuels and 
associated additives which contain one or more elements other than 
carbon, hydrogen, oxygen, nitrogen, and/or sulfur, or exceed 16 ppm by 
volume of sulfur.
    (vi) Propane Categories. (A) The Baseline Propane category is 
comprised of propane fuels and associated additives (including at least 
an odorant additive) which contain no elements other than carbon, 
hydrogen, oxygen, nitrogen, and/or sulfur, and contain no more than 20 
percent by volume non-propane hydrocarbons. Baseline Propane 
formulations shall not contain more than 123 ppm by weight of sulfur, 
including any sulfur which may be contributed by the odorant additive.
    (B) The Non-Baseline Propane category consists of propane fuels and 
associated additives which conform to the specifications in paragraph 
(e)(3)(vi)(A) of this section for the Baseline Propane category, except 
that they exceed the 20 percent by volume limit for butane and higher 
hydrocarbons.
    (C) The Atypical Propane category consists of propane fuels and 
associated additives which contain elements other than carbon, 
hydrogen, oxygen, nitrogen, and/or sulfur, or exceed 123 ppm by weight 
of sulfur.
    (4) Fuel/Additive Groups. Fuel/additive groups are subdivisions of 
the fuel/additive categories. One or more group(s) are defined within 
each category in each fuel family according to the presence of 
differing characteristics in the fuel or additive/base fuel mixture. 
For each group, one formulation (either a base fuel or a member fuel or 
additive product) is chosen to represent all the member products in the 
group in any tests required under this subpart. The section which 
follows describes the fuel/additive groups.
    (i) Baseline Groups. (A) The Baseline Gasoline category comprises a 
single group. The gasoline base fuel specified in Sec. 79.55(b) shall 
serve as the representative of this group.
    (B) The Baseline Diesel category comprises a single group. The 
diesel base fuel specified in Sec. 79.55(c) shall serve as the 
representative of this group.
    (C) The Baseline Methanol category includes two groups: M100 and 
M85. The M100 group consists of methanol-gasoline formulations 
containing at least 96 percent methanol by volume. These formulations 
must contain odorants and bitterants (limited in elemental composition 
to carbon, hydrogen, oxygen, nitrogen, sulfur, and chlorine) for 
prevention of purposeful or inadvertent consumption. The M100 base fuel 
specified in Sec. 79.55(d) shall serve as the representative for this 
group. The M85 group consists of methanol-gasoline formulations 
containing at least 50 percent by volume but less than 96 percent by 
volume methanol. The M85 base fuel specified in Sec. 79.55(d) shall 
serve as the representative of this group.
    (D) The Baseline Ethanol category comprises a single group. The E85 
base fuel specified in Sec. 79.55(e) shall serve as the representative 
of this group.
    (E) The Baseline Methane category comprises a single group. The CNG 
base fuel specified in Sec. 79.55(f) shall serve as the representative 
of this group.
    (F) The Baseline Propane category comprises a single group. The LPG 
base fuel specified in Sec. 79.55(g) shall serve as the representative 
of this group.
    (ii) Non-Baseline Groups-- (A) Non-Baseline Gasoline. The Non-
Baseline gasoline fuels and associated additives shall sort into groups 
according to the following criteria:
    (1) For gasoline fuel and additive products which contain 1.5 
percent oxygen by weight or more, a separate non-baseline gasoline 
group shall be defined by each oxygenate compound or methanol/co-
solvent blend listed as a component in the registration application or 
basic registration data of any such fuel or additive.
    (i) Examples of oxygenates occurring in non-baseline gasoline 
formulations include ethanol, methyl tertiary butyl ether (MTBE), ethyl 
tertiary butyl ether (ETBE), tertiary amyl methyl ether (TAME), 
diisopropyl ether (DIPE), dimethyl ether (DME), tertiary amyl ethyl 
ether (TAEE), and any other compound(s) which increase the oxygen 
content of the gasoline formulation. A separate non-baseline gasoline 
group is defined for each such oxygenating compound.
    (ii) Each unique methanol and co-solvent combination (whether one, 
two, or more additional oxygenate compounds) used in a non-baseline 
fuel shall also define a separate group. An oxygenate compound used as 
a co-solvent for methanol in a non-baseline gasoline formulation must 
be identified as such in its registration. If the oxygenate is not 
identified as a methanol co-solvent, then the compound shall be 
regarded by EPA as defining a separate non-baseline gasoline group. 
Examples of methanol/co-solvent combinations occurring in non-baseline 
gasoline formulations include methanol/isopropyl alcohol, methanol/
butanol, and methanol with alcohols up to C8/octanol (Octamix).
    (iii) For each such group, the representative to be used in testing 
shall be a formulation consisting of the gasoline base fuel blended 
with the relevant oxygenate compound (or methanol/co-solvent 
combination) in an amount equivalent to the highest actual or 
recommended concentration-in-use of the oxygenate (or methanol/co-
solvent combination) recorded in the basic registration data of any 
member fuel or additive product. In the event that two or more products 
in the same group contain the same and highest amount of the oxygenate 
or methanol/co-solvent blend, then the representative shall be chosen 
at random for such candidate products.
    (2) An oxygenate compound or methanol/co-solvent combination to be 
blended with the gasoline base fuel for testing purposes shall be 
chemical-grade quality, at a minimum, and shall not contain a 
significant amount of other contaminating oxygenate compounds.
    (3) Separate non-baseline gasoline groups shall also be defined for 
gasoline formulations derived from each particular non-conventional 
petroleum source or process.
    (i) Such groups may include, but are not limited to, the following: 
coal-derived gasoline formulations; chemically-synthesized gasoline 
formulations (including those using recycled chemical/petrochemical 
products); tar sand-derived gasoline formulations; shale-derived 
gasoline formulations; and other types of soil-recovered products used 
in formulating gasolines.
    (ii) In any such group, the first product to be registered or to 
apply for EPA registration shall be the representative of that group. 
If two or more such products are registered or apply for first 
registration simultaneously, then the representative shall be chosen by 
a random method from among such candidate products.
    (4) Pursuant to Sec. 79.51(i), non-baseline gasoline products may 
belong to more than one fuel/additive group.
    (B) Non-Baseline Diesel. The Non-Baseline diesel fuels and 
associated additives shall sort into groups according to the following 
criteria:
    (1) For diesel fuel and additive products which contain 1.0 percent 
oxygen by weight or more, a separate non-baseline diesel group shall be 
defined by each individual alcohol or ether listed as a component in 
the registration application or basic registration data of any such 
fuel or additive. For each such group, the representative to be used in 
testing shall be a formulation consisting of the diesel base fuel 
blended with the relevant alcohol or ether in an amount equivalent to 
the highest actual or recommended concentration-in-use of the alcohol 
or ether recorded in the basic registration data of any member fuel or 
additive product.
    (2) A separate non-baseline diesel group is also defined for each 
of the following classes of oxygenating compounds: mixed nitroso- 
compounds; mixed nitro- compounds; mixed alkyl nitrates; mixed alkyl 
nitrites; peroxides; furans; mixed alkyl esters of plant origin; and 
mixed alkyl esters of animal origin. For each such group, the 
representative to be used in testing shall be formulated as follows:
    (i) From the class of compounds which defines the group, a 
particular oxygenate compound shall be chosen from among all such 
compounds recorded in the registration application or basic 
registration data of any fuel or additive in the group.
    (ii) The selected compound shall be the one recorded in any member 
product's registration application with the highest actual or 
recommended maximum concentration-in-use. This compound, when mixed 
into the diesel base fuel at the indicated maximum concentration, shall 
serve as the group representative.
    (iii) In the event that two or more oxygenate compounds in the 
relevant class have the highest recorded concentration-in-use, then the 
oxygenate compound to be used in the group representative shall be 
chosen at random from the qualifying candidate compounds.
    (3) A separate non-baseline diesel group shall also be defined for 
each diesel fuel derived from a particular synthetic petroleum source 
or process.
    (i) Such groups include, but shall not be limited to, the 
following: coal-derived diesel formulations; chemically-synthesized 
diesel formulations (including those using recycled chemical/
petrochemical products); tar sand-derived diesel formulations; shale-
derived diesel formulations; and other types of soil-recovered products 
used in formulating diesel fuel(s).
    (ii) In any such group, the first product to be registered or to 
apply for EPA registration shall be the representative of that group. 
If two or more products are registered or apply for first registration 
simultaneously, then the representative shall be chosen by a random 
method from among such candidate products.
    (4) Pursuant to Sec. 79.51(i), non-baseline diesel products may 
belong to more than one fuel/additive group.
    (C) Non-Baseline Methanol. The Non-Baseline methanol formulations 
are sorted into groups based on the non-methanol, non-gasoline 
component(s) of the blended fuel. Each such component occurring 
separately and each unique combination of such components shall define 
a separate group.
    (1) The representative of each such non-baseline methanol group 
shall be the group member with the highest percent by volume of non-
methanol, non-gasoline component(s).
    (2) In case two or more such members have the same and highest 
concentration of non-methanol, non-gasoline component(s), the 
representative of the group shall be chosen at random from among such 
equivalent member products.
    (D) Non-Baseline Ethanol. The Non-Baseline ethanol formulations are 
sorted into groups based on the non-ethanol, non-gasoline component(s) 
of the blended fuel. Each such component occurring separately and each 
unique combination of such components shall define a separate group.
    (1) The representative of each such non-baseline ethanol group 
shall be the group member with the highest percent by volume of non-
ethanol, non-gasoline component(s).
    (2) In case two or more such members have the same and highest 
concentration of non-ethanol, non-gasoline component(s), the 
representative of the group shall be chosen at random from among such 
equivalent member products.
    (E) Non-Baseline Methane. The Non-Baseline methane category 
consists of one group. The group representative shall be the member 
fuel or fuel/additive formulation containing the highest concentration-
in-use of non-methane hydrocarbons. If two or more member products have 
the same and the highest concentration-in-use, then the representative 
shall be chosen at random from such products.
    (F) Non-Baseline Propane. The Non-Baseline propane category 
consists of one group. The group representative shall be the member 
fuel or fuel/additive formulation containing the highest concentration-
in-use of butane and higher hydrocarbons. If two or more products have 
the same and the highest concentration-in-use, then the representative 
shall be chosen at random from such products.
    (iii) Atypical groups.
    (A) As defined for each individual fuel family in Sec. 79.56(e)(3), 
fuels and additives meeting any one of the following criteria are 
considered atypical.
    (1) Gasoline Atypical fuels and additives contain one or more 
elements in addition to carbon, hydrogen, oxygen, nitrogen, and sulfur.
    (2) Diesel Atypical fuels and additives contain one or more element 
in addition to carbon, hydrogen, oxygen, nitrogen, and sulfur.
    (3) Methanol Atypical fuels and additives contain:
    (i) one or more element in addition to carbon, hydrogen, oxygen, 
nitrogen, sulfur, and chlorine, and/or
    (ii) sulfur in excess of 0.004 percent by weight, and/or
    (iii) chlorine in excess of 0.0001 percent by weight.
    (4) Ethanol Atypical fuels and additives contain:
    (i) one or more element in addition to carbon, hydrogen, oxygen, 
nitrogen, sulfur, chlorine, and copper, and/or
    (ii) sulfur in excess of 0.004 percent by weight, and/or
    (iii) contain chlorine (as chloride) in excess of 0.0004 percent by 
weight, and/or
    (iv) contain copper in excess of 0.07 mg/L.
    (5) Methane Atypical fuels and additives contain:
    (i) one or more element in addition to carbon, hydrogen, oxygen, 
nitrogen, and sulfur, and/or
    (ii) sulfur in excess of 16 ppm by volume.
    (6) Propane Atypical fuels and additives contain:
    (i) one or more element in addition to carbon, hydrogen, oxygen, 
nitrogen, and sulfur, and/or
    (ii) sulfur in excess of 123 ppm by weight.
    (B) General rules for sorting these atypical fuels and additives 
into separate groups are as follows:
    (1) Pursuant to Sec. 79.51(j), a given atypical product may belong 
to more than one atypical group.
    (2) Fuels and additives in different fuel families may not be 
grouped together, even if they contain the same atypical element(s) or 
other atypical characteristic(s).
    (3) A fuel or additive containing one or more atypical elements 
attached to a polymer compound must be sorted into a separate group 
from atypical fuels or fuel additives containing the same atypical 
element(s) in non-polymer form. However, the occurrence of a polymer 
compound which does not contain an atypical element does not affect the 
grouping of a fuel or additive.
    (C) Specific rules for sorting each family's atypical fuels and 
additives into separate groups, and for choosing each such group's 
representative for testing, are as follows:
    (1) A separate group is created for each atypical element (or other 
atypical characteristic) occurring separately, i.e., in the absence of 
any other atypical element or characteristic, in one or more fuels and/
or additives within a given fuel family.
    (i) Consistent with the basic grouping guidelines provided in 
Sec. 79.56(d), a fuel product which is classified as atypical because 
its basic registration data or application lists a bulk additive 
containing an atypical characteristic, may be grouped with that 
additive and/or with other fuels and additives containing the same 
atypical characteristic.
    (ii) Within a group of products containing only one atypical 
element or characteristic, the fuel or additive/base fuel mixture with 
the highest concentration-in-use or recommended concentration-in-use of 
the atypical element or characteristic shall be the designated 
representative of that group. In the event that two or more fuels or 
additive/base fuel mixtures within the group contain the same and 
highest concentration of the single atypical element or characteristic, 
then the group representative shall be selected by a random method from 
among such candidate products.
    (2) A separate group is also created for each unique combination of 
atypical elements (and/or other specified atypical characteristics) 
occurring together in one or more fuels and/or additives within a given 
fuel family.
    (i) Consistent with the basic grouping guidelines provided in 
Sec. 79.56(d), a fuel which is classified as atypical because its basic 
registration data lists one bulk additive containing two or more 
atypical characteristics, may be grouped with that additive and/or with 
other fuels and/or additives containing the same combination of 
atypical characteristics. Grouping of fuels containing more than one 
atypical additive shall be guided by provisions of Sec. 79.51(j).
    (ii) Within a group of such products containing a unique 
combination of two or more atypical elements or characteristics, the 
designated representative shall be the product within the group which 
contains the highest total concentration of the atypical elements or 
characteristics.
    (iii) In the event that two or more products within a given 
atypical group contain the same and highest concentration of the same 
atypical elements or characteristics then, among such candidate 
products, the designated representative shall be the product which, 
first, has the highest total concentration of metals, followed in order 
by highest total concentration of halogens, highest total concentration 
of other atypical elements (including sulfur concentration, as 
applicable), highest total concentration of polymers containing 
atypical elements, and, lastly, highest total concentration of oxygen.
    (iv) If two or more products have the same and highest 
concentration of the variable identified in the preceding paragraph, 
then, among such products, the one with the greatest concentration of 
the next highest variable on the list shall be the group 
representative.
    (v) This decision-making process shall continue until a single 
product is determined to be the representative. If two or more products 
remain tied at the end of this process, then the representative shall 
be chosen by a random method from among such remaining products.


Sec. 79.57.  Emission generation.

