[Federal Register Volume 63, Number 49 (Friday, March 13, 1998)]
[Proposed Rules]
[Pages 12421-12426]
From the Federal Register Online via the Government Publishing Office [www.gpo.gov]
[FR Doc No: 98-6571]


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DEPARTMENT OF HEALTH AND HUMAN SERVICES

Food and Drug Administration

21 CFR Part 184

[Docket No. 89G-0393]


Direct Food Substances Affirmed as Generally Recognized as Safe; 
Egg White Lysozyme

AGENCY: Food and Drug Administration, HHS.

ACTION: Tentative final rule.

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SUMMARY: The Food and Drug Administration (FDA) is issuing a tentative 
final rule to amend its regulations to affirm that egg white lysozyme 
enzyme preparation, when labeled by the common or usual name ``egg 
white lysozyme'' to identify its source, is generally recognized as 
safe (GRAS) for use in preventing late blowing of cheese caused by the 
bacterium Clostridium tyrobutyricum during cheese production. This 
action is in response to a petition submitted by Fordras S.A. (formerly 
SPA-Societa Prodotti Antibiotici S.p.A.). FDA has tentatively concluded 
that this use of the egg white lysozyme enzyme preparation is GRAS only 
when the ingredient statement for both bulk and packaged food that 
contains cheese manufactured using egg white lysozyme includes the 
common or usual name ``egg white lysozyme'' to identify the source of 
the protein. To give interested persons an opportunity to comment on 
this condition of use required for GRAS status, FDA is issuing this 
tentative final rule.

DATES: Submit written comments by May 27, 1998.

ADDRESSES: Submit written comments to the Dockets Management Branch 
(HFA-305), Food and Drug Administration, 12420 Parklawn Dr., rm. 1-23, 
Rockville, MD 20857.

FOR FURTHER INFORMATION CONTACT: Linda S. Kahl, Center for Food Safety 
and Applied Nutrition (HFS-206), Food and Drug Administration, 200 C 
St. SW., Washington, DC 20204, 202-418-3101.

SUPPLEMENTARY INFORMATION: 

I. Background

    In accordance with the procedures described in Sec. 170.35 (21 CFR 
170.35), SPA-Societa Prodotti Antibiotici S.p.A., now Fordras S.A., 
Milan, Italy, submitted a petition (GRASP 9G0355) requesting that egg 
white lysozyme used to inhibit the bacterium C. tyrobutyricum to 
prevent late blowing of cheese during production be affirmed as GRAS as 
a direct human food ingredient. FDA published the notice of filing for 
this petition in the Federal Register of October 27, 1989 (54 FR 
43861), and gave interested persons until December 26, 1989, to submit 
written comments.

II. Standards for GRAS Affirmation

    Under Sec. 170.30 (21 CFR 170.30), general recognition of safety 
may be based only on the views of experts qualified by scientific 
training and experience to evaluate the safety of substances directly 
or indirectly added to food. The basis of such views may be either: (1) 
Scientific procedures, or (2) in the case of a substance used in food 
prior to January 1, 1958, through experience based on common use in 
food. General recognition of safety based upon scientific procedures 
requires the same quantity and quality of scientific evidence as is 
required to obtain approval of a food additive regulation and 
ordinarily is based upon published studies, which may be corroborated 
by unpublished studies and other data and information (Sec. 170.30(b)). 
General recognition of safety through experience based on common use in 
food prior to January 1, 1958, may be determined without the quantity 
or quality of scientific procedures required for approval of a food 
additive regulation, but ordinarily is based upon generally available 
data and information concerning the pre-1958 history of use of the 
substance.
    FDA has evaluated Fordras S.A.'s petition on the basis of 
scientific procedures to whether the petitioned use of egg white 
lysozyme enzyme preparation to prevent the late blowing of cheese 
caused by the bacterium C. tyrobutyricum during cheese production is 
GRAS. In evaluating the petition, FDA considered published and 
unpublished data and information relating to the identity of, 
characteristic properties of, and estimated dietary exposure to the 
enzyme component (i.e., lysozyme) of the petitioned enzyme preparation 
(Refs. 1 through 7). FDA also considered that the source of the 
petitioned enzyme preparation, egg white, has been safely consumed by 
humans as a source of food protein throughout recorded history, and, 
therefore, is GRAS (Sec. 170.30(d)), and that the methods used for 
extracting lysozyme from the egg white source do not ordinarily alter 
the chemical identity and characteristic properties of enzymes (Ref. 
8). FDA also considered published scientific review articles (Refs. 1 
and 2) and a generally available trade association bulletin (Ref. 7) 
discussing the use of egg white lysozyme enzyme preparation for its