    This section specifies the equipment and procedures that must be 
used in generating the emissions which are to be subjected to the 
characterization procedures and/or the biological tests specified in 
Secs. 79.52(b) and 79.53 of these regulations. When applicable, they 
may also be required in conjunction with testing under Secs. 79.54 and 
79.58(c). Additional requirements concerning emission generation, 
delivery, dilution, quality control, and safety practices are outlined 
in Sec. 79.61.
    (a) Vehicle and engine selection criteria. (1) All vehicles and 
engines used to generate emissions for testing a fuel or additive/fuel 
mixture must be new (i.e., never before titled) and placed into the 
program with less than 500 miles on the odometer or 12 hours on the 
engine chronometer. The vehicles and engines shall be unaltered from 
the specifications of the original equipment manufacturer.
    (2) The vehicle/engine type, vehicle/engine class, and vehicle/
engine subclass designated to generate emissions for a given fuel or 
additive shall be the same type, class, and subclass which, over the 
previous three years, has consumed the most gallons of fuel in the fuel 
family applicable to the given fuel or additive. No distinction shall 
be made between light-duty vehicles and light-duty trucks for purposes 
of this classification.
    (3) Within this vehicle/engine type, class, and subclass, the 
specific vehicles and engines acceptable for emission generation are 
those that represent the most common fuel metering system and the most 
common of the most important emission control system devices or 
characteristics with respect to emission reduction performance for the 
model year in which testing begins. These vehicles will be determined 
through a survey of the previous model year's vehicle/engine sales 
within the given subclass. These characteristics shall include, but 
need not be limited to, aftertreatment device(s), fuel aspiration, air 
injection, exhaust gas recirculation, and feedback type.
    (4) Within the applicable subclass, the five highest selling 
vehicle/engine models that contain the most common such equipment and 
characteristics shall be determined. Any of these five models of the 
current model year (at the time testing begins) may be selected for 
emission generation.
    (i) If one or more of the five models is not available for the 
current model year, the choice of model for emission generation shall 
be limited to those remaining among the five.
    (ii) If fewer than five models of the given vehicle/engine type are 
available for the current model year, all such models shall be 
eligible.
    (5) When the fuel or fuel additive undergoing testing is not 
commonly used or intended to be used in the vehicle/engine types 
prescribed by this selection procedure, or when rebuilding or 
alteration is required to obtain a suitable vehicle/engine for emission 
generation, the manufacturer may submit a request to EPA for a 
modification in test procedure requirements. Any such request must 
include objective test results which support the claim that a more 
appropriate vehicle/engine type is needed as well as a suggested 
substitute vehicle/engine type. The vehicle/engine selection in this 
case shall be approved by EPA prior to the start of testing.
    (6) Once a particular model has been chosen on which to test a fuel 
or additive product, all mileage accumulation and generation of 
emissions for characterization and biological testing of such product 
shall be conducted on that same model.
    (i) If the initial test vehicle/engine fails or must be replaced 
for any reason, emission generation shall continue with a second 
vehicle/engine which is identical to, or resembles to the greatest 
extent possible, the initial test vehicle/engine. If more than one 
replacement vehicle/engine is necessary, all such vehicles/engines 
shall be identical, or resemble to the greatest extent possible, the 
initial test vehicle/engine.
    (ii) Manufacturers are encouraged to obtain, at the start of a test 
program, more than one emission generation vehicle/engine of the 
identical model, to ensure the availability of back-up emission 
generator(s). All backup vehicles/engines must be conditioned and must 
have their emissions fully characterized, as done for the initial test 
vehicle/engine, prior to their use as emission generators for 
biological testing. Alternating between such vehicles/engines regularly 
during the course of testing is permissible and advisable, particularly 
to allow regular maintenance on such vehicles/engines during prolonged 
health effects testing.
    (b) Vehicle/engine operation and maintenance. (1) For the purpose 
of generating combustion emissions from a fuel or additive/base fuel 
mixture for which the relevant class is light duty, either a light-duty 
vehicle shall be operated on a chassis dynamometer or a light-duty 
engine shall be operated on an engine dynamometer. When the relevant 
class is heavy duty, the emissions shall be generated on a heavy-duty 
engine operated on an engine dynamometer. In both cases, the vehicle or 
engine model shall be selected as described in paragraph (a) of this 
section and shall have all applicable fuel and emission control systems 
intact.
    (2) Except as provided in Sec. 79.51(h)(2)(iii), the fuel or 
additive/base fuel mixture being tested shall be used at all times 
during operation of the test vehicle or engine. No other fuels or 
additives shall be used in the test vehicle or engine once mileage 
accumulation has begun until emission generation for emission 
characterization and biological testing purposes is completed.
    (3) Scheduled and unscheduled vehicle/engine maintenance.
    (i) During emission generation, vehicles and engines must be 
maintained in good condition by following the recommendations of the 
original equipment manufacturer (OEM) for scheduled service and parts 
replacement, with repairs performed only as necessary. Modifications, 
adjustments, and maintenance procedures contrary to procedures found in 
40 CFR part 86 for the maintenance of test vehicles/engines or 
performed solely for the purpose of emissions improvement are not 
allowed.
    (ii) If unscheduled maintenance becomes necessary, the vehicle or 
engine must be repaired to OEM specifications, using OEM or OEM-
approved parts. In addition, the tester is required to measure the 
basic emissions pursuant to Sec. 79.52(b)(2)(i) after the unscheduled 
maintenance and before resuming testing to ensure that the post-
maintenance emissions shall be within 20 percent of pre-maintenance 
emissions levels. If the basic emissions cannot be brought within 20 
percent of their previous levels, then the manufacturer shall restart 
the emissions characterization and health testing of its products 
combustion emissions using a new vehicle/engine.
    (c) Mileage accumulation. (1) A vehicle/engine break-in period is 
required prior to generating emissions for characterization and/or 
biological testing under this subpart. The required mileage 
accumulation may be accomplished on a test track, on the street, on a 
dynamometer, or using any other conventionally accepted method.
    (2) Vehicles to be used in the evaluation of baseline and non-
baseline fuels and fuel additives shall accumulate 4,000 miles prior to 
emission testing. Engines to be used in the evaluation of baseline and 
non-baseline fuels and fuel additives shall accumulate 125 hours of 
operation on an engine dynamometer prior to emission testing.
    (3) When the test formulation is classified as an atypical fuel or 
fuel additive formulation (pursuant to definitions in 
Sec. 79.56(e)(4)(iii)), the following additional mileage accumulation 
requirements apply:
    (i) The test vehicle/engine must be operated for a minimum of 4,000 
vehicle miles or 125 hours of engine operation.
    (ii) Thereafter, at intervals determined by the tester, all 
emission fractions (i.e., vapor, semi-volatile, and particulate) shall 
be sampled and analyzed for the presence and amount of the atypical 
element(s) and/or other atypical constituents. Pursuant to paragraph 
(d) of this section, the sampled emissions must be generated in the 
absence of an intact aftertreatment device. Immediately before the 
samples are taken, a brief warmup period (at least ten miles or the 
engine equivalent) is required.
    (iii) Mileage accumulation shall continue until either 50 percent 
or more of the mass of each atypical element (or other atypical 
constituent) entering the engine can be measured in the exhaust 
emissions (all fractions combined), or the vehicle/engine has 
accumulated mileage (or hours) equivalent to 40 percent of the average 
useful life of the applicable vehicle/engine class (pursuant to 
regulations in 40 CFR part 86). For example, the maximum mileage 
required for light-duty vehicles is 40 percent of 100,000 miles (i.e., 
40,000 miles), while the maximum time of operation for heavy-duty 
engines is the equivalent of 40 percent of 290,000 miles (i.e., the 
equivalent in engine hours of 116,000 miles).
    (iv) When either condition in paragraph (c)(3)(iii) of this section 
has been reached, additional emission characterization and biological 
testing of the emissions may begin.
    (d) Use of exhaust aftertreatment devices. (1) If the selected test 
vehicle/engine, as certified by EPA, does not come equipped with an 
emissions aftertreatment device (such as a catalyst or particulate 
trap), such device shall not be used in the context of this program.
    (2) Except as provided in paragraph (d)(3) of this section for 
certain specialized additives, the following provisions apply when the 
test vehicle/engine, as certified by EPA, comes equipped with an 
emissions aftertreatment device.
    (i) For mileage accumulation:
    (A) When the test formulation does not contain any atypical 
elements (pursuant to definitions in Sec. 79.56(e)(4)(iii)), an intact 
aftertreatment device must be used during mileage accumulation.
    (B) When the test formulation does contain atypical elements, then 
the manufacturer may choose to accumulate the required mileage using a 
vehicle/engine equipped with either an intact aftertreatment device or 
with a non-functional aftertreatment device (e.g., a blank catalyst 
without its catalytic wash coat). In either case, sampling and analysis 
of emissions for measurement of the mass of the atypical element(s) (as 
described in Sec. 79.57(c)(3)) must be done on emissions generated with 
a non-functional (blank) aftertreatment device.
    (1) If the manufacturer chooses to accumulate mileage without a 
functional aftertreatment device, and if the manufacturer wishes to do 
this outside of a laboratory/test track setting, then a memorandum of 
exemption for product testing must be obtained by applying to the 
Director of the Field Operations and Support Division (see 
Sec. 79.59(a)(1)).
    (2) [reserved]
    (ii) For Tier 1 (Sec. 79.52), the total set of requirements for the 
characterization of combustion emissions (Sec. 79.52(b)) must be 
completed two times, once using emissions generated with the 
aftertreatment device intact and a second time with the aftertreatment 
device rendered nonfunctional or replaced with a non-functional 
aftertreatment device as described in paragraph (d)(2)(i)(B) of this 
section.
    (iii) For Tier 2 (Sec. 79.53), the standard requirements for 
biological testing of combustion emissions shall be conducted using 
emissions generated with a non-functioning aftertreatment device as 
described in paragraph (d)(2)(i)(B) of this section.
    (iv) For alternative Tier 2 requirements (Sec. 79.58(c)) or Tier 3 
requirements (Sec. 79.54) which may be prescribed by EPA, the use of 
functional or nonfunctional aftertreatment devices shall be specified 
by EPA as part of the test guidelines.
    (v) In the case where an intact aftertreatment device is not in 
place, all other manufacturer-specified combustion characteristics 
(e.g., back pressure, residence time, and mixing characteristics) of 
the altered vehicle/engine shall be retained to the greatest extent 
possible.
    (3) Notwithstanding paragraphs (d)(1) and (d)(2) of this section, 
when the subject of testing is a fuel additive specifically intended to 
enhance the effectiveness of exhaust aftertreatment devices, the 
related aftertreatment device may be used on the emission generation 
vehicle/engine during all mileage accumulation and testing.
    (e) Generation of combustion emissions--
    (1) Generating combustion emissions for emission characterization. 
(i) Combustion emissions shall be generated according to the exhaust 
emission portion of the Federal Test Procedure (FTP) for the 
certification of new motor vehicles, found in 40 CFR part 86, subpart B 
for light-duty vehicles/engines, and subparts D, M and N for heavy-duty 
vehicles/engines. The Urban Dynamometer Driving Schedule (UDDS), 
pursuant to 40 CFR part 86, appendix I(a), shall apply to light-duty 
vehicles/engines and the Engine Dynamometer Driving Schedule (EDS), 
pursuant to 40 CFR part 86, appendix I(f)(2), shall apply to heavy-duty 
vehicles/engines. The motoring portion of the heavy-duty test cycle may 
be eliminated, at the manufacturer's option, for the generation of 
emissions.
    (A) For light-duty engines operated on an engine dynamometer, the 
tester shall determine the speed-torque equivalencies (``trace'') for 
its test engine from valid FTP testing performed on a chassis 
dynamometer, using a test vehicle with an engine identical to that 
being tested. The test engine must then be operated under these speed 
and torque specifications to simulate the FTP cycle.
    (B) Special procedures not included in the FTP may be necessary in 
order to characterize emissions from fuels and fuel additives 
containing atypical elements or to collect some types of emissions 
(e.g., particulate emissions from light-duty vehicles/engines, semi-
volatile emissions from both light-duty and heavy-duty vehicles/
engines). Such alterations to the FTP are acceptable.
    (ii) Pursuant to Sec. 79.52(b)(1)(i) and Sec. 79.57(d)(2)(ii), 
emission generation and characterization must be repeated three times 
when the selected vehicle/engine is normally operated without an 
emissions aftertreatment device and six times when the selected 
vehicle/engine is normally operated with an emissions aftertreatment 
device. In the latter case, the emission generation and 
characterization process shall be repeated three times with the intact 
aftertreatment device in place and three times with a non-functioning 
(blank) aftertreatment device in place.
    (iii) From both light-duty and heavy-duty vehicles/engines, samples 
of vapor phase, semi-volatile phase, and particulate phase emissions 
shall be collected, except that semi-volatile phase, and particulate 
emissions need not be sampled for fuels and additives in the methane 
and propane families (pursuant to Sec. 79.56(e)(1)(v) and (vi)). The 
number and type of samples to be collected and separately analyzed 
during one emission generation/characterization process are as follows:
    (A) In the case of combustion emissions generated from light-duty 
vehicles/engines, the samples consist of three bags of vapor emissions 
(one from each segment of the light-duty exhaust emission cycle) plus 
one sample of particulate-phase emissions and one sample of semi-
volatile-phase emissions (collected over all segments of the exhaust 
emission cycle). If the mass of particulate emissions or semi-volatile 
emissions obtained during one driving cycle is not sufficient for 
characterization, then the driving cycle may be performed again and the 
extracted fractions combined prior to chemical analysis. Particulate-
phase emissions shall not be combined with semi-volatile-phase 
emissions.
    (B) In the case of combustion emissions generated from heavy-duty 
engines, the samples consist of one sample of each emission phase 
(vapor, particulate, and semi-volatile) collected over the entire cold-
start cycle and a second sample of each such phase collected over the 
entire hot-start cycle (see 40 CFR 86.334 through 86.342).
    (iv) Emission collection and storage. (A) Vapor phase emissions 
shall be collected and stored in Tedlar bags for subsequent chemical 
analysis. Storage conditions are specified in Sec. 79.52(b)(2).
    (B) Particulate phase emissions shall be collected on a particulate 
filter (or more than one, if required) using methods described in 40 
CFR 86.1301 through 86.1344. These methods, ordinarily applied only to 
heavy-duty emissions, are to be adapted and used for collection of 
particulates from light-duty vehicles/engines, as well. The particulate 
matter may be stored on the filter in a sealed container, or the 
soluble organic fraction may be extracted and stored in a separate 
sealed container. Both the particulate and the extract shall be 
shielded from ultraviolet light and stored at -20  deg.C or less. 
Particulate emissions shall be tested no later than six months from the 
date they were generated.
    (C) Semi-volatile emissions shall be collected immediately 
downstream from the particulate collection filters using porous polymer 
resin beds, or their equivalent, designed for their capture. The 
soluble organic fraction of semi-volatile emissions shall be extracted 
immediately and tested within six months of being generated. The 
extract shall be stored in a sealed container which is shielded from 
ultraviolet light and stored at -20  deg.C or less.
    (D) Particulate and semi-volatile phase emission collection, 
handling and extraction methods shall not alter the composition of the 
collected material, to the extent possible.
    (v) Additional requirements for combustion emission sampling, 
storage, and characterization are specified in Sec. 79.52(b).
    (2) Generating whole combustion emissions for biological testing. 
(i) Biological tests requiring whole combustion emissions shall be 
conducted using emissions generated from the test vehicle or engine 
operated in general accordance with the FTP procedures cited in this 
section. The emissions shall be generated continuously throughout the 
animal exposure periods, diluted by an amount appropriate for the test 
being performed as specified in Sec. 79.61(d)(3), passed through a 
mixing chamber, and routed to the biological test chamber.
    (ii) Light-duty test vehicles/engines shall be operated over the 
Urban Dynamometer Driving Schedule (or equivalent engine dynamometer 
trace, per paragraph (e)(1)(i)(A) of this section) and heavy-duty test 
engines shall be operated over the Engine Dynamometer Schedule (see 40 
CFR part 86, appendix I).
    (A) The tolerances of the driving cycle shall be two times those of 
the Federal Test Procedure and must be met 95 percent of the time.
    (B) The driving cycle shall be repeated as many times as required 
for the biological test session.
    (C) Light-duty dynamometers shall be calibrated prior to the start 
of a biological test (40 CFR 86.118-78), verified weekly (40 CFR 
86.118-78), and recalibrated as required. Heavy-duty dynamometers shall 
be calibrated and checked prior to the start of a biological test (40 
CFR 86.1318-84), recalibrated every two weeks (40 CFR 86.1318-84(a)) 
and checked as stated in 40 CFR 86.1318-84(b) and (c).
    (D) The fuel reservoir for the test vehicle/engine shall be large 
enough to operate the test vehicle/engine throughout the daily 
biological exposure period, avoiding the need for refueling during 
testing.
    (iii) An apparatus to integrate the large concentration swings 
typical of transient-cycle exhaust is to be used between the FTP-
Constant Volume Sampler (CVS) source of emissions and the exposure 
chamber containing the animal test cage(s). The purpose of such 
apparatus is to decrease the variability of the biological exposure 
atmosphere.
    (A) A large mixing chamber is suggested for this purpose. The 
mixing chamber would be charged from the CVS at a constant rate 
determined by the exposure chamber purge rate. Flow to the exposure 
chamber would begin at the conclusion of the initial transient cycle 
with the associated mixing chamber charge.
    (B) A potential alternative apparatus is a mini-diluter (see, for 
example, AIGER/CRADA, February, 1994 in Sec. 79.57(g)).
    (C) The mixing chamber (or any alternative emission moderation 
apparatus) must function such that the average concentration of total 
hydrocarbons leaving the apparatus shall be within 10 percent of the 
average concentration of hydrocarbons entering the chamber.
    (iv) Emission dilution. (A) Dilution air can be pre-dried to lower 
the relative humidity, thus permitting a lower dilution rate and a 
higher concentration of hydrocarbons to be achieved without 
condensation of water vapor.
    (B) With gasoline fuels, a minimum dilution ratio of about 1:5 raw 
exhaust (dewpoint about 125  deg.F) with dry, clean filtered air is 
required to reduce the water concentration to a dewpoint of about 68 
deg.F. The minimum dilution ratio (maximum exhaust flow rate) occurs at 
about 200 seconds into the UDDS transient driving cycle. Larger minimum 
dilution ratios are required if the dilution air includes water vapor. 
However, the minimum dilution ratio will vary with fuel composition. 
Fuels which generate greater engine exhaust water concentrations (e.g., 
alcohol and natural gas fuels) will require greater initial dilutions. 
Heated transfer ducts or tubing can be used to avoid water condensation 
in much of the system, but the mixing chamber described in paragraph 
(e)(2)(iii) of this section will generally be at or near laboratory 
temperature, and CVS dilution will have to be adequate to assure that 
the cumulative dew point in the chamber remains below laboratory 
temperature at all times (further guidance on this topic may be found 
in Black and Snow, 1994 in Sec. 79.57(g)).
    (C) After the initial exhaust dilution to preserve the character of 
the exhaust, the exhaust stream can be further diluted in the mixing 
chamber (and/or after leaving the chamber) to achieve the desired 
biological exposure concentrations.
    (v) Verification procedures. (A) The entire system used to dilute 
and transport whole combustion emissions (i.e., from exhaust pipe to 
outlet in the biological testing chamber) shall be verified before any 
animal exposures begin, and verified at least weekly during testing. 
(See procedures at 40 CFR 86.119-90 for light-duty vehicles and 
Sec. 86.1319-90 for heavy-duty engines.) Verification testing shall be 
accomplished by introducing a known sample at the end of the vehicle/
engine exhaust pipe into the dilution system and measuring the amount 
exiting the system. For example, an injected hydrocarbon sample could 
be detected with a gas chromatograph (GC) and flame ionization detector 
(FID) to determine the recovery factor.
    (B) Verification of the integrity of the mixing chamber (or 
alternative apparatus) shall be determined before animal exposures 
begin and at least weekly thereafter. Composite values for weight 
percent total hydrocarbons shall be determined for the test vehicle/
engine's dilute exhaust stream entering and exiting the mixing chamber 
apparatus. These values must be within 10 percent of each other.
    (vi) Emission exposure quality control. (A) The tester shall 
incorporate the additional quality assurance and safety procedures 
outlined in Sec. 79.61(d) to control variability of emissions during 
the generation of exposure emissions during health effect testing.
    (B) These procedures include requirements that the mean exposure 
concentration in the inhalation test chamber shall be within 10 percent 
of the target concentration (established in the developmental phase of 
testing) on 90 percent or more of exposure days and that daily 
monitoring of CO, CO2, NOX, SOX, and total hydrocarbons 
in the exposure chamber shall be required. Analysis of the particle 
size distribution shall also be performed to establish the stability 
and consistency of particle size distribution in the test exposure.
    (C) The testing facility shall allow an audit of its premises, the 
qualifications, e.g., curriculum vitae, of its staff assigned to 
testing, and the specimens and records of the testing for registration 
purposes (as specified in Sec. 79.60).
    (vii) In order to allow for unforeseen problems with the emission 
generation or dilution equipment, emission generation may be 
interrupted for up to four hours on a maximum of two occasions in any 
four-week period of testing. The amount of time for which emission 
generation was interrupted shall subsequently be added after the 
equipment problem is corrected. If the equipment problem causes more 
than four consecutive hours of emission generation to be interrupted, 
or if more than two such occasions occurs in any four-week period 
during testing, the interrupted tests shall be void. Testers shall be 
aware of concerns for backup vehicles/engines cited in paragraph 
(a)(7)(ii) of this section.
    (3) Generating particulate and semi-volatile emissions for 
biological testing. (i) Salmonella mutagenicity testing, pursuant to 
Sec. 79.68, shall be conducted on extracts of the particulate and semi-
volatile emission phases separately. These emissions shall be generated 
by operating the test vehicle/engine over the appropriate FTP driving 
cycle (see paragraph (e)(2)(ii) of this section) and collected and 
analyzed according to methods described in 40 CFR 86.1301 through 1344 
(further information on this subject may be found in Perez, et al. CRC 
Report No. 551, 1987 listed in Sec. 79.57(g)).
    (A) Particulate emissions shall be collected on particulate filters 
and extracted from the collection equipment for use in biological 
tests. The particulate emissions from all segments of the FTP or from 
multiple FTP cycles may be collected on one or more filters, as 
necessary. The time spent collecting sufficient quantities of the test 
substances in emissions samples will vary, depending on the emission 
characteristics of the engine and fuel or additive/base fuel mixture 
and on the requirements of the biological test protocol.
    (B) Semi-volatile emissions shall be collected immediately 
downstream from the particulate collection filters using porous polymer 
resin beds, or their equivalent, designed for their capture. Semi-
volatile phase emissions shall be collected on one apparatus. The time 
spent collecting sufficient quantities of the test substances in 
emissions samples will vary, depending on the emission characteristics 
of the engine and fuel or additive/base fuel mixture and on the 
requirements of the biological test protocol.
    (ii) The extraction method shall be determined by the 
specifications of the biological test for which the emissions are used.
    (iii) Particulate and semi-volatile emission storage requirements 
are as specified in Sec. 79.57(e)(1)(iv).
    (iv) Particulate and semi-volatile phase emission collection, 
handling and extraction methods shall not alter the composition of the 
collected material, to the extent possible.
    (v) Particulate emissions shall not be combined with semi-volatile 
phase emissions.
    (f) Generation of evaporative emissions for characterization and 
biological testing. (1) Except as provided in paragraph (f)(5) of this 
section, an evaporative emissions generator shall be used to volatilize 
samples of a fuel or additive/base fuel mixture for evaporative 
emissions characterization and biological testing. Emissions shall be 
collected and sampled using equipment and methods appropriate for use 
with the compounds being characterized and the requirements of the 
emission characterization analysis. In the case of potentially 
explosive test substance concentrations, care must be taken to avoid 
generating explosive atmospheres. The tester is referred to 
Sec. 79.61(d)(8) for considerations involving explosivity.
    (2) Evaporative Emissions Generator (EEG) Description. An EEG is a 
fuel tank or vessel to which heat is applied causing a portion of the 
fuel to evaporate at a desired rate. The manufacturer has flexibility 
in designing an EEG for testing a particular fuel or fuel additive. The 
sample used to generate emissions in the EEG shall be renewed at least 
daily.
    (i) The evaporation chamber shall be made from materials compatible 
with the fuels and additives being tested and shall be equipped with a 
drain.
    (ii) The chamber shall be filled to 40 5 percent of its 
interior volume with the fuel or additive/base fuel mixture being 
tested, with the remainder of the volume containing air.
    (iii) The concentration of the evaporated fuel or additive/base 
fuel mixture in the vapor space of the evaporation chamber during the 
time emissions are being withdrawn for testing shall not vary by more 
than 10 percent from the equilibrium concentration in the vapor space 
of emissions generated from the fresh fuel or additive/base fuel 
mixture in the chamber.
    (A) During the course of a day's emission generation period, the 
level of fuel in the EEG shall be maintained to within 7 percent of its 
height at the start of the daily exposure period.
    (B) The fuel used in the EEG shall be drained at the end of each 
daily exposure. The EEG shall be refilled with a fresh supply of the 
test formulation before the start of each daily exposure.
    (C) The vapor space of the evaporation chamber shall be well mixed 
throughout the time emissions are being withdrawn for testing.
    (iv) The size of the evaporation chamber shall be determined by the 
rate at which evaporative emissions shall be needed in the test animal 
exposure chambers and the rate at which the fuel or the additive/base 
fuel mixture evaporates. The rate of evaporative emissions may be 
adjusted by altering the size of the EEG or by using one or more 
additional EEG(s). Emission rate modifications shall not be adjusted by 
temperature control or pressure control.
    (v) The temperature of the fuel or additive/base fuel mixture in 
the evaporation chamber shall be 130  deg.F5 deg.F. The 
vapors shall maintain this temperature up to the point in the system 
where the vapors are diluted.
    (vi) The pressure in the vapor space of the evaporation chamber and 
the dilution and sampling apparatus shall stay within 10 percent of 
ambient atmospheric pressure.
    (vii) There shall be no controls or equipment on the evaporation 
chamber system that change the concentration or composition of the 
vapors generated for testing.
    (viii) Manufacturers shall perform verification testing of 
evaporative emissions in a manner analogous to the verification testing 
performed for combustion emissions.
    (3) For biological testing, vapor shall be withdrawn from the EEG 
at a constant rate, diluted with air as required for the particular 
study, and conducted immediately to the biological testing chamber(s) 
in a manner similar to the method used in Sec. 79.57(e), excluding the 
mixing chamber therein. The rate of emission generation shall be high 
enough to supply the biological exposure chamber with sufficient 
emissions to allow for a minimum of fifteen air changes per exposure 
chamber per hour. Interruption of evaporative emissions exposures 
during biological testing for more than four consecutive hours, or on 
more than two separate occasions within a four-week period for less 
than four consecutive hours, shall cause the affected test(s) to be 
void.
    (4) For characterization of evaporative emissions, samples of 
equilibrated emissions to the vapor space of the EEG shall be withdrawn 
into Tedlar bags, then stored and analyzed as specified in 
Sec. 79.52(b).
    (5) A manufacturer (or group of manufacturers) may submit to EPA a 
request for approval of an alternative method of generating evaporative 
emissions for use in emission characterization and biological tests 
required under this subpart.
    (i) To be approved by EPA, the request must fully explain the 
rationale for the proposed method as well as the technical procedures, 
quality control, and safety precautions to be used, and must 
demonstrate that the proposed method will meet the following criteria:
    (A) The emission mixture generated by the proposed procedures must 
be reasonably similar to the equilibrium composition of the vapor which 
occurs in the vehicle fuel tank head space when the subject fuel or 
additive/base fuel mixture is in use and near-maximum in-use 
temperatures are encountered.
    (B) The emissions mixture generated by the proposed method must be 
sufficiently concentrated to provide adequate exposure levels in the 
context of the required toxicologic tests.
    (C) The proposed method must include procedures to ensure that the 
emissions delivered to the biologic exposure chambers will provide a 
reasonably constant exposure atmosphere over time.
    (ii) If EPA approves the request, EPA will place in the public 
record a copy of the request, together with all supporting procedural 
descriptions and justifications, and will notify the public of its 
availability by publishing a notice in the Federal Register.
    (g) References. For additional background information on the 
emission generation procedures outlined in this paragraph (g), the 
following references may be consulted. Additional references can be 
found in Sec. 79.61(f).
    (1) AIGER/CRADA (American Industry/Government Emissions Research 
Cooperative Research and Development Agreement, ``Specifications for 
Advanced Emissions Test Instrumentation'' AIGER PD-94-1, Revision 5.0, 
February, 1994
    (2) Black, F. and R. Snow, ``Constant Volume Sampling System Water 
Condensation'' SAE #940970 in ``Testing and Instrumentation'' SP-1039, 
Society of Automotive Engineers, Feb. 28-Mar. 3, 1994.
    (3) Perez, J.M., Jass, R.E., Leddy, D.G., eds. ``Chemical Methods 
for the Measurement of Unregulated Diesel Emissions (CRC-APRAC Project 
No. CAPI-1-64), Coordinating Research Council, CRC Report No. 551, 
August, 1987.
    (4) Phalen, R.F., ``Inhalation Studies: Foundations and 
Techniques'', CRC Press, Inc., Boca Raton, Florida, 1984.


Sec. 79.58  Special provisions.

    (a) Relabeled Additives. Sellers of relabeled additives (pursuant 
to Sec. 79.50) are not required to comply with the provisions of 
Secs. 79.52, 79.53 or 79.59, except that such sellers are required to 
comply with Sec. 79.59(b).
    (b) Low Vapor Pressure Fuels and Additives. Fuels which are not 
designated as ``evaporative fuels'' and fuel additives which are not 
designated as ``evaporative fuel additives'' pursuant to the 
definitions in Sec. 79.50 need not undergo the emission 
characterization or health effects testing specified in Secs. 79.52 and 
79.53 for evaporative emissions. At EPA's discretion, the evaporative 
emissions of such fuels and additives may be required to undergo Tier 3 
testing, pursuant to Sec. 79.54.
    (c) Alternative Tier 2 Provisions. At EPA's discretion, EPA may 
modify the standard Tier 2 health effects testing requirements for a 
fuel or fuel additive (or group). Such modification may encompass 
substitution, addition, or deletion of Tier 2 studies or study 
specifications, and/or changes in underlying engine or equipment 
requirements, except that a Tier 2 endpoint will not be deleted in the 
absence of existing information deemed adequate by EPA or alternative 
testing requirements for such endpoint. If warranted by the particular 
requirements, EPA will allow additional time for completion of the 
alternative Tier 2 testing program.
    (1) When EPA intends to require testing in lieu of or in addition 
to standard Tier 2 health testing, EPA will notify the responsible 
manufacturer (or group) by certified letter of the specific tests which 
EPA is proposing to require in lieu of or in addition to Tier 2, and 
the proposed schedule for completion and submission of such tests. A 
copy of the letter will be placed in the public record. EPA intends to 
send the notification prior to November 27, 1995, or in the case of new 
fuels and additives (as defined in Sec. 79.51(c)(3)), within 18 months 
of EPA's receipt of an intent to register such product. However, EPA's 
notification to the manufacturer (or group) may occur at any time up to 
EPA's receipt of Tier 2 data for the product(s) in question. EPA will 
provide the manufacturer with 60 days from the date of receipt of the 
notice to comment on the tests which EPA is proposing to require and on 
the proposed schedule. If the manufacturer believes that undue costs or 
hardships will occur as a result of EPA's delay in providing 
notification of alternative Tier 2 requirements, then the 
manufacturer's comments should describe and include evidence of such 
hardship. In particular, if the standard Tier 2 toxicology testing for 
the fuel or additive in question has already begun at the time the 
manufacturer receives EPA's notification of proposed alternative Tier 2 
requirements, then EPA shall refrain from requiring alternative Tier 2 
tests provided that EPA receives the standard Tier 2 data and report 
(pursuant to Sec. 79.59(c)) within one year of the date on which the 
toxicology testing began.
    (2) EPA will issue a notice in the Federal Register announcing its 
intent to require special testing in lieu of or in addition to the 
standard Tier 2 testing for a particular fuel or additive manufacturer 
or group, and that a copy of the letter to the manufacturer or group 
describing the proposed alternative Tier 2 testing for that 
manufacturer or group is available in the public record for review and 
comment. The public shall have a minimum of 30 days after the 
publication of this notice to comment on the proposed alternative Tier 
2 testing.
    (3) EPA will include in the public record a copy of any timely 
comments concerning the proposed alternative Tier 2 testing 
requirements received from the affected manufacturer or group or from 
the public, and the responses of EPA to such comments. After reviewing 
all such comments received, EPA may adopt final alternative Tier 2 
requirements by sending a certified letter describing such final 
requirements to the manufacturer or group. In that event, EPA will also 
issue a notice in the Federal Register announcing that it has adopted 
final alternative Tier 2 requirements and that a copy of the letter 
adopting the requirements has been included in the public record.
    (4) After EPA's receipt of a manufacturer's (or group's) 
submittals, EPA will notify the responsible manufacturer (or group) 
regarding the adequacy of the submittal and potential Tier 3 testing 
requirements according to the same relative time intervals and by the 
same procedures as specified in Sec. 79.51 (c) and (d) for routine Tier 
1 and Tier 2 submittals.
    (d) Small Business Provisions. (1) For purposes of these 
provisions, when subsidiary, divisional, or other complex business 
arrangements exist, manufacturer is defined as the business entity with 
ultimate ownership of all related parents, subsidiaries, divisions, 
branches, or other operating units. Total annual sales means the 
average of the manufacturer's total sales revenue in each of the three 
years prior to such manufacturer's submittal to EPA of the basic 
registration information pursuant to Sec. 79.59 (b)(2) through (b)(5).
    (2) Provisions Applicable to Baseline and Non-baseline Products. A 
manufacturer with total annual sales less than $50 million is not 
required to meet the requirements of Tier 1 and Tier 2 (specified in 
Secs. 79.52 and 79.53) with regard to such manufacturer's fuel and/or 
additive products which meet the criteria for inclusion in a Baseline 
or Non-baseline group pursuant to Sec. 79.56. Upon such manufacturer's 
satisfactory completion and submittal to EPA of basic registration data 
specified in Sec. 79.59(b), the manufacturer may request and EPA shall 
issue a registration for such product, subject to Sec. 79.51(c) and 
paragraphs (d)(4) and (d)(5) of this section.
    (3) Provisions Applicable to Atypical Products. A manufacturer with 
total annual sales less than $10 million is not required to meet the 
requirements of Tier 2 (specified in Sec. 79.53) in regard to such 
manufacturer's fuel and/or additive products which meet the criteria 
for inclusion in an Atypical group pursuant to Sec. 79.56. Upon such 
manufacturer's satisfactory completion and submittal to EPA of basic 
registration data specified in Sec. 79.59(b) and Tier 1 information 
specified in Sec. 79.52 for an Atypical fuel or additive, the 
manufacturer may request and EPA shall issue a registration for such 
product, subject to Sec. 79.51(c) and paragraphs (d)(4) and (d)(5) of 
this section. Compliance with Tier 1 requirements under this paragraph 
may be accomplished by the individual manufacturer or as a part of a 
group pursuant to Sec. 79.56.
    (4) Any registration granted by EPA under the provisions of this 
section are conditional upon satisfactory completion of any Tier 3 
requirements which EPA may subsequently impose pursuant to Sec. 79.54. 
In such circumstances, the Tier 3 requirements might include (but would 
not necessarily be limited to) information which would otherwise have 
been required under the provisions of Tier 1 and/or Tier 2.
    (5) The provisions in paragraphs (d)(2) and (d)(3) of this section 
are voluntary on the part of qualifying small manufacturers. Such 
manufacturers may choose to fulfill the standard requirements for their 
fuels and additives, individually or as a part of a group, rather than 
satisfying only the requirements specified in paragraphs (d)(2) and/or 
(d)(3) of this section. If a qualifying small manufacturer elects these 
special provisions rather than the standard requirements for a product, 
then EPA will generally assume that any additional information 
submitted by other manufacturers, for fuels and additives meeting the 
same grouping criteria (under Sec. 79.56) as that of the small 
manufacturer's product, is pertinent to further testing and/or 
regulatory decisions that may affect the small manufacturer's product.
    (e) Aftermarket Aerosol Additives. (1) To obtain registration for 
an aftermarket aerosol fuel additive, the manufacturer shall provide 
existing information in the form of a literature search, a discussion 
of the potential exposure(s) to such product, and the basic 
registration data specified in Sec. 79.59(b).
     (2) The literature search shall include existing data on potential 
health and welfare effects due to exposure to the aerosol product 
itself and its raw (uncombusted) components. The analysis for potential 
exposures shall be based on the actual or anticipated production volume 
and market distribution of the particular aerosol product, and its 
estimated frequency of use. Other Tier 1 and Tier 2 requirements are 
not routinely required for aerosol products. EPA will review the 
submitted information and, at EPA's discretion, may require from the 
manufacturer further information and/or testing under Tier 3 on a case-
by-case basis.


Sec. 79.59  Reporting requirements.

    (a) Timing. (1) The manufacturer of each designated fuel or fuel 
additive shall submit to EPA the basic registration data detailed in 
paragraph (b) of this section. Forms for submitting this data may be 
obtained from EPA at the following address: Director, Field Operations 
and Support Division, 6406J--Fuel/Additives Registration, U.S. 
Environmental Protection Agency, 401 M Street, S.W., Washington, DC 
20460.
    (i) For existing products (pursuant to Sec. 79.51(c)(1)), 
manufacturers shall submit the basic registration data as specified in 
Sec. 79.59(b) to EPA by November 28, 1994.
    (ii) For registrable products (pursuant to Sec. 79.51(c)(2)), 
manufacturers shall submit the basic registration data as specified in 
Sec. 79.59(b) to apply for registration for such product.
    (iii) For new products (pursuant to Sec. 79.51(c)(3)), 
manufacturers are strongly encouraged to notify EPA of an intent to 
obtain product registration by submitting the basic registration data 
as specified in Sec. 79.59(b) prior to starting Tiers 1 and 2.
    (2) The information specified in paragraph (c) of this section 
shall be submitted to the address in paragraph (a)(1) of this section 
at the conclusion of activities performed in compliance with Tiers 1 
and 2 under the provisions of Secs. 79.52 and 79.53, according to the 
time constraints specified in Sec. 79.51 (c) through (d).
    (3) The information specified in paragraph (d) of this section 
shall be submitted to EPA at the address in paragraph (a)(1) of this 
section at the conclusion of activities performed in compliance with 
Tier 3 under the provisions of Sec. 79.54.
    (b) Basic Registration Data. Each manufacturer of a designated fuel 
or fuel additive shall submit the following data in regard to such fuel 
or fuel additive:
    (1) The information specified in Sec. 79.11 or Sec. 79.21. If such 
information has already been submitted to EPA in compliance with 
subpart B or C of this part, and if such previous information is 
accurate and up-to-date, the manufacturer need not resubmit this 
information.
    (2) Annual production volume of the fuel or fuel additive product, 
in units of gallons per year if most commonly sold in liquid form or 
kilograms per year if most commonly sold in solid form. For fuels and 
fuel additives already in production, the most recent annual production 
volume and the volume projected to be produced in the third subsequent 
year shall be provided. For products not yet in production, the best 
estimate of expected annual volume during the third year of production 
shall be provided.
    (3) Market distribution of the product. For fuels and bulk 
additives, this information shall be presented as the percent of total 
annual sales volume marketed in each Petroleum Administration for 
Defense District (PADD). The states comprising each PADD are listed in 
the following section. For aftermarket additives, the distribution data 
shall be presented as the percent of total annual sales volume marketed 
in each state. For a product not yet in production, the manufacturer 
shall present the distribution (by PADD or state, as applicable) 
projected to occur during the third year of production.
    (i) The following states and jurisdictions are included in PADD I:

Connecticut
Delaware
District of Columbia
Florida
Georgia
Maine
Maryland
Massachusetts
New Hampshire
New Jersey
New York
North Carolina
Pennsylvania
Rhode Island
South Carolina
Vermont
Virginia
West Virginia

    (ii) The following states are included in PADD II:

Illinois
Indiana
Iowa
Kansas
Kentucky
Michigan
Minnesota
Missouri
Nebraska
North Dakota
Ohio
Oklahoma
South Dakota
Tennessee
Wisconsin

    (iii) The following states are included in PADD III:

Alabama
Arkansas
Louisiana
Mississippi
New Mexico
Texas

    (iv) The following states are included in PADD IV:

Colorado
Idaho
Montana
Utah
Wyoming

    (v) The following states are included in PADD V:

Alaska
Arizona
California
Hawaii
Nevada
Oregon
Washington

    (4) Any applicable information pursuant to the grouping provisions 
in Sec. 79.56, as follows:
    (i) If the manufacturer has enrolled or intends to enroll the 
product in a fuel/additive group, the relevant group and the person(s) 
or entity expected to submit information on behalf of the group must be 
identified.
    (ii) If the manufacturer intends to rely on registration 
information previously submitted by another manufacturer (or group) for 
registration of other product(s) in the same fuel/additive group, then 
the original submitter and its product (or product group) shall be 
identified. In such cases, the manufacturer shall provide evidence that 
the original submitter has been notified of the use of its registration 
data and that the manufacturer has complied or intends to comply with 
the proportional reimbursement required under Sec. 79.56(c) of this 
rule.
    (5) Any applicable information pursuant to the special provisions 
in Sec. 79.58, as follows:
    (i) If the manufacturer claims applicability of the special 
provisions for relabeled additives, pursuant to Sec. 79.58(a), then the 
manufacturer and brand name of the original product shall be given.
    (ii) If the manufacturer claims applicability of any small business 
provisions pursuant to Sec. 79.58(d), the average of the manufacturer's 
total annual sales revenue for the previous three years shall be given.
    (iii) If the manufacturer claims applicability of the special 
provisions for aerosol products, pursuant to Sec. 79.58(e), then the 
purpose and recommended frequency of use shall be given.
    (c) Tier 1 and Tier 2 Reports. If the results of Tiers 1 and 2 are 
reported to EPA at the same time, then the report shall include the 
following documents in paragraphs (c)(1) through (7) of this section. 
If Tier 1 and Tier 2 results are submitted to EPA separately, then the 
separate Tier 1 report shall include only documents in paragraphs 
(c)(1) through (4), (c)(6), and associated appendices in paragraphs 
(c)(7) of this section, and the separate Tier 2 report shall include 
only documents in paragraphs (c)(1) through (3), (c)(5), (c)(6), and 
associated appendices in paragrpah (c)(7) of this section. In addition, 
pursuant to the requirements in Sec. 79.51(c)(1)(ii)(B), if the Tier 2 
report for registered fuels and fuel additives is not submitted prior 
to May 27, 1997, then evidence of a suitable arrangement for completion 
of Tier 2 (e.g., a copy of a signed contract with a qualified 
laboratory for applicable Tier 2 services) must be submitted to EPA 
prior to that date.
    (1) Cover page. (i) Identification of test substance,
    (ii) Name and address of the manufacturer of the test substance,
    (iii) Name and phone number of a designated contact person,
    (iv) Group information, if applicable, including:
    (A) Group name or grouping criteria,
    (B) Name and address of responsible organization or entity 
reporting for the group,
    (C) Product trade name and manufacturer of each member fuel and 
additive to which the report pertains.
    (2) Executive Summary. Text overview of the significant results and 
conclusions obtained as a result of completing the requirements of Tier 
1 and/or Tier 2, including references if used to support such results 
and conclusions.
    (3) Test Substance Information. Test substance description, 
including, as applicable,
    (i) Base fuel parameter values (including types and concentrations 
of base fuel additives) or test fuel composition (if a fuel other than 
the base fuel is used in testing). These values must be provided for 
each of the fuel parameters specified in Sec. 79.55 for the applicable 
fuel family.
    (ii) Test additive composition and concentration
    (4) Summary of Tier 1 (i) Literature Search. Pursuant to 
Sec. 79.52(d), the literature search shall include a text summary of 
the methods and results of the literature search, including the 
following:
    (A) Identification of person(s) performing the literature search,
    (B) Description of data sources accessed, search strategy used, 
search period, and terms included in literature search,
    (C) Documentation of all unpublished in-house and other privately-
conducted studies,
    (D) Tables summarizing the protocols and results of all cited 
studies,
    (E) Summary of significant results and conclusions with respect to 
the effects of the emissions of the subject fuel or fuel additive on 
the public health and welfare, including references if used to support 
such results and conclusions.
    (F) Statement of the extent to which the literature search has 
produced adequate information comparable to that which would otherwise 
be obtained through the performance of applicable emission 
characterization requirements under Sec. 79.52(b) and/or health effects 
testing requirements under Sec. 79.53, including justifications and 
specific references.
    (ii) Emission Characterization. Pursuant to Sec. 79.52(b), the 
emission characterization shall include:
    (A) Name, address, and telephone number of the laboratory 
performing the characterization,
    (B) Name and description of analytic methods used for 
characterization.
    (iii) Exposure Analysis. Pursuant to Sec. 79.52(c), the exposure 
analysis shall include:
    (A) A qualitative discussion of the potential exposure(s) of the 
general and any special at-risk populations to the emission products, 
based on annual and projected production volume, and market 
distribution data. For group submittals, this discussion shall address 
the characteristics of the cumulative exposure from the potential use 
of all fuel or additive products in the group.
    (B) Identification of person(s) preparing the analysis.
    (5) Summary of Tier 2. For each health effects test performed 
pursuant to the provisions of Sec. 79.53, the Tier 2 summary shall 
contain the following information:
    (i) Name, address, and telephone number of the testing facility,
    (ii) Summary of procedures (including quality assurance, quality 
control and compliance with Good Laboratory Practice Standards as 
specified in Sec. 79.60), findings, and conclusions, including 
references if used to support such results and conclusions,
    (iii) Description of any problems and their resolution.
    (6) Conclusions. The conclusions shall identify the need for 
further testing, if that need exists, or justify that current testing 
and/or available information is adequate for the tier(s) included in 
the report.
    (7) Appendices. The appendices shall contain detailed documentation 
related to the summary information described in this section, 
including, at a minimum, the following five appendices:
    (i) Literature search appendices shall contain:
    (A) Copies of literature source outputs, including reference lists 
and associated abstracts from database searches, printed or on 3\1/2\ 
inch IBM-compatible computer diskettes;
    (B) Summary tables organized by health or welfare endpoint and type 
of emission (e.g., combustion, evaporation, individual emission 
product), presenting in tabular form the following information at a 
minimum: number and species of test subjects, exposure concentrations/
duration, positive (i.e., abnormal) findings including numbers of test 
subjects involved, and bibliographic references;
    (C) Complete documentation and/or reprints of articles for any 
previous study relied upon for satisfying emission characterization 
and/or Tier 2 test requirements; and
    (D) Full reports for unpublished/in-house studies.
    (ii) Emissions characterization appendices shall contain:
    (A) Complete laboratory reports, including documentation of 
calibration and verification procedures;
    (B) Documentation of the emissions generation procedures used; and
    (C) Lists of speciated emission products and their emission rates 
reported in units of grams/mile.
    (iii) Exposure analysis appendices may be submitted to report any 
detailed documentation of data used in the analyses and/or calculations 
determining potential exposures to population(s). If modeling data are 
used, these should be included in an appendix.
    (iv) Tier 2 appendices shall contain, for each test performed:
    (A) Complete protocol used;
    (B) Documentation of emission generation procedures; and
    (C) Complete laboratory report in compliance with the reporting 
standards in Sec. 79.60, including detailed test results and 
conclusions, and descriptions of any problems encountered and their 
resolution.
    (v) Laboratory certification/accreditation information, personnel 
credentials, and statements of compliance with the Good Laboratory 
Practices Standards specified in Sec. 79.60 and the requirements in 
Sec. 79.53(c)(1).
    (d) Tier 3 Report. Subject to applicability as specified in 
Sec. 79.54, each manufacturer of a designated fuel or fuel additive, or 
each group of such manufacturers pursuant to the provisions of 
Sec. 79.56, shall submit the following information with respect to each 
Tier 3 test conducted for such fuels or fuel additives:
    (1) The test objectives, including a summary of the reason(s) why 
such additional testing, beyond Tiers 1 and 2, was required;
    (2) Name, address, and telephone number of each testing facility;
    (3) Summary of test procedures, results and conclusions;
    (4) Complete documentation of test protocols and emission 
generation procedures, complete laboratory reports in compliance with 
the reporting standards of Sec. 79.60, detailed test results and 
conclusions, including references if used to support such results and 
conclusions, and descriptions of any problems encountered and their 
resolution; and
    (5) Laboratory certification information, personnel credentials, 
and statements of compliance with the Good Laboratory Practices 
Standards specified in Sec. 79.60.
    (e) Availability of Information. (1) All health and safety test 
data and other information concerning health and welfare effects which 
is submitted by any manufacturer or group pursuant to Secs. 79.52(c), 
79.53, or 79.54, shall be considered to be public information and shall 
be made available to the public by EPA upon request. A reasonable fee 
may be charged by EPA for copying such materials. Any manufacturer or 
group who claims that any information concerning the composition of a 
fuel or fuel additive product, or any other information, submitted 
under this subpart is confidential business information must state this 
claim in writing at the time of the submittal.
    (2) To assert a business confidentiality claim concerning any 
information submitted under this subpart, the submitter must:
     (i) Clearly mark the information as confidential at each location 
it appears in the submission; and
    (ii) Submit with the information claimed as confidential a separate 
document setting forth the claim and listing each location at which the 
information appears in the submission.
    (3) If any person subsequently requests access to information 
submitted under this subpart (other than health and safety test data 
and other information concerning health and welfare effects), and such 
information is subject to a claim of business confidentiality, the 
request and any subsequent disclosure shall be governed by the 
provisions of 40 CFR part 2.


Sec. 79.60  Good laboratory practices (GLP) standards for inhalation 
exposure health effects testing.

    (a) General Provisions--(1) Scope. (i) This section prescribes good 
laboratory practices (GLPs) for conducting inhalation exposure studies 
relating to motor vehicle emissions health effects testing under this 
part. These directions are intended to ensure the quality and integrity 
of health effects data submitted pursuant to registration regulations 
issued under sections 211(b) or 211(e) of the Clean Air Act (CAA) (42 
U.S.C. 7545).
    (ii) This section applies to any study described by paragraph 
(a)(1)(i) of this section which any person conducts, initiates, or 
supports on or after May 27, 1994.
    (iii) It is EPA's policy that all health effects data developed 
under sections 211(b) and (e) of CAA be in accordance with provisions 
of this section. If data are not developed in accordance with the 
provisions of this section, EPA may consider such data insufficient to 
evaluate the health effects of a motor vehicle's fuel or fuel additive 
emissions, unless the submitter provides additional information 
demonstrating that the data are reliable and adequate and EPA 
determines that the data are sufficient.
    (2) Definitions. As used in this section, the following terms shall 
have the meanings specified:
    Batch means a specific quantity or lot of a test fuel, additive/
base fuel mixture, or reference substance that has been characterized 
according to Sec. 79.60(f)(1)(i).
    CAA means the Clean Air Act.
    Carrier means any material which is combined with engine/motor 
vehicle emissions or a reference substance for administration to a test 
system. ``Carrier'' includes, but is not limited to, clean, filtered 
air, water, feed, and nutrient media.
    Control atmosphere means clean, filtered air which is administered 
to the test system in the course of a study for the purpose of 
establishing a basis for comparison with the test atmosphere for 
chemical or biological measurements.
    Experimental start date means the first date the test atmosphere is 
applied to the test system.
    Experimental termination date means the last date on which data are 
collected directly from the study.
    Person includes an individual, partnership, corporation, 
association, scientific or academic establishment, government agency, 
or organizational unit thereof, and any other legal entity.
    Quality assurance unit means any person or organizational element, 
except the study director, designated by testing facility management to 
perform the duties relating to quality assurance of the studies.
    Raw data means any laboratory worksheets, records, memoranda, 
notes, or exact copies thereof, that are the result of original 
observations and activities of a study and are necessary for the 
reconstruction and evaluation of the report of that study. In the event 
that exact transcripts of raw data have been prepared (e.g., tapes 
which have been transcribed verbatim, dated, and verified accurate by 
signature), the exact copy or exact transcript may be substituted for 
the original source as raw data. ``Raw data'' may include photographs, 
videotape, microfilm or microfiche copies, computer printouts, magnetic 
media, including dictated observations, and recorded data from 
automated instruments.
    Reference substance means any chemical substance or mixture, 
analytical standard, or material other than engine/motor vehicle 
emissions and/or its carrier, that is administered to or used in 
analyzing the test system in the course of a study. A ``reference 
substance'' is used to establish a basis for comparison with the test 
atmosphere for known chemical or biological measurements, i.e., 
positive or negative control substance.
    Specimen means any material derived from a test system for 
examination or analysis.
    Sponsor means person who initiates and supports, by provision of 
financial or other resources, a study or a person who submits a study 
to EPA in response to the CAA Section 211(b) or 211(e) Fuels and Fuel 
Additives Registration Rule or a testing facility, if it both initiates 
and actually conducts the study.
    Study means any experiment, at one or more test sites, in which a 
test system is exposed to a test atmosphere under laboratory conditions 
to determine or help predict the health effects of that exposure in 
humans, other living organisms, or media.
    Study completion date means the date the final report is signed by 
the study director.
    Study director means the individual responsible for the overall 
conduct of a study.
    Study initiation date means the date the protocol is signed by the 
study director.
    Test substance means a vapor and/or aerosol mixture composed of 
engine/motor vehicle emissions and clean, filtered air which is 
administered directly, or indirectly, by the inhalation route to a test 
system in a study which develops data to meet the registration 
requirements of CAA section 211(b) or (e).
    Test system means any animal, microorganism, chemical or physical 
matrix, to which the test, control, or reference substance is 
administered or added for study. This definition also includes 
appropriate groups or components of the system not treated with the 
test, control, or reference substance.
    Testing facility means a person who actually conducts a study, 
i.e., actually uses the test substance in a test system. ``Testing 
facility'' encompasses only those operational units that are being or 
have been used to conduct studies.
    TSCA means the Toxic Substances Control Act (15 U.S.C. 2601 et 
seq.).
    (3) Applicability to studies performed under grants and contracts. 
When a sponsor or other person utilizes the services of a consulting 
laboratory, contractor, or grantee to perform all or a part of a study 
to which this section applies, it shall notify the consulting 
laboratory, contractor, or grantee that the service is, or is part of, 
a study that must be conducted in compliance with the provisions of 
this section.
    (4) Statement of compliance or non-compliance. Any person who 
submits to EPA a test in compliance with registration regulations 
issued under CAA section 211(b) or section 211(e) shall include in the 
submission a true and correct statement, signed by the sponsor and the 
study director, of one of the following types:
    (i) A statement that the study was conducted in accordance with 
this section; or
    (ii) A statement describing in detail all differences between the 
practices used in the study and those required by this section; or
    (iii) A statement that the person was not a sponsor of the study, 
did not conduct the study, and does not know whether the study was 
conducted in accordance with this section.
    (5) Inspection of a testing facility. (i) A testing facility shall 
permit an authorized employee or duly designated representative of EPA, 
at reasonable times and in a reasonable manner, to inspect the facility 
and to inspect (and in the case of records also to copy) all records 
and specimens required to be maintained regarding studies to which this 
section applies. The records inspection and copying requirements shall 
not apply to quality assurance unit records of findings and problems, 
or to actions recommended and taken, except the EPA may seek production 
of these records in litigation or formal adjudicatory hearings.
    (ii) EPA will not consider reliable for purposes of showing that a 
test substance does or does not present a risk of injury to health or 
the environment any data developed by a testing facility or sponsor 
that refuses to permit inspection in accordance with this section. The 
determination that a study will not be considered reliable does not, 
however, relieve the sponsor of a required test of any obligation under 
any applicable statute or regulation to submit the results of the study 
to EPA.
    (6) Effects of non-compliance. (i) Pursuant to sections 114, 208, 
and 211(d) of the CAA, it shall be a violation of this section and a 
violation of this rule (40 CFR part 79, subpart F) if:
    (A) The test is not being or was not conducted in accordance with 
any requirement of this part; or
    (B) Data or information submitted to EPA under part 79, including 
the statement required by Sec. 79.60(a)(4), include information or data 
that are false or misleading, contain significant omissions, or 
otherwise do not fulfill the requirements of this part; or
    (C) Entry in accordance with Sec. 79.60(a)(5) for the purpose of 
auditing test data is denied.
    (ii) EPA, at its discretion, may not consider reliable for purposes 
of showing that a chemical substance or mixture does not present a risk 
of injury to health any study which was not conducted in accordance 
with this part. EPA, at its discretion, may rely upon such studies for 
purposes of showing adverse effects. The determination that a study 
will not be considered reliable does not, however, relieve the sponsor 
of a required test of the obligation under any applicable statute or 
regulation to submit the results of the study to EPA.
    (iii) If data submitted in compliance with registration regulations 
issued under CAA section 211(b) or section 211(e) are not developed in 
accordance with this section, EPA may determine that the sponsor has 
not fulfilled its obligations under 40 CFR part 79 and may require the 
sponsor to develop data in accordance with the requirements of this 
section in order to satisfy such obligations.
    (b) Organization and Personnel. (1) Personnel. (i) Each individual 
engaged in the conduct of or responsible for the supervision of a study 
shall have education, training, and experience, or combination thereof, 
to enable that individual to perform the assigned functions.
    (ii) Each testing facility shall maintain a current summary of 
training and experience and job description for each individual engaged 
in or supervising the conduct of a study.
    (iii) There shall be a sufficient number of personnel for the 
timely and proper conduct of the study according to the protocol.
    (iv) Personnel shall take necessary personal sanitation and health 
precautions designed to avoid contamination of test fuel and additive/
base fuel mixtures, test and reference substances, and test systems.
    (v) Personnel engaged in a study shall wear clothing appropriate 
for the duties they perform. Such clothing shall be changed as often as 
necessary to prevent microbiological, radiological, or chemical 
contamination of test systems and test, control, and reference 
substances.
    (vi) Any individual found at any time to have an illness that may 
adversely affect the quality and integrity of the study shall be 
excluded from direct contact with test systems, fuel and fuel/additive 
mixtures, test and reference substances and any other operation or 
function that may adversely affect the study until the condition is 
corrected. All personnel shall be instructed to report to their 
immediate supervisors any health or medical conditions that may 
reasonably be considered to have an adverse effect on a study.
    (2) Testing facility management. For each study, testing facility 
management shall:
    (i) Designate a study director as described in Sec. 79.60(b)(3) 
before the study is initiated.
    (ii) Replace the study director promptly if it becomes necessary to 
do so during the conduct of a study.
    (iii) Assure that there is a quality assurance unit as described in 
Sec. 79.60(b)(4).
    (iv) Assure that test fuels and fuel/additive mixtures and test and 
reference substances have been identified as to content, strength, 
purity, stability, and uniformity, as applicable.
    (v) Assure that personnel, resources, facilities, equipment, 
materials and methodologies are available as scheduled.
    (vi) Assure that personnel clearly understand the functions they 
are to perform.
    (vii) Assure that any deviations from these regulations reported by 
the quality assurance unit are communicated to the study director and 
corrective actions are taken and documented.
    (3) Study director. For each study, a scientist or other 
professional person with a doctorate degree or equivalent in toxicology 
or other appropriate discipline shall be identified as the study 
director. The study director has overall responsibility for the 
technical conduct of the study, as well as for the interpretation, 
analysis, documentation, and reporting of results, and represents the 
single point of study control. The study director shall assure that:
    (i) The protocol, including any changes, is approved as provided by 
Sec. 79.60(g)(1)(i) and is followed;
    (ii) All experimental data, including observations of unanticipated 
responses of the test system are accurately recorded and verified;
    (iii) Unforeseen circumstances that may affect the quality and 
integrity of the study are noted when they occur, and corrective action 
is taken and documented;
    (iv) Test systems are as specified in the protocol;
    (v) All applicable good laboratory practice regulations are 
followed; and
    (vi) All raw data, documentation, protocols, specimens, and final 
reports are archived properly during or at the close of the study.
    (4) Quality assurance unit. A testing facility shall have a quality 
assurance unit which shall be responsible for monitoring each study to 
assure management that the facilities, equipment, personnel, methods, 
practices, records, and controls are in conformance with the 
regulations in this section. For any given study, the quality assurance 
unit shall be entirely separate from and independent of the personnel 
engaged in the direction and conduct of that study. The quality 
assurance unit shall conduct inspections and maintain records 
appropriate to the study.
    (i) Quality assurance unit duties. (A) Maintain a copy of a master 
schedule sheet of all studies conducted at the testing facility indexed 
by test substance and containing the test system, nature of study, date 
study was initiated, current status of each study, identity of the 
sponsor, and name of the study director.
    (B) Maintain copies of all protocols pertaining to all studies for 
which the unit is responsible.
    (C) Inspect each study at intervals adequate to ensure the 
integrity of the study and maintain written and properly signed records 
of each periodic inspection showing the date of the inspection, the 
study inspected, the phase or segment of the study inspected, the 
person performing the inspection, findings and problems, action 
recommended and taken to resolve existing problems, and any scheduled 
date for re-inspection. Any problems which are likely to affect study 
integrity found during the course of an inspection shall be brought to 
the attention of the study director and management immediately.
    (D) Periodically submit to management and the study director 
written status reports on each study, noting any problems and the 
corrective actions taken.
    (E) Determine that no deviations from approved protocols or 
standard operating procedures were made without proper authorization 
and documentation.
    (F) Review the final study report to assure that such report 
accurately describes the methods and standard operating procedures, and 
that the reported results accurately reflect the raw data of the study.
    (G) Prepare and sign a statement to be included with the final 
study report which shall specify the dates inspections were made and 
findings reported to management and to the study director.
    (ii) The responsibilities and procedures applicable to the quality 
assurance unit, the records maintained by the quality assurance unit, 
and the method of indexing such records shall be in writing and shall 
be maintained. These items including inspection dates, the study 
inspected, the phase or segment of the study inspected, and the name of 
the individual performing the inspection shall be made available for 
inspection to authorized employees or duly designated representatives 
of EPA.
    (iii) An authorized employee or a duly designated representative of 
EPA shall have access to the written procedures established for the 
inspection and may request test facility management to certify that 
inspections are being implemented, performed, documented, and followed 
up in accordance with this paragraph.
    (c) Facilities--(1) General. Each testing facility shall be of 
suitable size and construction to facilitate the proper conduct of 
studies. Testing facilities which are not completely located within an 
indoor controlled environment shall be of suitable location/proximity 
to facilitate the proper conduct of studies. Testing facilities shall 
be designed so that there is a degree of separation that will prevent 
any function or activity from having an adverse effect on the study.
    (2) Test system care facilities. (i) A testing facility shall have 
a sufficient number of animal rooms or other test system areas, as 
needed, to ensure proper separation of species or test systems, 
quarantine or isolation of animals or other test systems, and routine 
or specialized housing of animals or other test systems.
    (ii) A testing facility shall have a number of animal rooms or 
other test system areas separate from those described in paragraph (a) 
of this section to ensure isolation of studies being done with test 
systems or test, control, and reference substances known to be 
biohazardous, including volatile atmospheres and aerosols, radioactive 
materials, and infectious agents. The animal handling facility must 
operate under the supervision of a veterinarian.
    (iii) Separate areas shall be provided, as appropriate, for the 
diagnosis, treatment, and control of laboratory test system diseases. 
These areas shall provide effective isolation for the housing of test 
systems either known or suspected of being diseased, or of being 
carriers of disease, from other test systems.
    (iv) Facilities shall have proper provisions for collection and 
disposal of contaminated air, water, or other spent materials. When 
animals are housed, facilities shall exist for the collection and 
disposal of all animal waste and refuse or for safe sanitary storage of 
waste before removal from the testing facility. Disposal facilities 
shall be so provided and operated as to minimize vermin infestation, 
odors, disease hazards, and environmental contamination.
    (v) Facilities shall have provisions to regulate environmental 
conditions (e.g., temperature, humidity, day length, etc.) as specified 
in the protocol.
    (3) Test system supply/operation areas. (i) There shall be storage 
areas, as needed, for feed, bedding, supplies, and equipment. Storage 
areas for feed and bedding shall be separated from areas where the test 
systems are located and shall be protected against infestation or 
contamination. Perishable supplies shall be preserved by appropriate 
means.
    (ii) Separate laboratory space and other space shall be provided, 
as needed, for the performance of the routine and specialized 
procedures required by studies.
    (4) Facilities for handling test fuels and fuel/additive mixtures 
and reference substances. (i) As necessary to prevent contamination or 
mixups, there shall be separate areas for:
    (A) Receipt and storage of the test fuels and fuel/additive 
mixtures and reference substances;
    (B) Mixing of the test fuels, fuel/additive mixtures, and reference 
substances with a carrier, i.e., liquid hydrocarbon; and
    (C) Storage of the test fuels, fuel/additive mixtures, and 
reference substance/carrier mixtures.
    (ii) Storage areas for test fuels and fuel/additive mixtures and 
reference substances and for reference mixtures shall be separate from 
areas housing the test systems and shall be adequate to preserve the 
identity, strength, purity, and stability of the substances and 
mixtures.
    (5) Specimen and data storage facilities. Space shall be secured 
for archives for the storage and retrieval of all raw data and 
specimens from completed studies.
    (d) Equipment--(1) Equipment design. Equipment used in the 
generation, measurement, or assessment of data and equipment used for 
facility environmental control shall be of appropriate design and 
adequate capacity to function according to the protocol and shall be 
suitably located for operation, inspection, cleaning, and maintenance.
    (2) Maintenance and calibration of equipment. (i) Equipment shall 
be adequately inspected, cleaned, and maintained. Equipment used for 
the generation, measurement, or assessment of data shall be adequately 
tested, calibrated, and/or standardized.
    (ii) The written standard operating procedures required under 
Sec. 79.60(e)(1)(ii)(K) shall set forth in sufficient detail the 
methods, materials, and schedules to be used in the routine inspection, 
cleaning, maintenance, testing, calibration, and/or standardization of 
equipment, and shall specify, when appropriate, remedial action to be 
taken in the event of failure or malfunction of equipment. The written 
standard operating procedures shall designate the person responsible 
for the performance of each operation.
    (iii) Written records shall be maintained of all inspection, 
maintenance, testing, calibrating, and/or standardizing operations. 
These records, containing the date of the operation, shall describe 
whether the maintenance operations were routine and followed the 
written standard operating procedures. Written records shall be kept of 
non-routine repairs performed on equipment as a result of failure and 
malfunction. Such records shall document the nature of the defect, how 
and when the defect was discovered, and any remedial action taken in 
response to the defect.
    (e) Testing Facilities Operation--(1) Standard operating 
procedures. (i) A testing facility shall have standard operating 
procedures in writing, setting forth study methods that management is 
satisfied are adequate to insure the quality and integrity of the data 
generated in the course of a study. All deviations in a study from 
standard operating procedures shall be authorized by the study director 
and shall be documented in the raw data. Significant changes in 
established standard operating procedures shall be properly authorized 
in writing by management.
    (ii) Standard operating procedures shall be established for, but 
not limited to, the following:
    (A) Test system room preparation;
    (B) Test system care;
    (C) Receipt, identification, storage, handling, mixing, and method 
of sampling of test fuels and fuel/additive mixtures and reference 
substances;
    (D) Test system observations;
    (E) Laboratory or other tests;
    (F) Handling of test animals found moribund or dead during study;
    (G) Necropsy or postmortem examination of test animals;
    (H) Collection and identification of specimens;
    (I) Histopathology
    (J) Data handling, storage and retrieval.
    (K) Maintenance and calibration of equipment.
    (L) Transfer, proper placement, and identification of test systems.
    (iii) Each laboratory or other study area shall have immediately 
available manuals and standard operating procedures relative to the 
laboratory procedures being performed. Published literature may be used 
as a supplement to standard operating procedures.
    (iv) A historical file of standard operating procedures, and all 
revisions thereof, including the dates of such revisions, shall be 
maintained.
    (2) Reagents and solutions. All reagents and solutions in the 
laboratory areas shall be labeled to indicate identity, titer or 
concentration, storage requirements, and expiration date. Deteriorated 
or outdated reagents and solutions shall not be used.
    (3) Animal and other test system care. (i) There shall be standard 
operating procedures for the housing, feeding, handling, and care of 
animals and other test systems.
    (ii) All newly received test systems from outside sources shall be 
isolated and their health status or appropriateness for the study shall 
be evaluated. This evaluation shall be in accordance with acceptable 
veterinary medical practice or scientific methods.
    (iii) At the initiation of a study, test systems shall be free of 
any disease or condition that might interfere with the purpose or 
conduct of the study. If during the course of the study, the test 
systems contract such a disease or condition, the diseased test systems 
shall be isolated, if necessary. These test systems may be treated for 
disease or signs of disease provided that such treatment does not 
interfere with the study. The diagnosis, authorization of treatment, 
description of treatment, and each date of treatment shall be 
documented and shall be retained.
    (iv) When laboratory procedures require test animals to be 
manipulated and observed over an extended period of time or when 
studies require test animals to be removed from and returned to their 
housing units for any reason (e.g., cage cleaning, treatment, etc.), 
these test systems shall receive appropriate identification (e.g., 
tattoo, color code, etc.). Test system identification shall conform 
with current laboratory animal handling practice. All information 
needed to specifically identify each test system within the test 
system-housing unit shall appear on the outside of that unit. Suckling 
animals are excluded from the requirement of individual identification 
unless otherwise specified in the protocol.
    (v) Except as specified in paragraph (e)(3)(v)(A) of this section, 
test animals of different species shall be housed in separate rooms 
when necessary. Test animals of the same species, but used in different 
studies, shall not ordinarily be housed in the same room when 
inadvertent exposure to the test or reference substances or test system 
mixup could affect the outcome of either study. If such mixed housing 
is necessary, adequate differentiation by space and identification 
shall be made.
    (A) Test systems that may be used in multispecies tests need not be 
housed in separate rooms, provided that they are adequately segregated 
to avoid mixup and cross-contamination.
    (B) [reserved]
    (vi) Cages, racks, pens, enclosures, and other holding, rearing, 
and breeding areas, and accessory equipment, shall be cleaned and 
sanitized at appropriate intervals.
    (vii) Feed and water used for the test animals shall be analyzed 
periodically to ensure that contaminants known to be capable of 
interfering with the study and reasonably expected to be present in 
such feed or water are not present at greater than trace levels. 
Documentation of such analyses shall be maintained as raw data.
    (viii) Bedding used in animal cages or pens shall not interfere 
with the purpose or conduct of the study and shall be changed as often 
as necessary to keep the animals dry and clean.
    (ix) If any pest control materials are used, the use shall be 
documented. Cleaning and pest control materials that interfere with the 
study shall not be used.
    (x) All test systems shall be acclimatized to the environmental 
conditions of the test, prior to their use in a study.
    (f) Test fuels, additive/base fuel mixtures, and reference 
substances--(1) Test fuel, fuel/additive mixture, and reference 
substance identity (i) The product brand name/service mark, strength, 
purity, content, or other characteristics which appropriately define 
the test fuel, fuel/additive mixture, or reference substance shall be 
reported for each batch and shall be documented before its use in a 
study. Methods of synthesis, fabrication, or derivation, as 
appropriate, of the test fuel, fuel/additive mixture, or reference 
substance shall be documented by the sponsor or the testing facility, 
and such location of documentation shall be specified.
    (ii) The stability of test fuel, fuel/additive mixture, and 
reference substances under storage conditions at the test site shall be 
known for all studies.
    (2) Test fuel, additive/base fuel mixture, and reference substance 
handling. Procedures shall be established for a system for the handling 
of the test fuel, fuel/additive mixture, and reference substance(s) to 
ensure that:
    (i) There is proper storage.
    (ii) Distribution is made in a manner designed to preclude the 
possibility of contamination, deterioration, or damage.
    (iii) Proper identification is maintained throughout the 
distribution process.
    (iv) The receipt and distribution of each batch is documented. Such 
documentation shall include the date and quantity of each batch 
distributed or returned.
    (3) Mixtures of test emissions or reference solutions with 
carriers.
    (i) For test emissions or each reference substance mixed with a 
carrier, tests by appropriate analytical methods shall be conducted:
    (A) To determine the uniformity of the test substance and to 
determine, periodically, the concentration of the test emissions or 
reference substance in the mixture;
    (B) When relevant to the conduct of the experiment, to determine 
the solubility of each reference substance in the carrier mixture 
before the experimental start date; and
    (C) To determine the stability of test emissions or a reference 
solution in the test substance before the experimental start date or 
concomitantly according to written standard operating procedures, which 
provide for periodic analysis of each batch.
    (ii) Where any of the components of the reference substance/carrier 
mixture has an expiration date, that date shall be clearly shown on the 
container. If more than one component has an expiration date, the 
earliest date shall be shown.
    (iii) If a chemical or physical agent is used to facilitate the 
mixing of a test substance with a carrier, assurance shall be provided 
that the agent does not interfere with the integrity of the test.
    (g) Protocol for and conduct of a study--(1) Protocol. (i) Each 
study shall have a written protocol that clearly indicates the 
objectives and all methods for the conduct of the study. The protocol 
shall contain but shall not be limited to the following information:
    (A) A descriptive title and statement of the purpose of the study.
    (B) Identification of the test fuel, fuel/additive mixture, and 
reference substance by name, chemical abstracts service (CAS) number or 
code number, as applicable.
    (C) The name and address of the sponsor and the name and address of 
the testing facility at which the study is being conducted.
    (D) The proposed experimental start and termination dates.
    (E) Justification for selection of the test system, as necessary.
    (F) Where applicable, the number, body weight, sex, source of 
supply, species, strain, substrain, and age of the test system.
    (G) The procedure for identification of the test system.
    (H) A description of the experimental design, including methods for 
the control of bias.
    (I) Where applicable, a description and/or identification of the 
diet used in the study. The description shall include specifications 
for acceptable levels of contaminants that are reasonably expected to 
be present in the dietary materials and are known to be capable of 
interfering with the purpose or conduct of the study if present at 
levels greater than established by the specifications.
    (J) Each concentration level, expressed in milligrams per cubic 
meter of air or other appropriate units, of the test or reference 
substance to be administered and the frequency of administration.
    (K) The type and frequency of tests, analyses, and measurements to 
be made.
    (L) The records to be maintained.
    (M) The date of approval of the protocol by the sponsor and the 
dated signature of the study director.
    (N) A statement of the proposed statistical method.
    (ii) All changes in or revisions of an approved protocol and the 
reasons therefor shall be documented, signed by the study director, 
dated, and maintained with the protocol.
    (2) Conduct of a study. (i) The study shall be conducted in 
accordance with the protocol.
    (ii) The test systems shall be monitored in conformity with the 
protocol.
    (iii) Specimens shall be identified by test system, study, nature, 
and date of collection. This information shall be located on the 
specimen container or shall accompany the specimen in a manner that 
precludes error in the recording and storage of data.
    (iv) In animal studies where histopathology is required, records of 
gross findings for a specimen from postmortem observations shall be 
available to a pathologist when examining that specimen 
histopathologically.
    (v) All data generated during the conduct of a study, except those 
that are generated by automated data collection systems, shall be 
recorded directly, promptly, and legibly in ink. All data entries shall 
be dated on the day of entry and signed or initialed by the person 
entering the data. Any change in entries shall be made so as not to 
obscure the original entry, shall indicate the reason for such change, 
and shall be dated and signed or identified at the time of the change. 
In automated data collection systems, the individual responsible for 
direct data input shall be identified at the time of data input. Any 
change in automated data entries shall be made so as not to obscure the 
original entry, shall indicate the reason for change, shall be dated, 
and the responsible individual shall be identified.
    (h) Records and Reports--(1) Reporting of study results. (i) A 
final report shall be prepared for each study and shall include, but 
not necessarily be limited to, the following:
    (A) Name and address of the facility performing the study and the 
dates on which the study was initiated and was completed, terminated, 
or discontinued.
    (B) Objectives and procedures stated in the approved protocol, 
including any changes in the original protocol.
    (C) Statistical methods employed for analyzing the data.
    (D) The test fuel, additive/base fuel mixture, and test and 
reference substances identified by name, chemical abstracts service 
(CAS) number or code number, strength, purity, content, or other 
appropriate characteristics.
    (E) Stability, and when relevant to the conduct of the study, the 
solubility of the test emissions and reference substances under the 
conditions of administration.
    (F) A description of the methods used.
    (G) A description of the test system used. Where applicable, the 
final report shall include the number of animals or other test 
organisms used, sex, body weight range, source of supply, species, 
strain and substrain, age, and procedure used for identification.
    (H) A description of the concentration regimen as daily exposure 
period, i.e., number of hours, and exposure duration, i.e., number of 
days.
    (I) A description of all circumstances that may have affected the 
quality or integrity of the data.
    (J) The name of the study director, the names of other scientists 
or professionals and the names of all supervisory personnel, involved 
in the study.
    (K) A description of the transformations, calculations, or 
operations performed on the data, a summary and analysis of the data, 
and a statement of the conclusions drawn from the analysis.
    (L) The signed and dated reports of each of the individual 
scientists or other professionals involved in the study, including each 
person who, at the request or direction of the testing facility or 
sponsor, conducted an analysis or evaluation of data or specimens from 
the study after data generation was completed.
    (M) The locations where all specimens, raw data, and the final 
report are to be kept or stored.
    (N) The statement, prepared and signed by the quality assurance 
unit, as described in Sec. 79.60(b)(4)(i)(G).
    (ii) The final report shall be signed and dated by the study 
director.
    (iii) Corrections or additions to a final report shall be in the 
form of an amendment by the study director. The amendment shall clearly 
identify that part of the final report that is being added to or 
corrected and the reasons for the correction or addition, and shall be 
signed and dated by the person responsible. Modification of a final 
report to comply with the submission requirements of EPA does not 
constitute a correction, addition, or amendment to a final report.
    (iv) A copy of the final report and of any amendment to it shall be 
maintained by the sponsor and the test facility.
    (2) Storage and retrieval of records and data. (i) All raw data, 
documentation, records, protocols, specimens, and final reports 
generated as a result of a study shall be retained. Specimens obtained 
from mutagenicity tests, wet specimens of blood, urine, feces, and 
biological fluids, do not need to be retained after quality assurance 
verification. Correspondence and other documents relating to 
interpretation and evaluation of data, other than those documents 
contained in the final report, also shall be retained.
    (ii) All raw data, documentation, protocols, specimens, and interim 
and final reports shall be archived for orderly storage and expedient 
retrieval. Conditions of storage shall minimize deterioration of the 
documents or specimens in accordance with the requirements for the time 
period of their retention and the nature of the documents of specimens. 
A testing facility may contract with commercial archives to provide a 
repository for all material to be retained. Raw data and specimens may 
be retained elsewhere provided that the archives have specific 
reference to those other locations.
    (iii) An individual shall be identified as responsible for the 
archiving of records.
    (iv) Access to archived material shall require authorization and 
documentation.
    (v) Archived material shall be indexed to permit expedient 
retrieval.
    (3) Retention of records. (i) Record retention requirements set 
forth in this section do not supersede the record retention 
requirements of any other regulations in this subchapter.
    (ii) Except as provided in paragraph (h)(3)(iii) of this section, 
documentation records, raw data, and specimens pertaining to a study 
and required to be retained by this part shall be archived for a period 
of at least ten years following the completion of the study.
    (iii) Wet specimens, samples of test fuel, additive/base fuel 
mixtures, or reference substances, and specially prepared material 
which are relatively fragile and differ markedly in stability and 
quality during storage, shall be retained only as long as the quality 
of the preparation affords evaluation. Specimens obtained from 
mutagenicity tests, wet specimens of blood, urine, feces, biological 
fluids, do not need to be retained after quality assurance 
verification. In no case shall retention be required for a longer 
period than that set forth in paragraph (h)(3)(ii) of this section.
    (iv) The master schedule sheet, copies of protocols, and records of 
quality assurance inspections, as required by Sec. 79.60(b)(4)(iii) 
shall be maintained by the quality assurance unit as an easily 
accessible system of records for the period of time specified in 
paragraph (h)(3)(ii) of this section.
    (v) Summaries of training and experience and job descriptions 
required to be maintained by Sec. 79.60(b)(1)(ii) may be retained along 
with all other testing facility employment records for the length of 
time specified in paragraph (h)(3)(ii) of this section.
    (vi) Records and reports of the maintenance and calibration and 
inspection of equipment, as required by Sec. 79.60(d)(2) (ii) and 
(iii), shall be retained for the length of time specified in paragraph 
(h)(3)(ii) of this section.
    (vii) If a facility conducting testing or an archive contracting 
facility goes out of business, all raw data, documentation, and other 
material specified in this section shall be transferred to the sponsor 
of the study for archival.
    (viii) Records required by this section may be retained either as 
original records or as true copies such as photocopies, microfilm, 
microfiche, or other accurate reproductions of the original records.