[[Page 12422]]

technical effect of preventing late blowing of cheese contaminated with 
C. tyrobutyricum as well as generally available information documenting 
that this intended use of the petitioned enzyme preparation has been 
approved in several European countries (Refs. 9 through 13). Finally, 
FDA considered generally available and accepted information relating to 
processing aids used in the manufacture of the enzyme preparation and 
generally available and accepted specifications for food-grade enzyme 
preparations (Ref. 14).

III. Safety Evaluation

    When present as a contaminant in milk used for cheesemaking, the 
pasteurization-resistant bacterium C. tyrobutyricum ferments lactate to 
produce carbon dioxide, hydrogen, and volatile organic acids. This 
fermentation causes a defect in cheese manufacture known as ``late 
blowing,'' which is typified by abnormal levels of open texture 
accompanied by undesirable odors and flavors. Late blowing can be a 
serious economic problem in the manufacture of several varieties of 
cheese (Refs. 1, 2, and 7).
    The contamination by C. tyrobutyricum of milk used for 
cheesemaking, although reducible by good husbandry and hygienic milking 
practices, is unavoidable. Although treatment with certain chemical 
agents has been shown to be effective against the problems raised by 
this contamination, treatment with lysozyme enzyme preparation has been 
found to be the most effective method of managing the late blowing of 
cheese contaminated with C. tyrobutyricum (Refs. 1 and 2).