Sec. 79.61.  Vehicle emissions inhalation exposure guideline.

    (a) Purpose. This guideline provides additional information on 
methodologies required to conduct health effects tests involving 
inhalation exposures to vehicle combustion emissions from fuels or 
fuel/additive mixtures. Where this guideline and the other health 
effects testing guidelines in 40 CFR 79.62 through 79.68 specify 
differing values for the same test parameter, the specifications in the 
individual health test guideline shall prevail for that health effect 
endpoint.
    (b) Definitions. For the purposes of this section the following 
definitions apply.
    Acute inhalation study means a short-term toxicity test 
characterized by a single exposure by inhalation over a short period of 
time (at least 4 hours and less than 24 hours), followed by at least 14 
days of observation.
    Aerodynamic diameter means the diameter of a sphere of unit density 
that has the same settling velocity as the particle of the test 
substance. It is used to compare particles of different sizes, 
densities and shapes, and to predict where in the respiratory tract 
such particles may be deposited. It applies to the size of aerosol 
particles.
    Chronic inhalation study means a prolonged and repeated exposure by 
inhalation for the life span of the test animal; technically, two years 
in the rat.
    Concentration means an exposure level. Exposure is expressed as 
weight or volume of test aerosol/substance per volume of air, usually 
mg/m3 or as parts per million (ppm) over a given time period. 
Micrograms per cubic meter (g/m3) or parts per billion 
may be appropriate, as well.
    Cumulative toxicity means the adverse effects of repeated exposures 
occurring as a result of prolonged action or increased concentration of 
the administered test substance or its metabolites in the susceptible 
tissues.
    Inhalable diameter means that aerodynamic diameter of a particle 
which is considered to be inhalable for the organism. It is used to 
refer to particles which are capable of being inhaled and may be 
deposited anywhere within the respiratory tract from the trachea to the 
alveoli.
    Mass median aerodynamic diameter (MMAD) means the calculated 
aerodynamic diameter, which divides the particles of an aerosol in half 
based on the mass of the particles. Fifty percent of the particles in 
mass will be larger than the median diameter, and fifty percent will be 
smaller than the median diameter. MMAD describes the particle 
distribution of any aerosol based on the weight and size of the 
particles. MMAD and the geometric standard deviation describe the 
particle-size distribution.
    Material safety data sheet (MSDS) means documentation or 
information on the physical, chemical, and hazardous characteristics of 
a given chemical, usually provided by the product's manufacturer.
    Reynolds number means a dimensionless number that is proportional 
to the ratio of inertial forces to frictional forces acting on a fluid. 
It quantitatively provides a measure of whether flow is laminar or 
turbulent. A fluid traveling through a pipe is fully developed into a 
laminar flow for a Reynolds number less than 2000, and fully developed 
into a turbulent flow for a Reynolds number greater than 4000.
    Subacute inhalation toxicity means the adverse effects occurring as 
a result of the repeated daily exposure of experimental animals to a 
chemical by inhalation for part (less than 10 percent) of a lifespan; 
generally, less than 90 days.
    Subchronic inhalation study means a repeated exposure by inhalation 
for part (approximately 10 percent) of a life span of the exposed test 
animal.
    Toxic effect means an adverse change in the structure or function 
of an experimental animal as a result of exposure to a chemical 
substance.
    (c) Principles and design criteria of inhalation exposure systems. 
Proper conduct of inhalation toxicity studies of the emissions of fuels 
and additive/fuel mixtures requires that the exposure system be 
designed to ensure the controlled generation of the exposure 
atmosphere, the adequate dilution of the test emissions, delivery of 
the diluted exposure atmosphere to the test animals, and use of 
appropriate exposure chamber systems selected to meet criteria for a 
given exposure study.
    (1) Emissions generation. Emissions shall be generated according to 
the specifications in 40 CFR 79.57.
    (2) Dilution and delivery systems.
    (i) The delivery system is the means used to transport the 
emissions from the generation system to the exposure system. The 
dilution system is generally a component of the delivery system.
    (ii) Dilution provides control of the emissions concentration 
delivered to the exposure system, serving the function of diluting the 
associated combustion gases, such as carbon monoxide, carbon dioxide, 
nitrogen oxides, sulfur dioxide and other noxious gases and vapors, to 
levels that will ensure that there are no significant or measurable 
responses in the test animals as a result of exposure to the combustion 
gases. The formation of particle species is strongly dependent on the 
dilution rate, as well.
    (iii) The engine exhaust system shall connect to the first-stage-
dilution section at 90 deg. to the axis of the dilution section. This 
is then connected to a right angle elbow on the center line of the 
dilution section. Engine emissions are injected through the elbow so 
that exhaust flow is concurrent to dilution flow.
    (iv) Materials. In designing the dilution and delivery systems, the 
use of plastic, e.g., PVC and similar materials, copper, brass, and 
aluminum pipe and tubing shall be avoided if there exists a possibility 
of chemical reaction occurring between emissions and tubing. Stainless 
steel pipe and tubing is recommended as the best choice for most 
emission dilution and delivery applications, although glass and teflon 
may be appropriate, as well.
    (v) Flow requirements. (A) Conduit for dilute raw emissions shall 
be of such dimensions as to provide residence times for the emissions 
on the order of less than one second to several seconds before the 
emissions are further diluted and introduced to the test chambers. With 
the high flow rates in the dilute raw emissions conduit, it will be 
necessary to sample various portions of the dilute emissions for 
delivering differing concentrations to the test chambers. The unused 
portions of the emissions stream are normally exhausted to the 
atmosphere outside of the exposure facility.
    (B) Dimensions of the dilute raw exhaust conduit shall be such 
that, at a minimum, the flow Reynolds number is 70,000 or greater (see 
Mokler, et al., 1984 in paragraph (f)(13) of this section). This will 
maintain highly turbulent flow conditions so that there is more 
complete mixing of the exhaust emissions.
    (C) Wall losses. The delivery system shall be designed to minimize 
wall losses. This can be done by sizing the tubing or pipe to maintain 
laminar flow of the diluted emissions to the exposure chamber. A flow 
Reynolds number of 1000-3000 will ensure minimal wall losses. Also, the 
length of and number and degree of bends in the delivery lines to the 
exposure chamber system shall be minimized.
    (D) Whole-body exposure vs. nose-only exposure delivery systems. 
Flow rates through whole-body chamber systems are of the order of 100 
liters per minute to 500 liters per minute. Nose-only systems are on 
the order of less than 50 liters per minute. To maintain laminar flow 
conditions, the principles described in paragraph (c)(2)(v)(C) of this 
section apply to both systems.
    (vi) Dilution requirements. (A) To maintain the water vapor, and 
dissolved organic compounds, in the raw exhaust emissions stream, a 
manufacturer/tester will initially dilute one part emissions with a 
minimum of five parts clean, filtered air (see Hinners, et al., 1979 in 
paragraph (f)(11) of this section). Depending on the water vapor 
content of a particular fuel/additive mixture's combustion emissions 
and the humidity of the dilution air, initial exhaust dilutions as high 
as 1:15 or 1:20 may be necessary to maintain the general character of 
the exhaust as it cools, e.g., M100. At this point, it is expected that 
the exhaust stream would be further diluted to more appropriate levels 
for rodent health effects testing.
    (B) A maximum concentration (minimum dilution) of the raw exhaust 
going into the test animal cages is anticipated to lie in the range 
between 1:5 and 1:50 exhaust emissions to clean, filtered air. The 
minimum concentration (maximum dilution) of raw exhaust for health 
effects testing is anticipated to be in range between 1:100 and 1:150. 
Individual manufacturers will treat these ranges as approximations only 
and will determine the optimum range of emission concentrations to 
elicit effects in Tier 2 health testing for their particular fuel/fuel 
additive mixture.
    (3) Exposure chamber systems--(i) Referenced Guidelines. (A) The 
U.S. Department of Health and Human Services ``Guide for the Care and 
Use of Laboratory Animals'' (Guide), 1985 cited in paragraph 
(c)(3)(ii)(A)(4), and in paragraphs (d)(2)(i), (d)(2)(ii), (d)(2)(iii), 
(d)(4)(ii), and (d)(4)(iii) of this section, has been incorporated by 
reference.
    (B) This incorporation by reference was approved by the Director of 
the Federal Register in accordance with 5 U.S.C. 552(a) and 1 CFR part 
51. Copies may be purchased from the Superintendent of Documents, U.S. 
Government Printing Office, Washington, DC 20402. Copies may be 
inspected at U.S. EPA, OAR, 401 M Street SW., Washington, DC, 20460 or 
at the Office of the Federal Register, 800 North Capitol Street NW., 
suite 700, Washington, DC.
    (ii) Exposure chambers. There are two basic types of dynamic 
inhalation exposure chambers, whole-body chambers and nose-/head-only 
exposure chambers (see Cheng and Moss, 1989 in paragraph (f)(8) of this 
section).
    (A) Whole-body chambers. (1) The flow rate through a chamber shall 
be maintained at 15 air changes per hour.
    (2) The chambers are usually maintained at a slightly negative 
pressure (0.5 to 1.5 inch of water) to prevent leakage of test 
substance into the exposure room.
    (3) The exposure chamber shall be designed in such a way as to 
provide uniform distribution of exposure concentrations in all 
compartments (see Cheng et al., 1989 in paragraph (f)(7) of this 
section).
    (4) Animals are housed in separate compartments inside the chamber, 
where the whole surface area of an animal is exposed to the test 
material. The spaces required for different animal species shall follow 
the Guide. In general, the volume of animal bodies occupy less than 5 
percent of the chamber volume.
    (B) Head/nose-only exposure chambers. (1) In head/nose-only 
exposure chambers, only the head (oronasal) portion of the animal is 
exposed to the test material.
    (2) The chamber volume and flow rates are much less than in the 
whole-body exposure chambers because the subjects are usually 
restrained in a tube holder where the animal's breathing can be easily 
monitored. The head/nose-only exposure chamber is suitable for short-
term exposures or when use of a small amount of test material is 
required.
    (iii) Since whole-body exposure appears to be the least stressful 
mode of exposure, it is the preferred method. In general, head/nose 
only exposure, which is sometimes used to avoid concurrent exposure by 
the dermal or oral routes, i.e., grooming, is not recommended because 
of the stress accompanying the restraining of the animals. However, 
there may be specific instances where it may be more appropriate than 
whole-body exposure. The tester shall provide justification for its 
selection.
    (d) Inhalation exposure procedures--(1) Animal selection. (i) The 
rat is the preferred species for vehicle emission inhalation health 
effects testing. Commonly used laboratory strains shall be used. Any 
rodent species may be used, but the tester shall provide justification 
for the choice of that species.
    (ii) Young adult animals, approximately ten weeks of age for the 
rat, shall be used. At the commencement of the study, the weight 
variation of animals used shall not exceed 20 percent of 
the mean weight for each sex. Animals shall be randomly assigned to 
treatment and control groups according to their weight.
    (iii) An equal number of male and female rodents shall be used at 
each concentration level. Situations may arise where use of a single 
sex may be appropriate. Females, in general, shall be nulliparous and 
nonpregnant.
    (iv) The number of animals used at each concentration level and in 
the control group(s) depends on the type of study, number of biological 
end points used in the toxicity evaluation, the pre-determined 
sensitivity of detection and power of significance of the study, and 
the animal species. For an acute study, at least five animals of each 
sex shall be used in each test group. For both the subacute and 
subchronic studies, at least 10 rodents of each sex shall be used in 
each test group. For a chronic study, at least 20 male and 20 female 
rodents shall be used in each test group.
    (A) If interim sacrifices are planned, the number of animals shall 
be increased by the number of animals scheduled to be sacrificed during 
the course of the study.
    (B) For a chronic study, the number of animals at the termination 
of the study must be adequate for a meaningful and valid statistical 
evaluation of chronic effects.
    (v) A concurrent control group is required. This group shall be 
exposed to clean, filtered air under conditions identical to those used 
for the group exposed to the test atmosphere.
    (vi) The same species/strain shall be used to make comparisons 
between fuel-only and fuel/additive mixture studies. If another 
species/strain is used, the tester shall provide justification for its 
selection.
    (2) Animal handling and care. (i) A key element in the conduct of 
inhalation exposure studies is the proper handling and care of the test 
animal population. Therefore, the exposure conditions must conform 
strictly with the conditions for housing and animal care and use set 
forth in the Guide.
    (ii) In whole-body exposure chambers, animals shall be housed in 
individual caging. The minimum cage size per animal will be in 
accordance with instructions set forth in the Guide.
    (iii) Chambers shall be cleaned and maintained in accordance with 
recommendations and schedules set forth in the Guide.
    (A) Observations shall be made daily with appropriate actions taken 
to minimize loss of animals to the study (e.g., necropsy or 
refrigeration of animals found dead and isolation or sacrifice of weak 
or moribund animals). Exposure systems using head/nose-only exposure 
chambers require no special daily chamber maintenance. Chambers shall 
be inspected to ensure that they are clean, and that there are no 
obstructions in the chamber which would restrict air flow to the 
animals. Whole-body exposure chambers will be inspected on a minimum of 
twice daily, once before exposures and once after exposures.
    (B) Signs of toxicity shall be recorded as they are observed, 
including the time of onset, degree, and duration.
    (C) Cage-side observations shall include, but are not limited to: 
changes in skin, fur, eye and mucous membranes, respiratory, autonomic, 
and central nervous systems, somatomotor activity, and behavioral 
patterns. Particular attention shall be directed to observation of 
tremors, convulsions, salivation, diarrhea, lethargy, sleep, and coma.
    (iv) Food and water will be withheld from animals for head/nose-
only exposure systems. For whole-body-exposure systems, water only may 
be provided. When the exposure generation system is not operating, food 
will be available ad libitum. During operation of the generation 
system, food will be withheld to avoid possible contamination by 
emissions.
    (v) At the end of the study period, all survivors in the main study 
population shall be sacrificed. Moribund animals shall be removed and 
sacrificed when observed.
    (3) Concentration levels and selection. (i) In acute and subacute 
toxicity tests, at least three exposure concentrations and a control 
group shall be used and spaced appropriately to produce test groups 
with a range of toxic effects and mortality rates. The data shall be 
sufficient to produce a concentration-response curve and permit an 
acceptable estimation of the median lethal concentration.
    (ii) In subchronic and chronic toxicity tests, testers shall use at 
least three different concentration levels, with a control exposure 
group, to determine a concentration-response relationship. 
Concentrations shall be spaced appropriately to produce test groups 
with a range of toxic effects. The concentration-response data may also 
be sufficient to determine a NOAEL, unless the result of a limit test 
precludes such findings. The criteria for selecting concentration 
levels has been published (40 CFR 798.2450 and 798.3260).
    (A) The highest concentration shall result in toxic effects but not 
produce an incidence of fatalities which would prevent a meaningful 
evaluation of the study.
    (B) The lowest concentration shall not produce toxic effects which 
are directly attributable to the test exposure. Where there is a useful 
estimation of human exposure, the lowest concentration shall exceed 
this.
    (C) The intermediate concentration level(s) shall produce minimal 
observable toxic effects. If more than one intermediate concentration 
level is used, the concentrations shall be spaced to produce a 
gradation of toxic effects.
    (D) In the low, intermediate, and control exposure groups, the 
incidence of fatalities shall be low to absent, so as not to preclude a 
meaningful evaluation of the results.
    (4) Exposure chamber environmental conditions. The following 
environmental conditions in the exposure chamber are critical to the 
maintenance of the test animals: flow; temperature; relative humidity; 
lighting; and noise.
    (i) Filtered and conditioned air shall be used during exposure, to 
dilute the exhaust emissions, and during non- exposure periods to 
maintain environmental conditions that are free of trace gases, dusts, 
and microorganisms on the test animals. Twelve to fifteen air changes 
per hour will be provided at all times to whole-body-exposure chambers. 
The minimum air flow rate for head/nose-only exposure chambers will be 
a function of the number of animals and the average minute volume of 
the animals:

Qminimum(L/min)=2  x  number of animals  x  average minute volume

(see Cheng and Moss, 1989 in paragraph (f)(8) of this section).
    (ii) Recommended ranges of temperature for various species are 
given in the Guide. The recommended temperature ranges will be used for 
establishing temperature conditions of whole-body- exposure chambers. 
For rodents in whole-body-exposure chambers, the recommended 
temperature is 22  deg.C +/- 2  deg.C and for rabbits, it is 20  deg.C 
+/- 3  deg.C. Temperature ranges have not been established for head/
nose-only tubes; however, recommended maximum temperature limits have 
been established at the Inhalation Toxicology Research Institute (see 
Barr, 1988 in paragraph (f)(1) of this section). Maximum temperature 
for rats and mice in head/nose-only tubes is 23  deg.C.
    (iii) Relative humidity. The relative humidity in the chamber air 
is important for heat balance and shall be maintained between 40 
percent and 60 percent, but in certain instances, this may not be 
practicable. Testers shall follow Guide recommends for a 30 percent to 
70 percent relative humidity range for rodents in exposure chambers.
    (iv) Lighting. Light intensity of 30 foot candles at 3 ft. from the 
floor of the exposure facility is recommended (see Rao, 1986 in 
paragraph (f)(16) of this section).
    (5) Exposure Conditions. Study animals shall be exposed to the test 
atmosphere on a repeated basis for at least 6 hours per day on a 7-day 
per week basis for the exposure period. However, based primarily on 
practical considerations, exposure on a 5-day-per-week basis for a 
minimum of 6 hours per day is the minimum acceptable exposure period.
    (6) Exposure atmosphere. (i) The exposure atmosphere shall be held 
as constant as is practicable and must be monitored continuously or 
intermittently, depending on the method of analysis, to ensure that 
exposure levels are at the target values or within stated limits during 
the exposure period. Sampling methodology will be determined based on 
the type of generation system and the type of exposure chamber system 
specified for the exposure study.
    (A) Integrated samples of test atmosphere aerosol shall be taken 
daily during the exposure period from a single representative sample 
port in the chamber near the breathing zone of the animals. Gas samples 
shall be taken daily to determine concentrations (ppm) of the major 
vapor components of the test atmosphere including CO, CO2, 
NOX, SO2, and total hydrocarbons.
    (B) To ensure that animals in different locations of the chamber 
receive a similar exposure atmosphere, distribution of an aerosol or 
vapor concentration in exposure chambers can be determined without 
animals during the developmental phase of the study, or it can be 
determined with animals early in the study. For head/nose-only exposure 
chambers, it may not be possible to monitor the chamber distribution 
during the exposure, because the exposure port contains the animal.
    (C) During the development of the emissions generation system, 
particle size analysis shall be performed to establish the stability of 
an aerosol concentration with respect to particle size. Over the course 
of the exposure, analysis shall be conducted as often as is necessary 
to determine the consistency of particle size distribution.
    (D) Chamber rise and fall times. The rise time required for the 
exposure concentration to reach 90 percent of the stable concentration 
after the generator is turned on, and the fall time when the chamber 
concentration decreases to 10 percent of the stable concentration after 
the generation system is stopped shall be determined in the 
developmental phase of the study. Time-integrated samples collected for 
calculating exposure concentrations shall be taken after the rise time. 
The daily exposure time is exclusive of the rise or the fall time.
    (ii) Instrumentation used for a given study will be determined 
based on the type of generation system and the type of exposure chamber 
system specified for the exposure study.
    (A) For exhaust studies, combustion gases shall be sampled by 
collecting exposure air in bags and then analyzing the collected air 
sample to determine major components of the combustion gas using gas 
analyzers. Exposure chambers can also be connected to gas analyzers 
directly by using sampling lines and switching valves. Samples can be 
taken more frequently using the latter method. Aerosol instruments, 
such as photometers, or time-integrated gravimetric determination may 
be used to determine the stability of any aerosol concentration in the 
chamber.
    (B) For evaporative emission studies, concentration of fuel vapors 
can usually be determined by using a gas chromatograph (GC) and/or 
infrared (IR) spectrometry. Grab samples for intermittent sampling can 
be taken from the chamber by using bubble samplers with the appropriate 
solvent to collect the vapors, or by collecting a small volume of air 
in a syringe. Intermediate or continuous monitoring of the chamber 
concentration is also possible by connecting the chamber with a GC or 
IR detector.
    (7) Monitoring chamber environmental conditions may be performed by 
a computer system or by exposure system operating personnel.
    (i) The flow-metering device used for the exposure chambers must be 
a continuous monitoring device, and actual flow measurements must be 
recorded at least every 30 minutes. Accuracy must be 5 
percent of full scale range. Measurement of air flow through the 
exposure chamber may be accomplished using any device that has 
sufficient range to accurately measure the air flow for the given 
chamber. Types of flow metering devices include rotameters, orifice 
meters, venturi meters, critical orifices, and turbinemeters (see 
Benedict, 1984 in paragraph (f)(4) and Spitzer, 1984 in paragraph 
(f)(17) of this section).
    (ii) Pressure. Pressure measurement may be accomplished using 
manometers, electronic pressure transducers, magnehelics, or similar 
devices (see Gillum, 1982 in paragraph (f)(10) of this section). 
Accuracy of the pressure device must be 5 percent of full 
scale range. Pressure measurements must be continuous and recorded at 
least every 30 minutes.
    (iii) Temperature. The temperature of exposure chambers must be 
monitored continuously and recorded at least every 30 minutes. 
Temperature may be measured using thermometers, RTD's, thermocouples, 
thermistors, or other devices (see Benedict, 1984 in paragraph (f)(4) 
of this section). It is necessary to incorporate an alarm system into 
the temperature monitoring system. The exposure operators must be 
notified by the alarm system when the chamber temperature exceeds 26.7 
deg.C (80  deg.F). The exposure must be discontinued and emergency 
procedures enacted to immediately reduce temperatures or remove test 
animals from high temperature environment when chamber temperatures 
exceed 29  deg.C. Accuracy of the temperature monitoring device will be 
1  deg.C for the temperature range of 20-30  deg.C.
    (iv) Relative humidity. The relative humidity of exposure chambers 
must be monitored continuously and recorded at least every 30 minutes. 
Relative humidity may be measured using various devices (see Chaddock, 
1985 in paragraph (f)(6) of this section).
    (v) Lighting shall be measured quarterly, or once at the beginning, 
middle, and end of the study for shorter studies.
    (vi) Noise level in the exposure chamber(s) shall be measured 
quarterly, or once at the beginning, middle, and end of the study for 
shorter studies.
    (vii) Oxygen content is critical, especially in nose-only chamber 
systems, and shall be greater than or equal to 19 percent in the test 
cages. An oxygen sensor shall be located at a single position in the 
test chamber and a lower alarm limit of 18 percent shall be used to 
activate an alarm system.
    (8) Safety procedures and requirements. In the case of potentially 
explosive test substance concentrations, care shall be taken to avoid 
generating explosive atmospheres.
    (i) It is mandatory that the upper explosive limit (UEL) and lower 
explosive limit (LEL) for the fuel and/or fuel additive(s) that are 
being tested be determined. These limits can be found in the material 
safety data sheets (MSDS) for each substance and in various reference 
texts. The air concentration of the fuel or additive-base fuel mixture 
in the generation system, dilution/delivery system, and the exposure 
chamber system shall be calculated to ensure that explosive limits are 
not present.
    (ii) Storage, handling, and use of fuels or fuel/additive mixtures 
shall follow guidelines given in 29 CFR 1910.106.
    (iii) Monitoring for carbon monoxide (CO) levels is mandatory for 
combustion systems. CO shall be continuously monitored in the immediate 
area of the engine/vehicle system and in the exposure chamber(s).
    (iv) Air samples shall be taken quarterly in the immediate area of 
the vapor generation system and the exposure chamber system, or once at 
the beginning, middle, and end of the study for shorter studies. These 
samples shall be analyzed by methods described in paragraph 
(d)(6)(ii)(B) of this section.
    (v) With the presence of fuels and/or fuel additives, all 
electrical and electronic equipment must be grounded. Also, the 
dilution/delivery system and chamber exposure system must be grounded. 
Guidelines for grounding are given in 29 CFR 1910.304.
    (9) Quality control and quality assurance procedures--(i) Standard 
operating procedures (SOPs). SOPs for exposure operations, sampling 
instruments, animal handling, and analytical methods shall be written 
during the developmental phase of the study.
    (ii) Technicians/operators shall be trained in exposure operation, 
maintenance, and documentation, as appropriate, and their training 
shall be documented.
    (iii) Flow meters, sampling instruments, and balances used in the 
inhalation experiments shall be calibrated with standards during the 
developmental phase to determine their sensitivity, detection limits, 
and linearity. During the exposure period, instruments shall be checked 
for calibration and documented to ensure that each instrument still 
functions properly.
    (iv) The mean exposure concentration shall be within 10 percent of 
the target concentration on 90 percent or more of exposure days. The 
coefficient of variation shall be within 25 percent of target on 90 
percent or more of exposure days. For example, a manufacturer might 
determine a mean exposure concentration of its product's exposure 
emissions by identifying ``marker'' compound(s) typical of the 
emissions of the fuel or fuel/additive mixture under study as a 
surrogate for the total of individual compounds in those exposure 
emissions. The manufacturer would note any concentration changes in the 
level of the ``marker'' compound(s) in the sample's daily emissions for 
biological testing.
    (v) The spatial variation of the chamber concentration shall be 10 
percent, or less. If a higher spatial variation is observed during the 
developmental phase, then air mixing in the chamber shall be increased. 
In any case, animals shall be rotated among the various cages in the 
exposure chamber(s) to insure each animal's uniform exposure during the 
study.
    (e) Data and reporting. Data shall be summarized in tabular form, 
showing for each group the number of animals at the start of the test, 
the number of animals showing lesions, the types of lesions, and the 
percentage of animals displaying each type of lesion.
    (1) Treatment of results. All observed results, quantitative and 
incidental, shall be evaluated by an appropriate statistical method. 
Any generally accepted statistical method may be used; the statistical 
methods shall be selected during the design of the study.
    (2) Evaluation of results. The findings of an inhalation toxicity 
study should be evaluated in conjunction with the findings of preceding 
studies and considered in terms of the observed toxic effects and the 
necropsy and histopathological findings. The evaluation will include 
the relationship between the concentration of the test atmosphere and 
the duration of exposure, and the severity of abnormalities, gross 
lesions, identified target organs, body weight changes, effects on 
mortality and any other general or specific toxic effects.
    (3) Test conditions. (i) The exposure apparatus shall be described, 
including:
    (A) The vehicle/engine design and type, the dynamometer, the 
cooling system, if any, the computer control system, and the dilution 
system for exhaust emission generation;
    (B) The evaporative emissions generator model, type, or design and 
its dilution system; and
    (C) Other test conditions, such as the source and quality of mixing 
air, fuel or fuel/additive mixture used, treatment of exhaust air, 
design of exposure chamber and the method of housing animals in a test 
chamber shall be described.
    (ii) The equipment for measuring temperature, humidity, particulate 
aerosol concentrations and size distribution, gas analyzers, fuel vapor 
concentrations, chamber distribution, and rise and fall time shall be 
described.
    (iii) Daily exposure results. The daily record shall document the 
date, the start and stop times of the exposure, number of samples taken 
during the day, daily concentrations determined, calibration of 
instruments, and problems encountered during the exposure. The daily 
exposure data shall be signed by the exposure operator and reviewed and 
signed by the exposure supervisor responsible for the study.
    (4) Exposure data shall be tabulated and presented with mean values 
and a measure of variability (e.g., standard deviation), and shall 
include:
    (i) Airflow rates through the inhalation equipment;
    (ii) Temperature and humidity of air;
    (iii) Chamber concentrations in the chamber breathing zone;
    (iv) Concentration of combustion exhaust gases in the chamber 
breathing zone;
    (v) Particle size distribution (e.g., mass median aerodynamic 
diameter and geometric standard deviation from the mean);
    (vi) Rise and fall time;
    (vii) Chamber concentrations during the non-exposure period; and
    (viii) Distribution of test substance in the chamber.
    (5) Animal data. Tabulation of toxic response data by species, 
strain, sex and exposure level for:
    (i) Number of animals exposed;
    (ii) Number of animals showing signs of toxicity; and
    (iii) Number of animals dying.
    (f) References. For additional background information on this 
exposure guideline, the following references should be consulted.

    (1) Barr, E.B. (1988) Operational Limits for Temperature and 
Percent Oxygen During HM Nose-Only Exposures--Emergency Procedures 
[interoffice memorandum]. Albuquerque, NM: Lovelace Inhalation 
Toxicology Research Institute; May 13.
    (2) Barr, E.B.; Cheng, Y.S.; Mauderly, J.L. (1990) Determination 
of Oxygen Depletion in a Nose-Only Exposure Chamber. Presented at: 
1990 American Association for Aerosol Research; June; Philadelphia, 
PA: American Association for Aerosol Research; abstract no. P2e1.
    (3) Barrow, C.S. (1989) Generation and Characterization of Gases 
and Vapors. In: McClellan, R.O., Henderson, R.F. ed. Concepts in 
Inhalation Toxicology. New York, NY: Hemisphere Publishing Corp., 
63-84.
    (4) Benedict, R.P. (1984) Fundamentals of Temperature, Pressure, 
and Flow Measurements. 3rd ed. New York, NY: John Wiley and Sons.
    (5) Cannon, W.C.; Blanton, E.F.; McDonald, K.E. The Flow-Past 
Chamber. (1983) An Improved Nose-Only Exposure System for Rodents. 
Am. Ind. Hyg. Assoc. J. 44: 923-928.
    (6) Chaddock, J.B. ed. (1985) Moisture and humidity. Measurement 
and Control in Science and Industry: Proceedings of the 1985 
International Symposium on Moisture and Humidity; April 1985; 
Washington, D.C. Research Triangle Park, NC: Instrument Society of 
America.
    (7) Cheng, Y.S.; Barr, E.B.; Carpenter, R.L.; Benson, J.M.; 
Hobbs, C.H. (1989) Improvement of Aerosol Distribution in Whole-Body 
Inhalation Exposure Chambers. Inhal. Toxicol. 1: 153-166.
    (8) Cheng,Y.S.; Moss, O.R. (1989) Inhalation Exposure Systems. 
In: McClellan, R.O.; Henderson, R.F. ed. Concepts in Inhalation 
Toxicology. New York, NY: Hemisphere Publishing Corp., 19-62.
    (9) Cheng, Y.S.; Yeh, H.C.; Mauderly, J.L.; Mokler, B.V. (1984) 
Characterization of Diesel Exhaust in a Chronic Inhalation Study. 
Am. Ind. Hyg. Assoc. J. 45: 547-555.
    (10) Gillum, D.R. (1982) Industrial Pressure Measurement. 
Research Triangle Park, NC: Instrument Society of America.
    (11) Hinners, R.G.; Burkart, J.K.; Malanchuk, M. (1979) Animal 
Exposure Facility for Diesel Exhaust Studies.
    (12) Kittelson, D.B.; Dolan, D.F. (1979) Diesel exhaust 
aerosols. In Willeke, K. ed. Generation of Aerosols and Facilities 
for Exposure Experiments. Ann Arbor, MI: Ann Arbor Science 
Publishers Inc., 337-360.
    (13) Mokler, B.V.; Archibeque, F.A.; Beethe, R.L.; Kelly, 
C.P.J.; Lopez, J.A.; Mauderly, J.L.; Stafford, D.L. (1984) Diesel 
Exhaust Exposure System for Animal Studies. Fundamental and Applied 
Toxicology 4: 270-277.
    (14) Moore, W.; et al. (1978) Preliminary finding on the 
Deposition and Retention of Automotive Diesel Particulate in Rat 
Lungs. Proc. of Annual Meeting of the Air Pollution Control Assn, 3, 
paper 78-33.7.
    (15) Raabe, O.G., Bennick, J.E., Light, M.E., Hobbs, C.H., 
Thomas, R.L., Tillery, M.I. (1973) An Improved Apparatus for Acute 
Inhalation Exposure of Rodents to Radioactive Aerosols. Toxicol & 
Applied Pharmaco.; 1973; 26: 264-273.
    (16) Rao, G.N. (1986) Significance of Environmental Factors on 
the Test System. In: Hoover, B.K.; Baldwin, J.K.; Uelner, A.F.; 
Whitmire, C.E.; Davies, C.L.; Bristol, D.W. ed. Managing conduct and 
data quality of toxicology studies. Raleigh, NC: Princeton 
Scientific Publishing Co., Inc.: 173-185.
    (17) Spitzer, D.W. (1984) Industrial Flow Measurement. Research 
Triangle Park, NC: Instrument Society of America.
    (18) 40 CFR part 798, Health effects testing guidelines.
    (19) 29 CFR part 1910, Occupational safety and health standards 
for general industry.
    (20) Federal Register, 42 FR 26748, May 25, 1977.


Sec. 79.62  Subchronic toxicity study with specific health effect 
assessments.