A. The Enzyme Component

    Enzymes are proteins or conjugated proteins (i.e., a protein that 
contains a nonamino acid moiety such as a carbohydrate) produced by 
plants, animals, and microorganisms that function as biochemical 
catalysts (American Heritage Dictionary of the English Language). Most 
enzymes are very specific in their ability to catalyze only certain 
chemical reactions; this high degree of specificity and strong 
catalytic activity are the most important functional properties of 
enzymes (Ref. 15).
    The Commission on Enzymes of the International Union of 
Biochemistry has devised a systematic strategy for naming enzymes. This 
system combines a naming system and a numbering system. For most 
enzymes, the systematic name is derived from the names of the 
substrate, product, and type of reaction. The systematic number is 
based on the class and subclasses to which the enzyme belongs. The 
systematic name of lysozyme is peptidoglycan N-acetylmuramoylhydrolase. 
Its systematic number is EC No. 3.2.1.17 and its Chemical Abstracts 
Service Registry Number (CAS Reg. No.) is 9001-63-2.
    Lysozyme was first discovered by A. Fleming, who identified 
lysozyme as an antibacterial enzyme present in nasal mucus membrane 
(Ref. 3). Subsequently, it was learned that the antibacterial activity 
of lysozyme occurs because of its ability to catalyze the hydrolysis of 
the structural polysaccharide peptidoglycan present in cell walls of 
certain bacteria (Ref. 2). Lysozyme activity has been shown to be 
present in bacteria, fungi, plants, and almost all animal tissues, with 
the highest levels found in secretions (including milk, mucus, saliva, 
and tears) and eggs. Lysozyme is believed to function in all of these 
organisms and tissues as an endogenous antimicrobial substance (Refs. 1 
and 2).
    Lysozyme was the first enzyme to have the details of its three-
dimensional structure published (Ref. 4), and it has become one of the 
best characterized of all enzymes, serving as an example for studies of 
enzyme mechanism and molecular evolution (Refs. 5 and 6 ). Lysozymes 
from various organisms are very similar to one another. Egg white 
lysozyme differs very little in structure, amino acid sequence and 
composition, catalytic mechanism, and substrate specificity from the 
enzyme found in human milk, saliva, mucus, and tears (Refs. 3 and 6).
    The petitioner provided two published scientific review articles 
(Refs. 1 and 2) that discuss the use of egg white lysozyme in cheese 
and other food. The petitioner also provided a generally available 
trade association bulletin (Ref. 7) that focuses on the use of egg 
white lysozyme for its technical effect of preventing late blowing in 
cheese. This bulletin describes the late blowing defect and how it 
arises, traditional chemical control measures (other than the use of 
lysozyme) to reduce the problem, and the increasing interest in using 
lysozyme as a replacement for traditional chemical control measures. In 
addition, the petitioner provided generally available information 
documenting that this intended use of the petitioned enzyme preparation 
has been approved in several countries, including Denmark, France, 
Germany, Italy, and Spain (Refs. 9 through 13).
    FDA considered the estimated dietary exposure to lysozyme for the 
proposed use in cheese (Refs. 16 and 17). Lysozyme accounts for 
approximately 3.5 percent of the total protein of domestic hen egg 
whites (Ref. 7). Whole eggs contain lysozyme at a level of 
approximately 3,300 parts per million (ppm). The petitioner reported 
that cheese manufactured using egg white lysozyme enzyme preparation 
contains a maximum of 400 ppm of lysozyme, or at least 8 times less 
than eggs on a weight basis. FDA has estimated a long-term mean intake 
of lysozyme to be 74 milligrams per person per day (mg/p/d) for 
consumers of eggs and 3.8 mg/p/d for consumers of cheese; the 
respective 90th percentile intakes are estimated to be 163 mg/p/day and 
8.1 mg/p/day. Egg whites from which lysozyme is extracted will be 
subsequently consumed in other food uses. Thus, there will be no long-
term net increase in lysozyme intake by the general population because 
egg whites without lysozyme will replace egg whites in current use that 
contain lysozyme (Ref. 16). On a per eating occasion basis, lysozyme 
intake for cheese consumers may be 16 mg on average, or 22 mg at the 
90th percentile level. For comparison, a per eating occasion lysozyme 
intake for egg consumers may be 264 mg on average, or 416 mg at the 
90th percentile level. Thus, lysozyme intake per eating occasion due to 
cheese consumption may constitute 5 to 6 percent of lysozyme intake due 
to egg consumption (Ref. 17).
    In general, issues relevant to a safety evaluation of proteins such 
as the enzyme component of an enzyme preparation are potential toxicity 
and allergenicity (Ref. 18). Proteins derived from egg whites do not 
raise toxicity concerns because egg whites have been safely consumed by 
humans as a source of food throughout recorded history without any 
reports of toxicity. However, proteins derived from egg whites do raise 
allergenicity concerns because, as with many common foods, there have 
been reports that consumption of egg whites can cause an allergic 
reaction in certain individuals, particularly children (Ref. 19). 
Therefore, FDA considered the question of whether the lysozyme 
component of egg whites is allergenic.
    In evaluating this question, FDA considered a report of an in vitro 
study of the binding of antibodies to specific egg proteins, where the 
antibodies were derived from the serum of patients known to be allergic 
to eggs (Ref. 20). This report suggests that lysozyme was an allergen 
for some individuals who became sensitive to egg whites. Although this 
study does not establish that ingestion of egg white lysozyme in cheese 
will actually cause a clinically

[[Page 12423]]

significant allergic reaction in such sensitive individuals, FDA is not 
aware of any data or information that would refute the study's 
inference that egg white lysozyme may be allergenic. Accordingly, FDA 
is proposing labeling, as discussed below, to alert the sensitive 
population to the presence of egg white lysozyme in cheese.
    A related question is whether egg white lysozyme, when present in 
cheese, is capable of inducing an allergenic response in susceptible 
individuals who have not previously consumed egg whites, e.g., because 
their customary diet excludes eggs. This question is no different than 
for any other food containing egg white when consumed by individuals 
with unknown susceptibility to eggs. The proposed label declaration 
would provide such individuals with the same protection as that 
provided by other egg-containing products with ingredient labeling. 
Thus, individuals who experience an allergic reaction to lysozyme-
containing cheese could identify egg white lysozyme as a possible cause 
of the reaction.