    (a) Purpose--(1) General toxicity. This subchronic inhalation study 
is designed to determine a concentration-response relationship for 
potential toxic effects in rats resulting from continuous or repeated 
inhalation exposure to vehicle/engine emissions over a period of 90 
days. A subgroup of perfusion-fixed animals is required, in addition to 
the main study population, for more exacting organ and tissue 
histology. This test will provide screening information on target organ 
toxicities and on concentration levels useful for running chronic 
studies and establishing exposure criteria. Initial information on 
effective concentrations/exposures of the test atmosphere may be 
determined from the literature of previous studies or through 
concentration range-finding trials prior to starting this study. This 
health effects screening test is not capable of directly determining 
those effects which have a long latency period for development (e.g., 
carcinogenicity and life-shortening), though it may permit the 
detremination of a no-observed-adverse-effect level, or NOAEL.
    (2) Specific health effects assessments (HEAs). These supplemental 
studies are designed to determine the potential for reproductive/
teratologic, carcinogenic, mutagenic, and neurotoxic health effect 
outcomes from vehicle/engine emission exposures. They are done in 
combination with the subchronic toxicity study and paragraph (c) of 
this section or may be done separately as outlined by the appropriate 
test guideline.
    (i) Fertility assessment/teratology. The fertility assessment is an 
in vivo study designed to provide information on potential health 
hazards to the fetus arising from the mother's repeated exposure to 
vehicle/engine emissions before and during her pregnancy. By including 
a mating of test animals, the study provides preliminary data on the 
effects of repeated vehicle/engine emissions exposure on gonadal 
function, conception, and fertility. The fertility assessment/
teratology guideline is found in Sec. 79.63.
    (ii) Micronucleus (MN) Assay. The MN assay is an in vivo 
cytogenetic test which gives information on potential carcinogenic and/
or mutagenic effects of exposure to vehicle/engine emissions. The MN 
assay detects damage to the chromosomes or mitotic apparatus of cells 
in the tissues of a test subject exposed repeatedly to vehicle/engine 
emissions. The assay is based on an increase in the frequency of 
micronucleated erythrocytes found in bone marrow from treated animals 
compared to that of control animals. The guideline for the MN assay is 
found in Sec. 79.64.
    (iii) Sister Chromatid Exchange (SCE) Assay. The SCE assay is an in 
vivo analysis which gives information on potential mutagenic and/or 
carcinogenic effects of exposure to vehicle/engine emissions. The assay 
detects the ability of a chemical to enhance the exchange of DNA 
between two sister chromatids of a duplicating chromosome. This assay 
uses peripheral blood lymphocytes isolated from an exposed rodent test 
species and grown to confluence in cell culture. The guideline for the 
SCE assay is found in Sec. 79.65.
    (iv) Neurotoxicity (NTX) measures. NTX measures include (A) 
histopathology of specified central and peripheral nervous system 
tissues taken from emission-exposed rodents, and (B) an assay of brain 
tissue levels of glial fibrillary acidic protein (GFAP), a major 
filament protein of astrocytes, from emission-exposed rodents. The 
guidelines for the neurohistopathology and GFAP studies are found in 
Sec. 79.66 and Sec. 79.67, respectively.
    (b) Definitions. For the purposes of this section, the following 
definitions apply:
    No-observed-adverse-effect-level (NOAEL) means the maximum 
concentration used in a test which produces no observed adverse 
effects. A NOAEL is expressed in terms of weight or volume of test 
substance given daily per unit volume of air (g/L or ppm).
    Subchronic inhalation toxicity means the adverse effects occurring 
as a result of the continuous or repeated daily exposure of 
experimental animals to a chemical by inhalation for part 
(approximately 10 percent) of a life span.
    (c) Principle of the test method. As long as none of the 
requirements of any study are violated by the combination, one or more 
HEAs may be combined with the general toxicity study through concurrent 
exposures of their study populations and/or by sharing the analysis of 
the same animal subjects. Requirements duplicated in combined studies 
need not be repeated. Guidelines for combining HEAs with the general 
toxicity study are as follows.
    (1) Fertility assessment. (i) The number of study animals in the 
test population is increased when the fertility assessment is run 
concurrently with the 90-day toxicity study. A minimum of 40 females 
per test group shall undergo vaginal lavage daily for two weeks before 
the start of the exposure period. The resulting wet smears are examined 
to cull those animals which are acyclic. Twenty-five females shall be 
randomly assigned to a for-breeding group with the balance of females 
assigned to a group for histopathologic examination.
    (ii) All test groups are exposed over a period of 90 days to 
various concentrations of the test atmosphere for a minimum of six 
hours per day. After seven weeks of exposures, analysis of vaginal cell 
smears shall resume on a daily basis for the 25 for-breeding females 
and shall continue for a period of four weeks or until each female in 
the group is confirmed pregnant. Following the ninth week of exposures, 
each for-breeding female is housed overnight with a single study male. 
Matings shall continue for as long as two weeks, or until pregnancy is 
confirmed (pregnancy day 0). Pregnant females are only exposed through 
day 15 of their pregnancy while daily exposures continue throughout the 
course of the study for non-pregnant females and study males.
    (iii) On pregnancy day 20, pregnant females are sacrificed and 
their uteri are examined. Pregnancy status and fetal effects are 
recorded as described in Sec. 79.63. At the end of the exposure period, 
all males and non-pregnant females are sacrificed and necropsied. 
Testes and epididymal tissue samples are taken from five perfusion-
fixed test subjects and histopathological examinations are carried out 
on the remainder of the non-pregnant females and study males.
    (2) Carcinogenicity/mutagenicity
(C/M) assessment. When combined with the subchronic toxicity study, the 
main study population is used to perform both the in vivo MN and SCE 
assays. Because of the constant turnover of the cells to be analyzed in 
these assays, a separate study population may be used for this 
assessment. A study population needs only to be exposed a minimum of 
four weeks. At exposure's end, ten animals per exposure and control 
groups are anaesthetized and heart punctures are performed on all 
members. After separating blood components, individual lymphocyte cell 
cultures are set up for SCE analysis. One femur from each study subject 
is also removed and the marrow extracted. The marrow is smeared onto a 
glass slide, and stained for analysis of micronuclei in erythrocytes.
    (3) Neurotoxicity (NTX) measures. (i) When combined with this 
subchronic toxicity study, test animals designated for whole-body 
perfusion fixation/lung histology and exposed as part of the main 
animal population are used to perform the neurohistology portion of 
these measures. After the last exposure period, a minimum of ten 
animals from each exposure group shall be preserved in situ with 
fixative. Sections of brain, spinal cord, and proximal sciatic or 
tibial nerve are then cut, processed further in formalin, and mounted 
for viewing under a light microscope. Fibers from the sciatic or tibial 
nerve sample are teased apart for further analysis under the 
microscope.
    (ii) GFAP assay. After the last exposure period, a minimum of ten 
rodents from each exposure group shall be sacrificed, and their brains 
excised and divided into regions. The tissue samples are then applied 
to filter paper, washed with anti-GFAP antibody, and visualized with a 
radio-labelled Protein A. The filters are quantified for degree of 
immunoreactivity between the antibody and GFAP in the tissue samples. A 
non-radioactive ELISA format is also referenced in the GFAP guideline 
cited in paragraph (a)(2)(iv) of this section. Note: Because the GFAP 
assay requires fresh, i.e., non-preserved, brain tissue, the number of 
test animals may need to be increased to provide an adequate number of 
test subjects to complete the histopathology requirements of both the 
GFAP and the general toxicity portion of the 90-day inhalation study.
    (iii) The start of the exposure period for the NTX measures study 
population may be staggered from that of the main study group to more 
evenly distribute the analytical work required in both study 
populations. The exposures would remain the same in all other respects.
    (d) Test procedures--(1) Animal selection--(i) Species and sex. The 
rat is the recommended species. If another rodent species is used, the 
tester shall provide justification for its selection. Both sexes shall 
be used in any assessment unless it is demonstrated that one sex is 
refractory to the effects of exposure.
    (ii) Age and number. Rats shall be at least ten weeks of age at the 
beginning of the study exposure. The number of animals necessary for 
individual health effect outcomes is as follows:
    (A) Thirty rodents per concentration level/group, fifteen of each 
sex, shall be used to satisfy the reporting requirements of the 90-day 
toxicity study. Ten animals per concentration level/group shall be 
designated for whole body perfusion with fixative (by gravity) for lung 
studies, and neurohistology and testes studies, as appropriate.
    (B) Forty rodents, 25 females and ten males shall be added for each 
test concentration or control group when combining a 90-day toxicity 
study with a fertility assessment.
    (C) The tester shall provide a group of 10 animals (five animals 
per sex per experimental/control groups) in addition to the main test 
population when performing the GFAP neurotoxicity HEA.
    (2) Recovery group. The manufacturer shall include an group of 20 
animals (10 animals per sex) in the test population, exposing them to 
the highest concentration level for the entire length of the study's 
exposure period. This group shall then be observed for reversibility, 
persistence, or delayed occurrence of toxic effects during a post-
exposure period of not less than 28 days.
    (3) Inhalation exposure. (i) All data developed within this study 
shall be in accordance with good laboratory practice provisions under 
Sec. 79.60.
    (ii) The general conduct of this study shall be in accordance with 
the vehicle emissions inhalation exposure guideline in Sec. 79.61.
    (4) Observation of animals. (i) All toxicological (e.g., weight 
loss) and neurological signs (e.g., motor disturbance) shall be 
recorded frequently enough to observe any abnormality, and not less 
than weekly for all study animals. Animals shall be weighed weekly.
    (ii) The following is a minimal list of measures that shall be 
noted:
    (A) Body weight;
    (B) Subject's reactivity to general stimuli such as removal from 
the cage or handling;
    (C) Description, incidence, and severity of any convulsions, 
tremors, or abnormal motor movements in the home cage;
    (D) Descriptions and incidence of posture and gait abnormalities 
observed in the home cage;
    (E) Description and incidence of any unusual or abnormal behaviors, 
excessive or repetitive actions (stereotypies), emaciation, 
dehydration, hypotonia or hypertonia, altered fur appearance, red or 
crusty deposits around the eyes, nose, or mouth, and any other 
observations that may facilitate interpretation of the data.
    (iii) Any animal which dies during the test is necropsied as soon 
as possible after discovery.
    (5) Clinical examinations. (i) The following examinations shall be 
performed on the twenty animals designated as the 90-day study 
population, exclusive of pregnant dams and those study animals targeted 
for perfusion by gravity:
    (A) The following hematology determinations shall be carried out at 
least two times during the test period (after 30 days of exposure and 
just prior to terminal sacrifice at the end of the exposure period): 
hematocrit, hemoglobin concentration, erythrocyte count, total and 
differential leukocyte count, and a measure of clotting potential such 
as prothrombin time, thromboplastin time, or platelet count.
    (B) Clinical biochemistry determinations on blood shall be carried 
out at least two times during the test period, after 30 days of 
exposure and just prior to terminal sacrifice at the end of the 
exposure period, on all groups of animals including concurrent 
controls. Clinical biochemical testing shall include assessment of 
electrolyte balance, carbohydrate metabolism, and liver and kidney 
function. The selection of specific tests will be influenced by 
observations on the mode of action of the substance. In the absence of 
more specific tests, the following determinations may be made: calcium, 
phosphorus, chloride, sodium, potassium, fasting glucose (with period 
of fasting appropriate to the species), serum alanine aminotransferase, 
serum aspartate aminotransferase, sorbitol dehydrogenase, gamma 
glutamyl transpeptidase, urea nitrogen, albumen, blood creatinine, 
methemoglobin, bile acids, total bilirubin, and total serum protein 
measurements. Additional clinical biochemistry shall be employed, where 
necessary, to extend the investigation of observed effects, e.g., 
analyses of lipids, hormones, acid/base balance, and cholinesterase 
activity.
    (ii) The following examinations shall initially be performed on the 
high concentration and control groups only:
    (A) Ophthalmological examination, using an ophthalmoscope or 
equivalent suitable equipment, shall be made prior to exposure to the 
test substance and at the termination of the study. If changes in the 
eyes are detected, all animals shall be examined.
    (B) Urinalysis is not required on a routine basis, but shall be 
done when there is an indication based on expected and/or observed 
toxicity.
    (iii) Preservation by whole-body perfusion of fixative into the 
anaesthetized animal for lung histology of ten animals from the 90-day 
study population for each experimental and control group.
    (6) Gross pathology. With the exception of the whole body 
perfusion-fixed test animals cited in paragraph (d)(1)(ii)(A) of this 
section, all rodents shall be subjected to a full gross necropsy which 
includes examination of the external surface of the body, all orifices 
and the cranial, thoracic, and abdominal cavities and their contents. 
Gross pathology shall be performed on the following organs and tissues:
    (i) The liver, kidneys, lungs, adrenals, brain, and gonads, 
including uterus, ovaries, testes, epididymides, seminal vesicles (with 
coagulating glands), and prostate, constitute the group of target 
organs for histology and shall be weighed as soon as possible after 
dissection to avoid drying. In addition, for other than rodent test 
species, the thyroid with parathyroids, when present, shall also be 
weighed as soon as possible after dissection to avoid drying.
    (ii) The following organs and tissues, or representative samples 
thereof, shall be preserved in a suitable medium for possible future 
histopathological examination: All gross lesions; lungs--which shall be 
removed intact, weighed, and treated with a suitable fixative to ensure 
that lung structure is maintained (perfusion with the fixative is 
considered to be an effective procedure); nasopharyngeal tissues; 
brain--including sections of medulla/pons, cerebellar cortex, and 
cerebral cortex; pituitary; thyroid/parathyroid; thymus; trachea; 
heart; sternum with bone marrow; salivary glands; liver; spleen; 
kidneys; adrenals; pancreas; reproductive organs: uterus; cervix; 
ovaries; vagina; testes; epididymides; prostate; and, if present, 
seminal vesicles; aorta; (skin); gall bladder (if present); esophagus; 
stomach; duodenum; jejunum; ileum; cecum; colon; rectum; urinary 
bladder; representative lymph node; (mammary gland); (thigh 
musculature); peripheral nerve/tissue; (eyes); (femur--including 
articular surface); (spinal cord at three levels--cervical, 
midthoracic, and lumbar); and (zymbal and exorbital lachrymal glands).
    (7) Histopathology. Histopathology shall be performed on the 
following organs and tissues from all rodents:
    (i) All gross lesions.
    (ii) Respiratory tract and other organs and tissues, listed in 
paragraph (d)(6)(ii) of this section (except organs/tissues in 
parentheses), of all animals in the control and high dose groups.
    (iii) The tissues mentioned in parentheses, listed in paragraph 
(d)(6)(ii) of this section, if indicated by signs of toxicity or target 
organ involvement.
    (iv) Lungs of animals in the low and intermediate dose groups shall 
also be subjected to histopathological examination, primarily for 
evidence of infection since this provides a convenient assessment of 
the state of health of the animals.
    (v) Lungs and trachea of the whole-body perfusion-fixed test 
animals cited in paragraph (d)(1)(ii)(A) of this section are examined 
for inhaled particle distribution.
    (e) Interpretation of results. All observed results, quantitative 
and incidental, shall be evaluated by an appropriate statistical 
method. The specific methods, including consideration of statistical 
power, shall be selected during the design of the study.
    (f) Test report. In addition to the reporting requirements as 
specified under Secs. 79.60 and 79.61(e), the following individual 
animal data information shall be reported:
    (1) Date of death during the study or whether animals survived to 
termination.
    (2) Date of observation of each abnormal sign and its subsequent 
course.
    (3) Individual body weight data, and group average body weight data 
vs. time.
    (4) Feed consumption data, when collected.
    (5) Hematological tests employed and all results.
    (6) Clinical biochemistry tests employed and all results.
    (7) Necropsy findings.
    (8) Type of stain/fixative and procedures used in preparing tissue 
samples.
    (9) Detailed description of all histopathological findings.
    (10) Statistical treatment of the study results, where appropriate.
    (g) References. For additional background information on this test 
guideline, the following references should be consulted.
    (1) 40 CFR 798.2450, Inhalation toxicity.
    (2) 40 CFR 798.2675, Oral Toxicity with Satellite Reproduction and 
Fertility Study.
    (3) General Statement of Work for the Conduct of Toxicity and 
Carcinogenicity Studies in Laboratory Animals (revised April, 1987/
modifications through January, 1990) appendix G, National Toxicology 
Program--U.S. Dept. of Health and Human Services (Public Health 
Service), P.O. Box 12233, Research Triangle Park, NC 27709.


Sec. 79.63  Fertility assessment/teratology.

    (a) Purpose. Fertility assessment/teratology is an in vivo study 
designed to provide information on potential health hazards to the 
fetus arising from the mother's repeated inhalation exposure to 
vehicle/engine emissions before and during her pregnancy. By including 
a mating of test animals, the study provides preliminary data on the 
effects of repeated vehicle/engine emissions exposure on gonadal 
function, conception, and fertility. Since this is a one-generation 
test that ends with examination of full-term fetuses, but not of live 
pups, it is not capable of determining effects on reproductive 
development which would only be detected in viable offspring of treated 
parents.
    (b) Definitions. For the purposes of this section, the following 
definitions apply:
    Developmental toxicity means the ability of an agent to induce in 
utero death, structural or functional abnormalities, or growth 
retardation after contact with the pregnant animal.
    Estrous cycle means the periodic recurrence of the biological 
phases of the female reproductive system which prepare the animal for 
conception and the development of offspring. The phases of the estrous 
cycle for a particular animal can be characterized by the general 
condition of the cells present in the vagina and the presence or 
absence of various cell types.
    Vaginal cytology evaluation means the use of wet vaginal cell 
smears to determine the phase of a test animal's estrous cycle and the 
potential for adverse exposure effects on the regularity of the 
animal's cycle. In the rat, common cell types found in the smears 
correlate well with the various stages of the estrous cycle and to 
changes occurring in the reproductive tract.
    (c) Principle of the test method. (1) For a two week period before 
exposures start, daily vaginal cell smears are examined from a surplus 
of female test animals to identify and cull those females which are 
acyclic. After culling, testers shall randomly assign at each exposure 
concentration (including unexposed) a minimum of twenty-five females 
for breeding and fifteen non-bred females for later histologic 
evaluation. Test animals shall be exposed by inhalation to graduated 
concentrations of the test atmosphere for a minimum of six hours per 
day over the next 13 weeks. Males and females in both test and control 
groups are mated after nine weeks of exposure. Exposures for pregnant 
females continue through gestation day 15, while exposures for males 
and all non- pregnant females shall continue for the full exposure 
period.
    (2) Beginning two weeks before the start of the mating period, 
daily vaginal smears resume for all to-be-bred females to characterize 
their estrous cycles. This will continue for four weeks or until a 
rat's pregnancy is confirmed, i.e., day 0, by the presence of sperm in 
the cell smear. On pregnancy day 20, shortly before the expected date 
of delivery, each pregnant female is sacrificed, her uterus removed, 
and the contents examined for embryonic or fetal deaths, and live 
fetuses. At the end of the exposure period, males and all non-pregnant 
females shall be weighed, and various organs and tissues, as 
appropriate, shall be removed and weighed, fixed with stain, and 
sectioned for viewing under a light microscope.
    (3) This assay may be done separately or in combination with the 
subchronic toxicity study, pursuant to the provisions in Sec. 79.62.
    (d) Limit test. If a test at one dose level of the highest 
concentration that can be achieved while maintaining a particle size 
distribution with a mass median aerodynamic diameter (MMAD) of 4 
micrometers (m) or less, using the procedures described in 
section 79.60 of this part produces no observable toxic effects and if 
toxicity would not be expected based upon data of structurally related 
compounds, then a full study using three dose levels might not be 
necessary. Expected human exposure though may indicate the need for a 
higher dose level.
    (e) Test procedures--(1) Animal selection--(i) Species and strain. 
The rat is the preferred species. Strains with low fecundity shall not 
be used and the candidate species shall be characterized for its 
sensitivity to developmental toxins. If another rodent species is used, 
the tester shall provide justification for its selection.
    (ii) Animals shall be a minimum of 10 weeks old at the start of the 
exposure period.
    (iii) Number and sex. Each test and control group shall have a 
minimum of 25 males and 40 females. In order to ensure that sufficient 
pups are produced to permit meaningful evaluation of the potential 
developmental toxicity of the test substance, twenty pregnant test 
animals are required for each exposure and control level.
    (2) Observation period. The observation period shall be 13 weeks, 
at a minimum.
    (3) Concentration levels and concentration selection. (i) To select 
the appropriate concentration levels, a pilot or trial study may be 
advisable. Since pregnant animals have an increased minute ventilation 
as compared to non-pregnant animals, it is recommended that the trial 
study be conducted in pregnant animals. Similarly, since presumably the 
minute ventilation will vary with progression of pregnancy, the animals 
should be exposed during the same period of gestation as in the main 
study. It is not always necessary, though, to carry out a trial study 
in pregnant animals. Comparisons between the results of a trial study 
in non-pregnant animals, and the main study in pregnant animals will 
demonstrate whether or not the test substance is more toxic in pregnant 
animals. In the trial study, the concentration producing embryonic or 
fetal lethalities or maternal toxicity should be determined.
    (ii) The highest concentration level shall induce some overt 
maternal toxicity such as reduced body weight or body weight gain, but 
not more than 10 percent maternal deaths.
    (iii) The lowest concentration level shall not produce any grossly 
observable evidence of either maternal or developmental toxicity.
    (4) Inhalation exposure. (i) All data developed within this study 
shall be in accordance with good laboratory practice provisions under 
Sec. 79.60.
    (ii) The general conduct of this study shall be in accordance with 
the vehicle emissions inhalation exposure guideline in Sec. 79.61.
    (f) Test performance--(1) Study conduct. Directions specific to 
this study are:
    (i) The duration of exposure shall be at least six hours daily, 
allowing appropriate additional time for chamber equilibrium.
    (ii) Where an exposure chamber is used, its design shall minimize 
crowding of the test animals. This is best accomplished by individual 
caging.
    (iii) Pregnant animals shall not be subjected to beyond the minimum 
amount of stress. Since whole-body exposure appears to be the least 
stressful mode of exposure, it is the preferred method. In general 
oronasal or head-only exposure, which is sometimes used to avoid 
concurrent exposure by the dermal or oral routes, is not recommended 
because of the associated stress accompanying the restraining of the 
animals. However, there may be specific instances where it may be more 
appropriate than whole-body exposure. The tester shall provide 
justification/reasoning for its selection.
    (iv) Measurements shall be made at least every other day of food 
consumption for all animals in the study. Males and females shall be 
weighed on the first day of exposure and 2-3 times per week thereafter, 
except for pregnant dams.
    (v) The test animal housing, mating, and exposure chambers shall be 
operated on a twenty-four hour lighting schedule, with twelve hours of 
light and twelve hours of darkness. Test animal exposure shall only 
occur during the light portion of the cycle.
    (vi) Signs of toxicity shall be recorded as they are observed 
including the time of onset, degree, and duration.
    (vii) Females showing signs of abortion or premature delivery shall 
be sacrificed and subjected to a thorough macroscopic examination.
    (viii) Animals that die or are euthanized because of morbidity will 
be necropsied promptly.
    (2) Vaginal cytology. (i) For a two week period before the mating 
period starts, each female in the to-be-bred population shall undergo a 
daily saline vaginal lavage. Two wet cell smears from this lavage shall 
be examined daily for each subject to determine a baseline pattern of 
estrus. Testers shall avoid excessive handling and roughness in 
obtaining the vaginal cell samples, as this may induce a condition of 
pseudo-pregnancy in the test animals.
    (ii) This will continue for four weeks or until day 0 of a rat's 
pregnancy is confirmed by the presence of sperm in the cell smear.
    (3) Mating and fertility assessment. (i) Beginning nine weeks after 
the start of exposure, each exposed and control group female (exclusive 
of the histology group females) shall be paired during non-exposure 
hours with a male from the same exposure concentration group. Matings 
shall continue for a period of two weeks, or until all mated females 
are determined to be pregnant. Mating pairs shall be clearly 
identified.
    (ii) Each morning, including weekends, cages shall be examined for 
the presence of a sperm plug. When found, this shall mark gestation day 
0 and pregnancy shall be confirmed by the presence of sperm in the 
day's wet vaginal cell smears.
    (iii) Two weeks after mating is begun, or as females are determined 
to be pregnant, bred animals are returned to pre-mating housing. Daily 
exposures continues through gestation day 15 for all pregnant females 
or through the balance of the exposure period for non-pregnant females 
and all males.
    (iv) Those pairs which fail to mate shall be evaluated in the 
course of the study to determine the cause of the apparent infertility. 
This may involve such procedures as additional opportunities to mate 
with a proven fertile partner, histological examination of the 
reproductive organs, and, in males, examination of the spermatogenic 
cycles. The stage of estrus for each non-pregnant female in the 
breeding group will be determined at the end of the exposure period.
    (4) All animals in the histology group shall be subject to 
histopathologic examination at the end of the study's exposure period.
    (g) Treatment of results. (1) All observed results, quantitative 
and incidental, shall be evaluated by an appropriate statistical 
method. The specific methods, including consideration of statistical 
power, shall be selected during the design of the study.
    (2) Data and reporting. In addition to the reporting requirements 
specified under Secs. 79.60 and 79.61, the final test report must 
include the following information:
    (i) Gross necropsy. (A) All animals shall be subjected to a full 
necropsy which includes examination of the external surface of the 
body, all orifices, and the cranial, thoracic, and abdominal cavities 
and their contents. Special attention shall be directed to the organs 
of the reproductive system.
    (B) The liver, kidneys, adrenals, pituitary, uterus, vagina, 
ovaries, testes, epididymides and seminal vesicles (with coagulating 
glands), and prostate shall be weighed wet, as soon as possible after 
dissection, to avoid drying.
    (i) At the time of sacrifice on gestation day 20 or at death during 
the study, each dam shall be examined macroscopically for any 
structural abnormalities or pathological changes which may have 
influenced the pregnancy.
    (ii) The contents of the uterus shall be examined for embryonic or 
fetal deaths and the number of viable fetuses. Gravid uterine weights 
need not be obtained from dead animals where decomposition has 
occurred. The degree of resorption shall be described in order to help 
estimate the relative time of death.
    (iii) The number of corpora lutea shall be determined in each 
pregnant dam.
    (iv) Each fetus shall be weighed, all weights recorded, and mean 
fetal weights determined.
    (v) Each fetus shall be examined externally and the sex determined.
    (vi) One-half of the rat fetuses in each litter shall be examined 
for skeletal anomalies, and the remaining half shall be examined for 
soft tissue anomalies, using appropriate methods.
    (ii) Histopathology. (A) Histopathology on vagina, uterus, ovaries, 
testes, epididymides, seminal vesicles, and prostate as appropriate for 
all males and histology group females in the control and high 
concentration groups and for all animals that died or were euthanized 
during the study. If abnormalities or equivocal results are seen in any 
of these organs/tissues, the same organ/tissue from test animals in 
lower concentration groups shall be examined.

    Note: Testes, seminal vesicles, epididymides, and ovaries, at a 
minimum, shall be examined in perfusion-fixed (pressure or gravity 
method) test subjects, when available.

    (B) All gross lesions in all study animals shall be examined.
    (C) As noted under mating procedures, reproductive organs of 
animals suspected of infertility shall be subject to microscopic 
examination.
    (D) The following organs and tissues, or representative samples 
thereof, shall be preserved in a suitable medium for future 
histopathological examination: all gross lesions; vagina; uterus; 
ovaries; testes; epididymides; seminal vesicles; prostate; liver; and 
kidneys/adrenals.
    (3) Evaluation of results. (i) The findings of a developmental 
toxicity study shall be evaluated in terms of the observed effects and 
the exposure levels producing effects. It is necessary to consider the 
historical developmental toxicity data on the species/strain tested.
    (ii) There are several criteria for determining a positive result 
for reproductive/teratologic effects; a statistically significant dose-
related decrease in the weight of the testes for treated subjects over 
control subjects, a decrease in neonatal viability, a significant 
change in the presence of soft tissue or skeletal abnormalities, or an 
increased rate of embryonic or fetal resorption or death. Other 
criteria, e.g., lengthening of the estrous cycle or the time spent in 
any one stage of estrus, changes in the proportion of viable male vs 
female fetuses or offspring, the number and type of cells in vaginal 
smears, or pathologic changes found during gross or microscopic 
examination of male or female reproductive organs may be based upon 
detection of a reproducible and statistically significant positive 
response for that evaluation parameter. A positive result indicates 
that, under the test conditions, the test substance does induce 
reproductive organ or fetal toxicity in the test species.
    (iii) A test substance which does not produce either a 
statistically significant dose-related change in the reproductive 
organs or cycle or a statistically significant and reproducible 
positive response at any one of the test points may not induce 
reproductive organ toxicity in this test species, but further 
investigation , e.g., to establish absorption and bioavailability of 
the test substance, should be considered.
    (h) Test report. In addition to the reporting requirements as 
specified under 40 CFR 79.60 and the vehicle emissions inhalation 
toxicity guideline as published in 40 CFR 79.61, the following specific 
information shall be reported:
    (1) Individual animal data. (i) Time of death during the study or 
whether animals survived to termination.
    (ii) Date of onset and duration of each abnormal sign and its 
subsequent course.
    (iii) Feed and body weight data.
    (iv) Necropsy findings.
    (v) Male test subjects.
    (A) Testicle weight, and body weight: testicle weight ratio.
    (B) Detailed description of all histopathological findings, 
especially for the testes and the epididymides.
    (vi) Female test subjects.
    (A) Uterine weight data.
    (B) Beginning and ending collection dates for vaginal cell smears.
    (C) Estrous cycle length compared within and between groups 
including mean cycle length for groups.
    (D) Percentage of time spent in each stage of cycle.
    (E) Stage of estrus at time of mating/sacrifice and proportion of 
females in estrus between concentration groups.
    (F) Detailed description of all histopathological findings, 
especially for uterine/ovary samples.
    (vii) Pregnancy and litter data. Toxic response data by exposure 
level, including but not limited to, indices of fertility and time-to-
mating, including the number of days until mating and the number of 
full or partial estrous cycles until mating.
    (A) Number of pregnant animals,
    (B) Number and percentage of live fetuses, resorptions.
    (viii) Fetal data. (A) Numbers of each sex.
    (B) Number of fetuses with any soft tissue or skeletal 
abnormalities.
    (2) Type of stain/fixative and procedures used in preparing tissue 
samples.
    (3) Statistical treatment of the study results.
    (i) References. For additional background information on this test 
guideline, the following references should be consulted.

    (1) 40 CFR 798.2675, Oral Toxicity with Satellite Reproduction 
and Fertility Study.
    (2) 40 CFR 798.4350, Inhalation Developmental Toxicity Study.
    (3) Chapin, R.E. and J.J. Heindel (1993) Methods in Toxicology, 
Vol. 3, Parts A and B: Reproductive Toxicology, Academic Press, 
Orlando, FL.
    (4) Gray, L.E., et al. (1989) ``A Dose-Response Analysis of 
Methoxychlor-Induced Alterations of Reproductive Development and 
Function in the Rat'' Fund. App. Tox. 12, 92-108.
    (5) Leblond, C.P. and Y. Clermont (1952) ``Definition of the 
Stages of the Cycle of the Seminiferous Epithelium of the Rat.'' 
Ann. N. Y. Acad. Sci. 55:548-73.
    (6) Morrissey, R.E., et al. (1988) ``Evaluation of Rodent Sperm, 
Vaginal Cytology, and Reproductive Organ Weight Data from National 
Toxicology Program 13-week Studies.'' Fundam. Appl. Toxicol. 11:343-
358.
    (7) Russell, L.D., Ettlin, R.A., Sinhattikim, A.P., and Clegg, 
E.D (1990) Histological and Histopathological Evaluation of the 
Testes, Cache River Press, Clearwater, FL.


Sec. 79.64   In vivo micronucleus assay.

    (a) Purpose. The micronucleus assay is an in vivo cytogenetic test 
which uses erythrocytes in the bone marrow of rodents to detect 
chemical damage to the chromosomes or mitotic apparatus of mammalian 
cells. As the erythroblast develops into an erythrocyte (red blood 
cell), its main nucleus is extruded and may leave a micronucleus in the 
cell body; a few micronuclei form under normal conditions in blood 
elements. This assay is based on an increase in the frequency of 
micronucleated erythrocytes found in bone marrow from treated animals 
compared to that of control animals. The visualization of micronuclei 
is facilitated in these cells because they lack a main nucleus.
    (b) Definitions. For the purposes of this section the following 
definitions apply:
    Micronuclei mean small particles consisting of acentric fragments 
of chromosomes or entire chromosomes, which lag behind at anaphase of 
cell division. After telophase, these fragments may not be included in 
the nuclei of daughter cells and form single or multiple micronuclei in 
the cytoplasm.
    Polychromatic erythrocyte (PCE) means an immature red blood cell 
that, because it contains RNA, can be differentiated by appropriate 
staining techniques from a normochromatic erythrocyte (NCE), which 
lacks RNA. In one to two days, a PCE matures into a NCE.
    (c) Test method--(1) Principle of the test method. (i) Groups of 
rodents are exposed by the inhalation route for a minimum of 6 hours/
day over a period of not less than 28 days to three or more 
concentrations of a test substance in air. Groups of animals are 
sacrificed at the end of the exposure period and femoral bone marrow is 
extracted. The bone marrow is then smeared onto glass slides, stained, 
and PCEs are scored for micronuclei. Researchers may need to run a 
trial at the highest tolerated concentration of the test atmosphere to 
optimize the sample collection time for micronucleated cells.
    (ii) This assay may be done separately or in combination with the 
subchronic toxicity study, pursuant to the provisions in Sec. 79.62.
    (2) Species and strain. (i) The rat is the recommended test animal. 
Other rodent species may be used in this assay, but use of that species 
will be justified by the tester.
    (ii) If a strain of mouse is used in this assay, the tester shall 
sample peripheral blood from an appropriate site on the test animal, 
e.g., the tail vein, as a source of normochromatic erythrocytes. 
Results shall be reported as outlined later in this guideline with 
``normochromatic'' interchanged for ``polychromatic'', where specified.
    (3) Animal number and sex. At least five female and five male 
animals per experimental/sample and control group shall be used. The 
use of a single sex or a smaller number of animals shall be justified.
    (4) Positive control group. A single concentration of a compound 
known to produce micronuclei in vivo is adequate as a positive control 
if it shows a significant response at any one time point; additional 
concentration levels may be used. To select an appropriate 
concentration level, a pilot or trial study may be advisable. 
Initially, one concentration of the test substance may be used, the 
maximum tolerated dose or that producing some indication of toxicity, 
e.g., a drop in the ratio of polychromatic to normochromatic 
erythrocytes. Intraperitoneal injection of 1,2-dimethyl-benz-anthracene 
or benzene are examples of positive control exposures. A concentration 
of 50-80 percent of an LD50 may be a suitable guide.
    (d) Test performance--(1) Inhalation exposure. (i) All data 
developed within this study shall be in accordance with good laboratory 
practice provisions under Sec. 79.60.
    (ii) The general conduct of this study shall be in accordance with 
the vehicle emissions inhalation exposure guideline in Sec. 79.61.
    (2) Preparation of slides and sampling times. Within twenty-four 
hours of the last exposure, test animals will be sacrificed. One femur 
from each test animal will be removed and placed in fetal bovine serum. 
The bone marrow is removed, cells processed, and two bone marrow smears 
are made for each animal on glass microscope slides. The slides are 
stained with acridine- orange (AO) or another appropriate stain (Giemsa 
+ Wright's, etc.) and examined under a microscope.
    (3) Analysis. Slides shall be coded for study before microscopic 
analysis. At least 1,000 first-division erythrocytes per animal shall 
be scored for the incidence of micronuclei. Sexes will be analyzed 
separately.
    (e) Data and report--(1) Treatment of results. In addition to the 
reporting requirements specified under Secs. 79.60 and 79.61, the final 
test report must include the criteria for scoring micronuclei. 
Individual data shall be presented in a tabular form including both 
positive and negative controls and experimental groups. The number of 
polychromatic erythrocytes scored, the number of micronucleated 
erythrocytes, the percentage of micronucleated cells, and, where 
applicable, the percentage of micronucleated erythrocytes shall be 
listed separately for each experimental and control animal. Absolute 
numbers shall be included if percentages are reported.
    (2) Interpretation of data. (i) There are several criteria for 
determining a positive response, one of which is a statistically 
significant dose-related increase in the number of micronucleated 
polychromatic erythrocytes. Another criterion may be based upon 
detection of a reproducible and statistically significant positive 
response for at least one of the test substance concentrations.
    (ii) A test substance which does not produce either a statistically 
significant dose-related increase in the number of micronucleated 
polychromatic erythrocytes or a statistically significant and 
reproducible positive response at any one of the test points is 
considered nonmutagenic in this system.
    (3) Test evaluation. (i) Positive results in the micronucleus test 
provide information on the ability of a chemical to induce micronuclei 
in erythrocytes of the test species under the conditions of the test. 
This damage may have been the result of chromosomal damage or damage to 
the mitotic apparatus.
    (ii) Negative results indicate that under the test conditions the 
test substance does not produce micronuclei in the bone marrow of the 
test species.
    (f) Test report. In addition to the reporting recommendations as 
specified under Sec. 79.60, the following specific information shall be 
reported:
    (1) Test atmosphere concentration(s) used and rationale for 
concentration selection.
    (2) Rationale for and description of treatment and sampling 
schedules, toxicity data, negative and positive controls.
    (3) Historical control data (negative and positive), if available.
    (4) Details of the protocol used for slide preparation.
    (5) Criteria for identifying micronucleated erythrocytes.
    (6) Micronucleus analysis by animal and by group for each 
concentration (sexes analyzed separately).
    (i) Ratio of polychromatic to normochromatic erythrocytes.
    (ii) Number of polychromatic erythrocytes with micronuclei.
    (iii) Number of polychromatic erythrocytes scored.
    (7) Statistical methodology chosen for test analysis.
    (g) References. For additional background information on this test 
guideline, the following references should be consulted.

    (1) 40 CFR 798.5395, In Vivo, Mammalian Bone Marrow Cytogenetics 
Tests: Micronucleus Assay.
    (2) Cihak, R. ``Evaluation of Benzidine by the Micronucleus 
Test.'' Mutation Research, 67: 383-384 (1979).
    (3) Evans, H.J. ``Cytological Methods for Detecting Chemical 
Mutagens.'' Chemical Mutagens: Principles and Methods for Their 
Detection, Vol. 4. Ed. A. Hollaender (New York and London: Plenum 
Press, 1976) pp. 1-29.
    (4) Heddle, J.A., et al. ``The Induction of Micronuclei as a 
Measure of Genotoxicity. A Report of the U.S. Environmental 
Protection Agency Gene-Tox Program.'' Mutation Research, 123:61-118 
(1983).
    (5) Preston, J.R. et al. ``Mammalian In Vivo and In Vitro 
Cytogenetics Assays: Report of the Gene-Tox Program.'' Mutation 
Research, 87:143-188 (1981).
    (6) Schmid, W. ``The micronucleus test for cytogenetic 
analysis'', Chemical Mutagens, Principles and Methods for their 
Detection. Vol. 4 Hollaender A, (Ed. A ed. (New York and London: 
Plenum Press, (1976) pp. 31-53.
    (7) Tice, R.E., and Al Pellom ``User's guide: Micronucleus assay 
data management and analysis system'', NTIS Order no. PB-90-212-
598AS.


Sec. 79.65  In vivo sister chromatid exchange assay.

    (a) Purpose. The in vivo sister chromatid exchange (SCE) assay 
detects the ability of a chemical to enhance the exchange of DNA 
between two sister chromatids of a duplicating chromosome. The most 
commonly used assays employ mammalian bone marrow cells or peripheral 
blood lymphocytes, often from rodent species.
    (b) Definitions. For the purposes of this section, the following 
definitions apply:
    C-metaphase means a state of arrested cell growth typically seen 
after treatment with a spindle inhibitor, i.e., colchicine.
    Sister chromatid exchange means a reciprocal interchange of the two 
chromatid arms within a single chromosome. This exchange is visualized 
during the metaphase portion of the cell cycle and presumably requires 
the enzymatic incision, translocation and ligation of at least two DNA 
helices.
    (c) Test method--(1) Principle of the test method. (i) Groups of 
rodents are exposed by the inhalation route for a minimum of 6 hours/
day over a period of not less than 28 days to three or more 
concentrations of a test substance in air. Groups of animals are 
sacrificed at the end of the exposure period and blood lymphocyte cell 
cultures are prepared from study animals. Cell growth is suspended 
after a time and cells are harvested, fixed and stained before scoring 
for SCEs. Researchers may need to run a trial at the highest tolerated 
concentration of the test atmosphere to optimize the sample collection 
time for second division metaphase cells.
    (ii) This assay may be done separately or in combination with the 
subchronic toxicity study, pursuant to the provisions in Sec. 79.62.
    (2) Description. (i) The method described here employs peripheral 
blood lymphocytes (PBL) of laboratory rodents exposed to the test 
atmosphere.
    (ii) Within twenty-four hours of the last exposure, test animal 
lymphocytes are obtained by heart puncture and duplicate cell cultures 
are started for each animal. Cultures are grown in bromo-deoxyuridine 
(BrdU), and then a spindle inhibitor (e.g., colchicine) is added to 
arrest cell growth. Cells are harvested, fixed, and stained and their 
chromosomes are scored for SCEs.
    (3) Species and strain. The rat is the recommended test animal. 
Other rodent species may be used in this assay, but use of that species 
will be justified by the tester.
    (4) Animal number and sex. At least five female and five male 
animals per experimental and control group shall be used. The use of a 
single sex or different number of animals shall be justified.
    (5) Positive control group. A single concentration of a compound 
known to produce SCEs in vivo is adequate as a positive control if it 
shows a significant response at any one time point; additional 
concentration levels may be used. To select an appropriate 
concentration level, a pilot or trial study may be advisable. 
Initially, one concentration of the test substance may be used, the 
maximum tolerated dose or that producing some indication of toxicity as 
evidenced by animal morbidity (including death) or target cell 
toxicity. Intraperitoneal injection of 1,2-dimethyl-benz-anthracene or 
benzene are examples of positive control exposures. A concentration of 
50-80 percent of an LD50 would also be a suitable guide.
    (6) Inhalation exposure. (i) All data developed within this study 
shall be in accordance with good laboratory practice provisions under 
Sec. 79.60.
    (ii) The general conduct of this study shall be in accordance with 
the vehicle emissions inhalation exposure guideline in Sec. 79.61.
    (d) Test performance--(1) Treatment. At the conclusion of the 
exposure period, all test animals are anaesthetized and heart punctures 
are performed. Lymphocytes are isolated over a Ficoll gradient and 
replicate cell cultures are started for each animal. After some 21 
hours, the cells are treated with BrdU and returned to incubation. The 
following day, a spindle inhibitor (e.g., colchicine) is added to 
arrest cell growth in c-metaphase. Cells are harvested 4 hours later 
and second-division metaphase cells are washed and fixed in 
methanol:acetic acid, stained, and chromosome preparations are scored 
for SCEs.
    (2) Staining method. Staining of slides to reveal SCEs can be 
performed according to any of several protocols. However, the 
fluorescence plus Giemsa method is recommended.
    (3) Number of cells scored. (i) A minimum of 25 well-stained, 
second-division metaphase cells shall be scored for each animal for 
each cell type.
    (ii) At least 100 consecutive metaphase cells shall be scored for 
the number of first, second, and third division metaphases for each 
animal for each cell type.
    (iii) At least 1000 consecutive PBL's shall be scored for the 
number of metaphase cells present.
    (iv) The number of cells to be analyzed per animal shall be based 
upon the number of animals used, the negative control frequency, the 
pre-determined sensitivity and the power chosen for the test. Slides 
shall be coded before microscopic analysis.
    (e) Data and report--(1) Treatment of results. In addition to the 
reporting requirements specified under Secs. 79.60 and 61, data shall 
be presented in tabular form, providing scores for both the number of 
SCE for each metaphase. Differences among animals within each group 
shall be considered before making comparisons between treated and 
control groups.
    (2) Statistical evaluation. Data shall be evaluated by appropriate 
statistical methods.
     (3) Interpretation of results. (i) There are several criteria for 
determining a positive result, one of which is a statistically 
significant dose-related increase in the number of SCE. Another 
criterion may be based upon detection of a reproducible and 
statistically significant positive response for at least one of the 
test concentrations.
    (ii) A test substance which does not produce either a statistically 
significant dose-related increase in the number of SCE or a 
statistically significant and reproducible positive response at any one 
of the test concentrations is considered not to induce rearrangements 
of DNA segments in this system.
    (iii) Both biological and statistical significance shall be 
considered together in the evaluation.
    (4) Test evaluation. (i) A positive result in the in vivo SCE assay 
for either, or both, the lung or lymphocyte cultures indicates that 
under the test conditions the test substance induces reciprocal 
interchanges of DNA in duplicating chromosomes from lung or lymphocyte 
cells of the test species.
    (ii) Negative results indicate that under the test conditions the 
test substance does not induce reciprocal interchanges in lung or 
lymphocyte cells of the test species.
    (5) Test report. In addition to the reporting recommendations as 
specified under Secs. 79.60 and 79.61, the following specific 
information shall be reported:
    (i) Test concentrations used, rationale for concentration 
selection, negative and positive controls;
    (ii) Toxic response data by concentration;
    (iii) Schedule of administration of test atmosphere, BrdU, and 
spindle inhibitor;
    (iv) Time of harvest after administration of BrdU;
    (v) Identity of spindle inhibitor, its concentration and timing of 
treatment;
    (vi) Details of the protocol used for cell culture and slide 
preparation;
    (vii) Criteria for scoring SCE;
    (viii) Replicative index, i.e., [percent 1st division+(2 x percent 
2nd division) + (3 x percent 3rd division) metaphases]/100; and
    (ix) Mitotic activity, i.e., # of metaphases/1000 cells.
    (f) References. For additional background information on this test 
guideline, the following references should be consulted.

    (1) 40 CFR 798.5915, In vivo Sister Chromatid Exchange Assay.
    (2) Kato, H. ``Spontaneous Sister Chromatid Exchanges Detected 
by a BudR-Labeling Method.'' Nature, 251:70-72 (1974).
    (4) Kligerman, A. D., et al. ``Sister Chromatid Exchange 
Analysis in Lung and Peripheral Blood Lymphocytes of Mice Exposed to 
Methyl Isocyanate by Inhalation.'' Environmental Mutagenesis 9:29-36 
(1987).
    (5) Kligerman, A.D., et al., ``Cytogenetic Studies of Rodents 
Exposed to Styrene by Inhalation'', IARC Monographs no. 127 
``Butadiene and Styrene: Assesment of Health Hazards'' (Sorsa, et 
al., eds), pp 217-224, 1993.
    (6) Kligerman, A., et al., ``Cytogenetic Studies of Mice Exposed 
to Styrene by Inhalation.'', Mutation Research, 280:35-43, 1992.
    (7) Wolff, S., and P. Perry. ``Differential Giemsa Staining of 
Sister Chromatids and the Study of Sister Chromatid Exchanges 
Without Autoradiography.'' Chromosoma 48: 341-53 (1974).


Sec. 79.66  Neuropathology assessment.

    (a) Purpose. (1) The histopathological and biochemical techniques 
in this guideline are designed to develop data in animals on 
morphologic changes in the nervous system associated with repeated 
inhalation exposures to motor vehicle emissions. These tests are not 
intended to provide a detailed evaluation of neurotoxicity. 
Neuropathological evaluation should be complemented by other 
neurotoxicity studies, e.g. behavioral and neurophysiological studies 
and/or general toxicity testing, to more completely assess the 
neurotoxic potential of an exposure.
    (2) [Reserved]
    (b) Definition. Neurotoxicity (NTX) or a neurotoxic effect is an 
adverse change in the structure or function of the nervous system 
following exposure to a chemical substance.
    (c) Principle of the test method. (1) Laboratory rodents are 
exposed to one of several concentration levels of a test atmosphere for 
at least six hours daily over a period of 90 days. At the end of the 
exposure period, the animals are anaesthetized, perfused in situ with 
fixative, and tissues in the nervous system are examined grossly and 
prepared for microscopic examination. Starting with the highest dosage 
level, tissues are examined under the light microscope for morphologic 
changes, until a no-observed-adverse-effect level is determined. In 
cases where light microscopy has revealed neuropathology, the NOAEL may 
be confirmed by electron microscopy.
    (2) The tests described herein may be combined with any other 
toxicity study, as long as none of the requirements of either are 
violated by the combination. Specifically, this assay may be combined 
with a subchronic toxicity study, pursuant to provisions in Sec. 79.62.
    (d) Limit test. If a test at one dose level of the highest 
concentration that can be achieved while maintaining a particle size 
distribution with a mass median aerodynamic diameter (MMAD) of 4 
micrometers (m) or less, using the procedures described in 
paragraph (a) of this section, produces no observable toxic effects and 
if toxicity would not be expected based upon data of structurally 
related compounds, then a full study using three dose levels might not 
be necessary. Expected human exposure though may indicate the need for 
a higher dose level.
    (e) Test procedures--(1) Animal selection--(i) Species and strain. 
Testing shall be performed in the species being used in other NTX 
tests. A standard strain of laboratory rat is recommended. The choice 
of species shall take into consideration such factors as the 
comparative metabolism of the chemical and species sensitivity to the 
toxic effects of the test substance, as evidenced by the results of 
other studies, the potential for combined studies, and the availability 
of other toxicity data for the species.
    (ii) Age. Animals shall be at least ten weeks of age at the start 
of exposure.
    (iii) Sex. Both sexes shall be used unless it is demonstrated that 
one sex is refractory to the effects of exposure.
    (2) Number of Animals. A minimum of ten animals per group shall be 
used. The tissues from each animal shall be examined separately.
    (3) Control Groups. (i) A concurrent control group, exposed to 
clean, filtered air only, is required.
    (ii) The laboratory performing the testing shall provide positive 
control data, e.g., results from repeated acrylamide exposure, as 
evidence of the ability of their histology procedures to detect 
neurotoxic endpoints. Positive control data shall be collected at the 
time of the test study unless the laboratory can demonstrate the 
adequacy of historical data for the planned study.
    (iii) A satellite group of 10 female and 10 male test subjects 
shall be treated with the highest concentration level for the duration 
of the exposure and observed thereafter for reversibility, persistence, 
or delayed occurrence of toxic effects during a post-treatment period 
of not less than 28 days.
    (4) Inhalation exposure. (i) All data developed within this study 
shall be in accordance with good laboratory practice provisions under 
Sec. 79.60.
    (ii) The general conduct of this study shall be in accordance with 
the vehicle emissions inhalation exposure guideline in Sec. 79.61.
    (5) Study conduct--(i) Observation of animals. All toxicological 
(e.g., weight loss) and neurological signs (e.g., motor disturbance) 
shall be recorded frequently enough to observe any abnormality, and not 
less than weekly.
    (ii) The following is a minimal list of measures that shall be 
noted:
    (A) Body weight;
    (B) Subject's reactivity to general stimuli such as removal from 
the cage or handling;
    (C) Description, incidence, and severity of any convulsions, 
tremors, or abnormal motor movements in the home cage;
    (D) Descriptions and incidence of posture and gait abnormalities 
observed in the home cage; and
    (E) Description and incidence of any unusual or abnormal behaviors, 
excessive or repetitive actions (stereotypies), emaciation, 
dehydration, hypotonia or hypertonia, altered fur appearance, red or 
crusty deposits around the eyes, nose, or mouth, and any other 
observations that may facilitate interpretation of the data.
    (iii) Sacrifice of animals--(A) General. The goal of the techniques 
outlined for sacrifice of animals and preparation of tissues is 
preservation of tissue morphology to simulate the living state of the 
cell.
    (B) Perfusion technique. Animals shall be perfused in situ by a 
generally recognized technique. For fixation suitable for light or 
electronic microscopy, saline solution followed by buffered 2.5 percent 
glutaraldehyde or buffered 4.0 percent paraformaldehyde, is 
recommended. While some minor modifications or variations in procedures 
are used in different laboratories, a detailed and standard procedure 
for vascular perfusion may be found in the text by Zeman and Innes 
(1963), Hayat (1970), and Spencer and Schaumburg (1980) under paragraph 
(g) of this section. A more sophisticated technique is described by 
Palay and Chan-Palay (1974) under paragraph (g) of this section.
    (C) Removal of brain and cord. After perfusion, the bony structure 
(cranium and vertebral column) shall be exposed. Animals shall then be 
stored in fixative-filled bags at 4  deg.C for 8-12 hours. The cranium 
and vertebral column shall be removed carefully by trained technicians 
without physical damage of the brain and cord. Detailed dissection 
procedures may be found in the text by Palay and Chan-Palay (1974) 
under paragraph (g) of this section. After removal, simple measurement 
of the size (length and width) and weight of the whole brain (cerebrum, 
cerebellum, pons-medulla) shall be made. Any abnormal coloration or 
discoloration of the brain and cord shall also be noted and recorded.
    (D) Sampling. Cross-sections of the following areas shall be 
examined: The forebrain, the center of the cerebrum, the midbrain, the 
cerebellum, and the medulla oblongata; the spinal cord at the cervical 
swelling (C3-C6), and proximal sciatic nerve (mid-thigh and 
sciatic notch) or tibial nerve (at knee). Other sites and tissue 
elements (e.g., gastrocnemius muscle) shall be examined if deemed 
necessary. Any observable gross changes shall be recorded.
    (iv) Specimen storage. Tissue samples from both the central and 
peripheral nervous system shall be further immersion fixed and stored 
in appropriate fixative (e.g., 10 percent buffered formalin for light 
microscopy; 2.5 percent buffered gluteraldehyde or 4.0 percent buffered 
paraformaldehyde for electron microscopy) for future examination. The 
volume of fixative versus the volume of tissues in a specimen jar shall 
be no less than 25:1. All stored tissues shall be washed with buffer 
for at least 2 hours prior to further tissue processing.
    (v) Histopathology examination--(A) Fixation. Tissue specimens 
stored in 10 percent buffered formalin may be used for this purpose. 
All tissues must be immersion fixed in fixative for at least 48 hours 
prior to further tissue processing.
    (B) Dehydration. All tissue specimens shall be washed for at least 
1 hour with water or buffer, prior to dehydration. (A longer washing 
time is needed if the specimens have been stored in fixative for a 
prolonged period of time.) Dehydration can be performed with increasing 
concentration of graded ethanols up to absolute alcohol.
    (C) Clearing and embedding. After dehydration, tissue specimens 
shall be cleared with xylene and embedded in paraffin or paraplast. 
Multiple tissue specimens (e.g. brain, cord, ganglia) may be embedded 
together in one single block for sectioning. All tissue blocks shall be 
labelled showing at least the experiment number, animal number, and 
specimens embedded.
    (D) Sectioning. Tissue sections, 5 to 6 microns in thickness, shall 
be prepared from the tissue blocks and mounted on standard glass 
slides. It is recommended that several additional sections be made from 
each block at this time for possible future needs for special 
stainings. All tissue blocks and slides shall be filed and stored in 
properly labeled files or boxes.
    (E) Histopathological techniques. The following general testing 
sequence is proposed for gathering histopathological data:
    (1) General staining. A general staining procedure shall be 
performed on all tissue specimens in the highest treatment group. 
Hematoxylin and eosin (H&E) shall be used for this purpose. The 
staining shall be differentiated properly to achieve bluish nuclei with 
pinkish background.
    (2) Peripheral nerve teasing. Peripheral nerve fiber teasing shall 
be used. Detailed staining methodology is available in standard 
histotechnological manuals such as AFIP (1968), Ralis et al. (1973), 
and Chang (1979) under paragraph (g) of this section. The nerve fiber 
teasing technique is discussed in Spencer and Schaumberg (1980) under 
paragraph (g) of this section. A section of normal tissue shall be 
included in each staining to assure that adequate staining has 
occurred. Any changes shall be noted and representative photographs 
shall be taken. If a lesion(s) is observed, the special techniques 
shall be repeated in the next lower treatment group until no further 
lesion is detectable.
    (F) Examination. All stained microscopic slides shall be examined 
with a standard research microscope. Examples of cellular alterations 
(e.g., neuronal vacuolation, degeneration, and necrosis) and tissue 
changes (e.g., gliosis, leukocytic infiltration, and cystic formation) 
shall be recorded and photographed.
    (f) Data collection, reporting, and evaluation. In addition to 
information meeting the requirements stated under 40 CFR 79.60 and 
79.61, the following specific information shall be reported:
    (1) Description of test system and test methods. (i) A description 
of the general design of the experiment shall be provided. This shall 
include a short justification explaining any decisions where 
professional judgment is involved such as fixation technique and choice 
of stains; and
    (ii) Positive control data from the laboratory performing the test 
that demonstrate the sensitivity of the procedures being used. 
Historical data may be used if all essential aspects of the 
experimental protocol are the same.
    (2) Results. All observations shall be recorded and arranged by 
test groups. This data may be presented in the following recommended 
format:
    (i) Description of signs and lesions for each animal. For each 
animal, data must be submitted showing its identification (animal 
number, treatment, dose, duration), neurologic signs, location(s) 
nature of, frequency, and severity of lesion(s). A commonly-used scale 
such as 1+, 2+, 3+, and 4+ for degree of severity ranging from very 
slight to extensive may be used. Any diagnoses derived from neurologic 
signs and lesions including naturally occurring diseases or conditions, 
shall also be recorded;
    (ii) Counts and incidence of lesions, by test group. Data shall be 
tabulated to show:
    (A) The number of animals used in each group, the number of animals 
displaying specific neurologic signs, and the number of animals in 
which any lesion was found; and
    (B) The number of animals affected by each different type of 
lesion, the average grade of each type of lesion, and the frequency of 
each different type and/or location of lesion.
    (iii) Evaluation of data. (A) An evaluation of the data based on 
gross necropsy findings and microscopic pathology observations shall be 
made and supplied. The evaluation shall include the relationship, if 
any, between the animal's exposure to the test atmosphere and the 
frequency and severity of any lesions observed; and
    (B) The evaluation of dose-response, if existent, for various 
groups shall be given, and a description of statistical method must be 
presented. The evaluation of neuropathology data shall include, where 
applicable, an assessment in conjunction with any other neurotoxicity 
studies, electrophysiological, behavioral, or neurochemical, which may 
be relevant to this study.
    (g) References. For additional background information on this test 
guideline, the following references should be consulted.

(1) 40 CFR 798.6400, Neuropathology.
(2) AFIP Manual of Histologic Staining Methods. (New York: McGraw-
Hill (1968).
(3) Chang, L.W. A Color Atlas and Manual for Applied Histochemistry. 
(Springfield, IL: Charles C. Thomas, 1979).
(4) Dunnick, J.K., et.al. Thirteen-week Toxicity Study of N-Hexane 
in B6C3F1 Mice After Inhalation Exposure (1989) Toxicology, 57, 163-
172.
(5) Hayat, M.A. ``Vol. 1. Biological applications,'' Principles and 
techniques of electron microscopy. (New York: Van Nostrand Reinhold, 
1970).
(6) Palay S.L., Chan-Palay, V. Cerebellar Cortex: Cytology and 
Organization. (New York: Springer-Verlag, 1974).
(7) Ralis, H.M., Beesley, R.A., Ralis, Z.A. Techniques in 
Neurohistology. (London: Butterworths, 1973).
(8) Sette, W. ``Pesticide Assessment Guidelines, Subdivision F, 
Neurotoxicity Test Guidelines.'' Report No. 540/09-91-123 U.S. 
Environmental Protection Agency 1991 (NTIS #PB91-154617).
(9) Spencer, P.S., Schaumburg, H.H. (eds). Experimental and Clinical 
Neurotoxicology. (Baltimore: Williams and Wilkins, 1980).
(10) Zeman, W., Innes, J.R.M. Craigie's Neuroanatomy of the Rat. 
(New York: Academic, 1963).