B. Enzyme Source, Manufacturing Methods, and Processing Aids

    Commercial preparations of lysozyme are derived from domestic hen 
egg whites using ion exchange methods and selective precipitation to 
isolate a highly purified protein fraction that contains mainly 
lysozyme but also may contain small amounts of other egg white 
proteins. Consistent with the agency's finding in its GRAS affirmation 
of microparticulated protein product (55 FR 6384, February 23, 1990), 
FDA finds that egg whites have been safely consumed by humans 
throughout recorded history and, therefore, are GRAS (Sec. 170.30(d)). 
The agency evaluated the methods used to isolate the enzyme lysozyme 
from egg whites. These methods are based on generally available and 
accepted principles of protein purification (Ref. 8). Such methods, if 
appropriately selected, do not ordinarily alter the chemical identity 
and characteristic properties of enzymes. Therefore, these methods do 
not materially change the quality, utility, functionality, or safety of 
enzymes. Moreover, the retention of the antibacterial activity that is 
characteristic of egg white lysozyme when egg white-derived lysozyme 
enzyme preparation is used in cheese evidences that lysozyme in the 
manufactured enzyme preparation remains unaltered from the lysozyme in 
egg whites. This is corroborative evidence of the fact that the methods 
used to isolate lysozyme from egg whites do not materially change the 
quality, utility, functionality or safety of the enzyme lysozyme.
     Enzyme preparations used in food processing are usually not 
chemically pure but contain, in addition to the enzyme component, 
materials that derive from the enzyme source. As mentioned above, egg 
white lysozyme enzyme preparation may contain small amounts of other 
egg white proteins. A related question is whether such proteins that 
may be present in the enzyme preparation are allergenic. Even if 
present, other source-derived proteins would not be a concern because 
the proposed label declaration for egg white lysozyme would alert 
individuals who are sensitive to egg whites to the possible presence of 
other proteins derived from egg whites.
    In addition to source-derived materials, enzyme preparations used 
in food processing usually contain materials that derive from the 
manufacturing methods used to generate the finished enzyme preparation. 
The egg white lysozyme enzyme preparation that is the subject of this 
document complies with the general requirements and additional 
requirements for enzyme preparations in the Food Chemicals Codex, 4th 
ed. (Ref. 14). The egg white lysozyme enzyme preparation that is the 
subject of this document may contain substances that are added to the 
enzyme preparation, such as preservatives, stabilizers or diluents, and 
trace amounts of processing aids that are used in its preparation. 
These substances must be acceptable for general use in foods (Refs. 14 
and 15).