Sec. 79.67  Glial fibrillary acidic protein assay.

    (a) Purpose. Chemical-induced injury of the nervous system, i.e., 
the brain, is associated with astrocytic hypertrophy at the site of 
damage (see O'Callaghan, 1988 in paragraph (e)(3) in this section). 
Assays of glial fibrillary acidic protein (GFAP), the major 
intermediate filament protein of astrocytes, can be used to document 
this response. To date, a diverse variety of chemical insults known to 
be injurious to the central nervous system have been shown to increase 
GFAP. Moreover, increases in GFAP can be seen at concentrations below 
those necessary to produce cytopathology as determined by routine Nissl 
stains (standard neuropathology). Thus it appears that assays of GFAP 
represent a sensitive approach for documenting the existence and 
location of chemical-induced injury of the central nervous system. 
Additional functional, histopathological, and biochemical tests are 
necessary to assess completely the neurotoxic potential of any 
chemical. This biochemical test is intended to be used in conjunction 
with neurohistopathological studies.
    (b) Principle of the test method. (1) This guideline describes the 
conduct of a radioimmunoassay for measurement of the amount of GFAP in 
the brain of vehicle emission-exposed and unexposed control animals. It 
is based on modifications (O'Callaghan & Miller 1985 in paragraph 
(e)(5), O'Callaghan 1987 in paragraph (e)(1) of this section) of the 
dot-immunobinding procedure described by Jahn et al. (1984) in 
paragraph (e)(2) of this section. Briefly, brain tissue samples from 
study animals are assayed for total protein, diluted in dot-
immunobinding buffer, and applied to nitrocellulose sheets. The spotted 
sheets are then fixed, blocked, washed and incubated in anti-GFAP 
antibody and [I125] Protein A. Bound protein A is then quantified 
by gamma spectrometry. In lieu of purified protein standards, standard 
curves are constructed from dilution of a single control sample. By 
comparing the immunoreactivity of individual samples (both control and 
exposed groups) with that of the sample used to generate the standard 
curve, the relative immunoreactivity of each sample is obtained. The 
immunoreactivity of the control groups is normalized to 100 percent and 
all data are expressed as a percentage of control. A variation on this 
radioimmunoassay procedure has been proposed (O'Callaghan 1991 in 
paragraph (e)(4) of this section) which uses a ``sandwich'' of GFAP, 
anti-GFAP, and a chromophore in a microtiter plate format enzyme-link 
immunosorbent assay (ELISA). The use of this variation shall be 
justified.
    (2) This assay may be done separately or in combination with the 
subchronic toxicity study, pursuant to the provisions of Sec. 79.62.
    (c) Test procedure--(1) Animal selection--(i) Species and strain. 
Test shall be performed on the species being used in concurrent testing 
for neurotoxic or other health effect endpoints. This will generally be 
a species of laboratory rat. The use of other rodent or non-rodent 
species shall be justified.
    (ii) Age. Based on other concurrent testing, young adult rats shall 
be used. Study rodents shall not be older than ten weeks at the start 
of exposures.
    (iii) Number of animals. A minimum of ten animals per group shall 
be used. The tissues from each animal shall be examined separately.
    (iv) Sex. Both sexes shall be used unless it is demonstrated that 
one sex is refractory to the effects.
    (2) Materials. The materials necessary to perform this study are 
[I125] Protein A (2-10 Ci/g), Anti-sera to GFAP, 
nitrocellulose paper (0.1 or 0.2 m pore size), sample 
application template (optional; e.g., ``Minifold II'', Schleicher & 
Schuell, Keene, NH), plastic sheet incubation trays.
    (3) Study conduct. (i) All data developed within this study shall 
be in accordance with good laboratory practice provisions under 
Sec. 79.60.
    (ii) Tissue Preparation. Animals are euthanized 24 hours after the 
last exposure and the brain is excised from the skull. On a cold 
dissecting platform, the following six regions are dissected freehand: 
cerebellum; cerebral cortex; hippocampus; striatum; thalamus/
hypothalamus; and the rest of the brain. Each region is then weighed 
and homogenized in 10 volumes of hot (70-90  deg.C) 1 percent (w/v) 
sodium dodecyl sulfate (SDS). Homogenization is best achieved through 
sonic disruption. A motor driven pestle inserted into a tissue grinding 
vessel is a suitable alternative. The homogenized samples can then be 
stored frozen at -70  deg.C for at least 4 years without loss of GFAP 
content.
    (iii) Total Protein Assay. Aliquots of the tissue samples are 
assayed for total protein using the method of Smith et al. (1985) in 
paragraph (e)(7) of this section. This assay may be purchased in kit 
form (e.g., Pierce Chemical Company, Rockford, IL).
    (iv) Sample Preparation. Dilute tissue samples in sample buffer 
(120 mM KCl, 20 mM NaCl, 2 mM MgCl2), 5 mM Hepes, pH 7.4, 0.7 
percent Triton X-100) to a final concentration of 0.25 mg total protein 
per ml (5 g/20 l).
    (v) Preparation of Standard Curve. Dilute a single control sample 
in sample buffer to give at least five standards, between 1 and 10 
g total protein per 20 l. The suggested values of 
total protein per 20 l sample buffer are 1.25, 2.50, 3.25, 
5.0, 6.25, 7.5, 8.75, and 10.0 g.
    (vi) Preparation of Nitrocellulose Sheets. Nitrocellulose sheets of 
0.1 or 0.2 micron pore size are rinsed by immersion in distilled water 
for 5 minutes and then air dried.
    (vii) Sample Application. Samples can be spotted onto the 
nitrocellulose sheets free-hand or with the aid of a template. For 
free-hand application, draw a grid of squares approximately 2 
centimeters by 2 centimeters (cm) on the nitrocellulose sheets using a 
soft pencil. Spot 5-10 l portions to the center of each square 
for a total sample volume of 20 l. For template aided sample 
application a washerless microliter capacity sample application 
manifold is used. Position the nitrocellulose sheet in the sample 
application device as recommended by the manufacturer and spot a 20 
l sample in one application. Do not wet the nitrocellulose or 
any support elements prior to sample application. Do not apply vacuum 
during or after sample application. After spotting samples (using 
either method), let the sheets air dry. The sheets can be stored at 
room temperature for several days after sample application.
    (viii) Standard Incubation Conditions. These conditions have been 
described by Jahn et al. (1984) in paragraph (e)(2) of this section. 
All steps are carried out at room temperature on a flat shaking 
platform (one complete excursion every 2-3 seconds). For best results, 
do not use rocking or orbital shakers. Perform the following steps in 
enough solution to cover the nitrocellulose sheets to a depth of 1 cm.
    (A) Incubate 20 minutes in fixer (25 percent (v/v) isopropanol, 10 
percent (v/v) acetic acid).
    (B) Discard fixer, wash several times in deionized water to 
eliminate the fixer, and then incubate for 5 minutes in Tris-buffered 
saline (TBS): 200 mM NaCL, 60 mM Tris-HCl to pH 7.4.
    (C) Discard TBS and incubate 1 hour in blocking solution (0.5 
percent gelatin (w/v)) in TBS.
    (D) Discard blocking solution and incubate for 2 hours in antibody 
solution (anti-GFAP antiserum diluted to the desired dilution in 
blocking solution containing 0.1 percent Triton X-100). Serum anti-
bovine GFAP, which cross reacts with GFAP from rodents and humans, can 
be obtained commercially (e.g., Dako Corp.) and used at a dilution of 
1:500.
    (E) Discard antibody solution, and wash in 4 changes of TBS for 5 
minutes each time. Then wash in TBS for 10 minutes.
    (F) Discard TBS and incubate in blocking solution for 30 minutes.
    (G) Discard blocking solution and incubate for 1 hour in Protein A 
solution ([I\125\]-labeled Protein A diluted in blocking solution 
containing 0.1 percent Triton X-100, sufficient to produce 2000 counts 
per minute (cpm) per 10 l of Protein A solution).
    (H) Remove Protein A solution (it may be reused once). Wash in 0.1 
percent Triton X-100 in TBS (TBSTX) for 5 minutes, 4 times. Then wash 
in TBSTX for 2-3 hours for 4 additional times. An overnight wash in a 
larger volume can be used to replace the last 4 washes.
    (I) Hang sheets to air-dry. Cut out squares or spots and count 
radioactivity in a gamma counter.
    (ix) Expression of data. Compare radioactivity counts for samples 
obtained from control and treated animals with counts obtained from the 
standard curve. By comparing the immunoreactivity (counts) of each 
sample with that of the standard curve, the relative amount of GFAP in 
each sample can be determined and expressed as a percent of control.
    (d) Data Reporting and Evaluation--(1) Test Report. In addition to 
information meeting the requirements stated under 40 CFR 79.60, the 
following specific information shall be reported:
    (i) Body weight and brain region weights at time of sacrifice for 
each subject tested;
    (ii) Indication of whether each subject survived to sacrifice or 
time of death;
    (iii) Data from control animals and blank samples; and
    (iv) Statistical evaluation of results;
    (2) Evaluation of Results. (i) Results shall be evaluated in terms 
of the extent of change in the amount of GFAP as a function of 
treatment and dose. GFAP assays (of any brain region) from a minimum of 
6 samples typically will result in a standard error of the mean of +/- 
5 percent. In this case, a chemically-induced increase in GFAP of 115 
percent of control is likely to be statistically significant.
    (ii) The results of this assay shall be compared to and evaluated 
with any relevant behavioral and histopathological data.
    (e) References. For additional background information on this test 
guideline the following references should be consulted.

(1) Brock, T.O and O'Callaghan, J.P. 1987. Quantitative changes in 
the synaptic vesicle proteins, synapsin I and p38 and the astrocyte 
specific protein, glial fibrillary acidic protein, are associated 
with chemical-induced injury to the rat central nervous system, J. 
Neurosci. 7:931-942.
(2) Jahn, R., Schiebler, W. Greengard, P. 1984. A quantitative dot-
immunobinding assay for protein using nitrocellulose membrane 
filters. Proc. Natl. Acad. Sci. U.S.A. 81:1684-1687.
(3) O'Callaghan, J.P. 1988. Neurotypic and gliotypic protein as 
biochemical markers of neurotoxicity. Neurotoxicol. Teratol. 10:445-
452.
(4) O'Callaghan, J.P. 1991. Quantification of glial fibrillary 
acidic protein: comparison of slot-immunobinding assays with a novel 
sandwich ELISA. Neurotoxicol. Teratol. 13:275-281.
(5) O'Callaghan, J.P. and Miller, D.B. 1985. Cerebellar hypoplasia 
in the Gunn rat is associated with quantitative changes in 
neurotypic and gliotypic proteins. J. Pharmacol. Exp. Ther. 234:522-
532.
(6) Sette, W.F. ``Pesticide Assessment Guidelines, Subdivision `F', 
Hazard Evaluation: Human and Domestic Animals, Addendum 10, 
Neurotoxicity, Series 81, 82, and 83'' US-EPA, Office of Pesticide 
Programs, EPA-540/09-91-123, March 1991.
(7) Smith, P.K., Krohn, R.I., Hermanson, G.T., Mallia, A.K., 
Gartner, F.H., Provenzano, M.D., Fujimoto, E.K., Goeke, N.M., Olson, 
B.J., Klenk, D.C. 1985. Measurement of protein using bicinchoninic 
acid. Annal. Biochem. 150:76-85.


Sec. 79.68  Salmonella typhimurium reverse mutation assay.

    (a) Purpose. The Salmonella typhimurium histidine (his) reversion 
system is a microbial assay which measures his- his+ 
reversion induced by chemicals which cause base changes or frameshift 
mutations in the genome of the microorganism Salmonella typhimurium.
    (b) Definitions. For the purposes of this section, the following 
definitions apply:

Base pair mutagen means an agent which causes a base change in DNA. In 
a reversion assay, this change may occur at the site of the original 
mutation or at a second site in the chromosome.
Frameshift mutagen is an agent which causes the addition or deletion of 
single or multiple base pairs in the DNA molecule.
Salmonella typhimurium reverse mutation assay detects mutation in a 
gene of a histidine-requiring strain to produce a histidine independent 
strain of this organism.

    (c) Reference substances. These may include, but need not be 
limited to, sodium azide, 2-nitrofluorene, 9-aminoacridine, 2-
aminoanthracene, congo red, benzopurpurin 4B, trypan blue or direct 
blue 1.
    (d) Test method.--(1) Principle. Motor vehicle combustion emissions 
from fuel or additive/base fuel mixtures are, first, filtered to trap 
particulate matter and, then, passed through a sorbent resin to trap 
semi-volatile gases. Bacteria are separately exposed to the extract 
from both the filtered particulates and the resin-trapped organics. 
Assays are conducted using both test mixtures with and without a 
metabolic activation system and exposed cells are plated onto minimal 
medium. After a suitable period of incubation, revertant colonies are 
counted in test cultures and compared to the number of spontaneous 
revertants in unexposed control cultures.
    (2) Description. Several methods for performing the test have been 
described. The procedures described here are for the direct plate 
incorporation method and the azo-reduction method. Among those used 
are:
    (i) Direct plate incorporation method;
    (ii) Preincubation method;
    (iii) Azo-reduction method;
    (iv) Microsuspension method; and
    (v) Spiral assay.
    (3) Strain selection--(i)Designation. Five tester strains shall be 
used in the assay. At the present time, TA1535, TA1537, TA98, and TA100 
are designated as tester strains. The fifth strain will be chosen from 
the pool of Salmonella strains commonly used to determine the degree to 
which nitrated organic compounds, i.e., nitroarenes, contribute to the 
overall mutagenic activity of a test substance. TA98/1,8-DNP6 or 
other suitable Rosenkranz nitro-reductase resistant strains will be 
considered acceptable. The choice of the particular strain is left to 
the discretion of the researcher. However, the researcher shall justify 
the use of the selected bacterial tester strains.
    (ii) Preparation and storage of bacterial tester strains. 
Recognized methods of stock culture preparation and storage shall be 
used. The requirement of histidine for growth shall be demonstrated for 
each strain. Other phenotypic characteristics shall be checked using 
such methods as crystal violet sensitivity and resistance to 
ampicillin. Spontaneous reversion frequency shall be in the range 
expected as reported in the literature and as established in the 
laboratory by historical control values.
    (iii) Bacterial growth. Fresh cultures of bacteria shall be grown 
up to the late exponential or early stationary phase of growth 
(approximately 108-109 cells per ml).
    (4) Exogenous metabolic activation. Bacteria shall be exposed to 
the test substance both in the presence and absence of an appropriate 
exogenous metabolic activation system. For the direct plate 
incorporation method, the most commonly used system is a cofactor-
supplemented postmitochondrial fraction prepared from the livers of 
rodents treated with enzyme-inducing agents, such as Aroclor 1254. For 
the azo-reduction method, a cofactor- supplemented postmitochondrial 
fraction (S-9) prepared from the livers of untreated hamsters is 
preferred. For this method, the cofactor supplement shall contain 
flavin mononucleotide, exogenous glucose 6-phosphate dehydrogenase, 
NADH and excess of glucose-6-phosphate.
    (5) Control groups--(i) Concurrent controls. Concurrent positive 
and negative (untreated) controls shall be included in each experiment. 
Positive controls shall ensure both strain responsiveness and efficacy 
of the metabolic activation system.
    (ii) Strain specific positive controls shall be included in the 
assay. Examples of strain specific positive controls are as follows:

(A) Strain TA1535, TA100: sodium azide;
(B) TA98: 2-nitrofluorene (without activation), 2-anthramine (with 
activation);
(C) TA1537: 9-aminoacridine; and
(D) TA98/1,8-DNP6: benzo(a)pyrene (with activation).
    The papers by Claxton et al., 1991 and 1992 in paragraph (g) in 
this section will provide helpful information for the selection of 
positive controls.
    (iii) Positive controls to ensure the efficacy of the activation 
system. The positive control reference substances for tests including a 
metabolic activation system shall be selected on the basis of the type 
of activation system used in the test. 2-Aminoanthracene is an example 
of a positive control compound in plate-incorporation tests using 
postmitochondrial fractions from the livers of rodents treated with 
enzyme-inducing agents such as Aroclor-1254. Congo red is an example of 
a positive control compound in the azo-reduction method. Other positive 
control reference substances may be used.
    (iv) Class-specific positive controls. The azo-reduction method 
shall include positive controls from the same class of compounds as the 
test agent wherever possible.
    (6) Sampling the test atmosphere.--(i) Extracts of test emissions 
are collected on Teflon-coated glass fiber filters using an 
exhaust dilution setup. The particulates are extracted with 
dichloromethane (DCM) using Soxhlet extraction techniques. Extracts in 
DCM can be stored at dry ice temperatures until use.
    (ii) Gaseous hydrocarbons passing through the filter are trapped by 
a porous, polymer resin, like XAD-2/styrene-divinylbenzene, or an 
equivalent product. Methylene chloride is used to extract the resin and 
the sample is evaporated to dryness before storage or use.
    (iii) Samples taken from this material are then used to expose the 
cells in this assay. Final concentration of extracts in solvent/
vehicle, or after solvent exchange, shall not interfere with cell 
viability or growth rate. The paper by Stump (1982) in paragraph (g) of 
this section is useful for preparing extracts of particulate and semi-
volatile organic compounds from diesel and gasoline exhaust stream.
    (iv) Exposure concentrations. (A) The test should initially be 
performed over a broad range of concentrations. Among the criteria to 
be taken into consideration for determining the upper limits of test 
substance concentration are cytotoxicity and solubility. Cytotoxicity 
of the test chemical may be altered in the presence of metabolic 
activation systems. Toxicity may be evidenced by a reduction in the 
number of spontaneous revertants, a clearing of the background lawn or 
by the degree of survival of treated cultures. Relatively insoluble 
samples shall be tested up to the limits of solubility. The upper test 
chemical concentration shall be determined on a case by case basis.
    (B) Generally, a maximum of 5 mg/plate for pure substances is 
considered acceptable. At least 5 different concentrations of test 
substance shall be used with adequate intervals between test points.
    (C) When appropriate, a single positive response shall be confirmed 
by testing over a narrow range of concentrations.
    (e) Test performance. All data developed within this study shall be 
in accordance with good laboratory practice provisions under 
Sec. 79.60.
    (1) Direct plate incorporation method. When testing with metabolic 
activation, test solution, bacteria, and 0.5 ml of activation mixture 
containing an adequate amount of postmitochondrial fraction shall be 
added to the liquid overlay agar and mixed. This mixture is poured over 
the surface of a selective agar plate. Overlay agar shall be allowed to 
solidify before incubation. At the end of the incubation period, 
revertant colonies per plate shall be counted. When testing without 
metabolic activation, the test sample and 0.1 ml of a fresh bacterial 
culture shall be added to 2.0 ml of overlay agar.
    (2) Azo-reduction method. When testing with metabolic activation, 
0.5 ml of activation mixture containing 150 l of 
postmitochondrial fraction and 0.1 ml of bacterial culture shall be 
added to a test tube kept on ice. 0.1 ml of test solution shall be 
added, and the tubes shall be incubated with shaking at 30  deg.C for 
30 minutes. At the end of the incubation period, 2.0 ml of agar shall 
be added to each tube, the contents mixed and poured over the surface 
of a selective agar plate. Overlay agar shall be allowed to solidify 
before incubation. At the end of the incubation period, revertant 
colonies per plate shall be counted. For tests without metabolic 
activation, 0.5 ml of buffer shall be used in place of the 0.5 ml of 
activation mixture. All other procedures shall be the same as those 
used for the test with metabolic activation.
    (3) Other methods/modifications may also be appropriate.
    (4) Media. An appropriate selective medium with an adequate overlay 
agar shall be used.
    (5) Incubation conditions. All plates within a given experiment 
shall be incubated for the same time period. This incubation period 
shall be for 48-72 hours at 37  deg.C.
    (6) Number of cultures. All plating shall be done at least in 
triplicate.
    (f) Data and report--(1) Treatment of results. Data shall be 
presented as number of revertant colonies per plate, revertants per 
kilogram (or liter) of fuel, and as revertants per kilometer (or mile) 
for each replicate and dose. These same measures shall be recorded on 
both the negative and positive control plates. The mean number of 
revertant colonies per plate, revertants per kilogram (or liter) of 
fuel, and revertants per kilometer (or mile), as well as individual 
plate counts and standard deviations shall be presented for the test 
substance, positive control, and negative control plates.
    (2) Statistical evaluation. Data shall be evaluated by appropriate 
statistical methods. Those methods shall include, at a minimum, means 
and standard deviations of the reversion data.
    (3) Interpretation of results. (i) There are several criteria for 
determining a positive result, one of which is a statistically 
significant dose-related increase in the number of revertants. Another 
criterion may be based upon detection of a reproducible and 
statistically significant positive response for at least one of the 
test substance concentrations.
    (ii) A test substance which does not produce either a statistically 
significant dose-related increase in the number of revertants or a 
statistically significant and reproducible positive response at any one 
of the test points is considered nonmutagenic in this system.
    (iii) Both biological and statistical significance shall be 
considered together in the evaluation.
    (4) Test evaluation. (i) Positive results from the Salmonella 
typhimurium reverse mutation assay indicate that, under the test 
conditions, the test substance induces point mutations by base changes 
or frameshifts in the genome of this organism.
    (ii) Negative results indicate that under the test conditions the 
test substance is not mutagenic in Salmonella typhimurium.
    (5) Test report. In addition to the reporting recommendations as 
specified under 40 CFR 79.60, the following specific information shall 
be reported:
    (i) Sampling method(s) used and manner in which cells are exposed 
to sample solution;
    (ii) Bacterial strains used;
    (iii) Metabolic activation system used (source, amount and 
cofactor); details of preparation of postmitochondrial fraction;
    (vi) Concentration levels and rationale for selection of 
concentration range;
    (v) Description of positive and negative controls, and 
concentrations used, if appropriate;
    (vi) Individual plate counts, mean number of revertant colonies per 
plate, number of revertants per mile (or kilometer), and standard 
deviation; and
    (vii) Dose-response relationship, if applicable.
    (g) References. For additional background information on this test 
guideline, the following references should be consulted.

(1) 40 CFR 798.5265, The Salmonella typhimurium reverse mutation 
assay.
(2) Ames, B.N., McCann, J., Yamasaki, E. ``Methods for detecting 
carcinogens and mutagens with the Salmonella/mammalian microsome 
mutagenicity test,'' Mutation Research 31:347-364 (1975).
(3) Huisingh, J.L., et al.,``Mutagenic and Carcinogenic Potency of 
Extracts of Diesel and Related Environmental Emissions: Study 
Design, Sample Generation, Collection, and Preparation''. In: Health 
Effects of Diesel Engine Emissions, Vol. II, W.E. Pepelko, R., M., 
Danner and N. A. Clarke (Eds.), US EPA, Cincinnati, EPA-600/9-80-
057b, pp. 788-800 (1980).
(5) Claxton, L.D., Allen, J., Auletta, A., Mortelmans, K., Nestmann, 
E., Zeiger, E. ``Guide for the Salmonella typhimurium/mammalian 
microsome tests for bacterial mutagenicity'' Mutation Research 
189(2):83-91 (1987).
(6) Claxton, L., Houk, V.S., Allison, J.C., Creason, J., 
``Evaluating the relationship of metabolic activation system 
concentrations and chemical dose concentrations for the Salmonella 
Spiral and Plate Assays'' Mutation Research 253:127-136 (1991).
(7) Claxton, L., Houk, V.S., Monteith, L.G., Myers, L.E., Hughes, 
T.J., ``Assessing the use of known mutagens to calibrate the 
Salmonella typhimurium mutagenicity assay: I. Without exogenous 
activation.'' Mutation Research 253:137-147 (1991).
(8) Claxton, L., Houk, V.S., Warner, J.R., Myers, L.E., Hughes, 
T.J., ``Assessing the use of known mutagens to calibrate the 
Salmonella typhimurium mutagenicity assay: II. With exogenous 
activation.'' Mutation Research 253:149-159 (1991).
(9) Claxton, L., Creason, J., Lares, B., Augurell, E., Bagley, S., 
Bryant, D.W., Courtois, Y.A., Douglas, G., Clare, C.B., Goto, S., 
Quillardet, P., Jagannath, D.R., Mohn, G., Neilsen, P.A., Ohnishi, 
Y., Ong, T., Pederson, T.C., Shimizu, H., Nylund, L., Tokiwa, H., 
Vink, I.G.R., Wang, Y., Warshawsky, D., ``Results of the IPCS 
Collaborative Study on Complex Mixtures'' Mutation Research 276:23-
32 (1992).
(10) Claxton, L., Douglas, G., Krewski, D., Lewtas, J., Matsushita, 
H., Rosenkranz, H., ``Overview, conclusions, and recommendations of 
the IPCS Collaborative Study on Complex Mixtures'' Mutation Research 
276:61-80 (1992).
(11) Houk, V.S., Schalkowsky, S., and Claxton, L.D., ``Development 
and Validation of the Spiral Salmonella Assay: An Automated Approach 
to Bacterial Mutagenicity Testing'' Mutation Research 223:49-64 
(1989).
(12) Jones, E., Richold, M., May, J.H., and Saje, A. ``The 
Assessment of the Mutagenic Potential of Vehicle Engine Exhaust in 
the Ames Salmonella Assay Using a Direct Exposure Method'' Mutation 
Research 97:35-40 (1985).
(13) Maron, D., and Ames, B. N., Revised methods for the Salmonella 
mutagenicity test, Mutation Research, 113:173-212 (1983).
(14) Prival, M.J., and Mitchell, V.D. ``Analysis of a method for 
testing azo dyes for mutagenic activity in Salmonella typhimurium in 
the presence of flavin mononucleotide and hamster liver S-9,'' 
Mutation Research 97:103-116 (1982).
(15) Rosenkranz, H.S., et.al. ``Nitropyrenes: Isolation, 
identification, and reduction of mutagenic impurities in carbon 
black and toners'' Science 209:1039-43 (1980).
(16) Stump, F., Snow, R., et.al., ``Trapping gaseous hydrocarbons 
for mutagenic testing'' SAE Technical Paper Series, No. 820776 
(1982).
(17) Vogel, H.J., Bonner, D.M. ``Acetylornithinase of E. coli: 
partial purification and some properties,'' Journal of Biological 
Chemistry. 218:97-106 (1956).

[FR Doc. 94-13784 Filed 6-24-94; 8:45 am]
BILLING CODE 6560-50-P