C. Labeling as a Condition of Use

    Egg whites are known to be an allergenic food source, particularly 
in children (Ref. 19). There is a literature report (Ref. 20) 
indicating that lysozyme may in fact have been an allergen for some 
individuals who became sensitive to egg whites. Although the reported 
in vitro study does not establish that ingestion of egg white lysozyme 
in cheese will actually cause a clinically significant allergic 
reaction in such sensitive individuals, FDA is not aware of any data or 
information that would refute the study's inference that egg white 
lysozyme may be allergenic. Therefore, FDA concludes that there is 
insufficient information in the current record to determine whether the 
ingestion of egg white lysozyme elicits an allergenic response when 
consumed by individuals who are sensitive to egg whites. Accordingly, 
as discussed below, FDA is proposing labeling to alert such individuals 
to the presence of egg white lysozyme in cheese. Such labeling also 
would alert the sensitive population to the possible presence of 
source-derived proteins other than lysozyme in the enzyme preparation.
    Under section 409(c)(1) of the Federal Food, Drug, and Cosmetic Act 
(the act) (21 U.S.C. 348(c)(1)), FDA is authorized, in approving the 
use of a food additive, to list the conditions under which the additive 
may be safely used. These conditions may include any labeling 
requirements that the agency deems necessary to ensure the safe use of 
the additive. Similarly, under Sec. 184.1(b)(3) (21 CFR 184.1(b)(3)), 
in affirming a substance as GRAS, FDA is authorized to set forth the 
particular conditions of use, including labeling, under which there is 
general recognition among qualified experts that the use of the 
substance is safe. After careful review of the evidence on the use of 
egg white lysozyme enzyme preparation in preventing late blowing in 
cheese, FDA has tentatively concluded that such use is GRAS only when 
the conditions of its use include a declaration on the label or 
labeling of the presence of egg white lysozyme in both bulk and 
packaged food containing such treated cheese. Therefore, this tentative 
final rule (Sec. 184.1550(c)(1)) establishes that the declaration of 
egg white lysozyme enzyme preparation by the common or usual name ``egg 
white lysozyme'' is a condition of use required for GRAS status, so 
that consumers who are allergic to egg white products can be alerted to 
the presence of the egg white-derived enzyme in treated cheese.

D. Summary and Conclusions

    The petitioner provided published data and information relating to 
the identity of, characteristic properties of, and estimated dietary 
exposure to the enzyme component (Refs. 1 through 7). The source of the 
petitioned enzyme preparation, egg white, has been safely consumed by 
humans as a source of food protein throughout recorded history, and, 
therefore, is GRAS (Sec. 170.30(d)). The petitioner provided generally 
available information showing that the methods used for extracting 
lysozyme from the egg white source do not ordinarily alter the chemical 
identity and characteristic properties of enzymes (Ref. 8). Moreover, 
there is corroborating evidence that the extraction of egg white 
lysozyme does not change its chemical identity or characteristics 
because the antibacterial activity of egg white lysozyme is retained. 
FDA concludes that the methods used to manufacture egg white lysozyme 
enzyme preparation do not change the safety for food use of the enzyme 
lysozyme and that toxicological

[[Page 12424]]

studies are not necessary to establish the safety of lysozyme or other 
source-derived proteins that may remain in the manufactured enzyme 
preparation. FDA also concludes that there will be no net increase in 
dietary exposure of the general population to the commonly consumed 
enzyme lysozyme due to the proposed use in cheese because lysozyme will 
simply be transferred from eggs to cheese (Ref. 16).
    The petitioner also provided generally available and accepted 
information relating to processing aids used in the manufacture of the 
enzyme preparation and generally available and accepted specifications 
for food grade enzyme preparations (Ref. 14). FDA concludes that 
substances added to the egg white lysozyme enzyme preparation or 
potential residues of processing aids used in the manufacturing process 
do not present a basis for concern about the safety of the egg white 
lysozyme enzyme preparation.
    The petitioner provided published scientific review articles (Refs. 
1 and 2) and a generally available trade bulletin (Ref. 7) that discuss 
the use of the egg white lysozyme enzyme preparation in cheese and 
other food, including its use for the intended effect of preventing 
late blowing of cheese contaminated with C. tyrobutyricum. The 
petitioner also provided generally available information documenting 
that this intended use of lysozyme has been approved in several 
European countries (Refs. 9 through 13). FDA concludes that generally 
available and accepted data and information establish that lysozyme 
will achieve the intended technical effect of preventing late blowing 
in cheese contaminated with C. tyrobutyricum.
    Finally, information in the petition and otherwise available to FDA 
raises the question of whether the lysozyme component of egg whites is 
allergenic. FDA is proposing labeling to alert individuals who may be 
sensitive to egg whites to the presence of egg white lysozyme in 
cheese, including the possible presence of other source-derived 
proteins that may be present in the enzyme preparation.

IV. Comments

    FDA received two comments in response to the filing notice. One 
comment expressed agreement that lysozyme is GRAS for use in preventing 
late blowing in cheese and supported the affirmation of GRAS status by 
the agency.
    One comment stated that use of lysozyme as a food preservative may 
lead to selection of lysozyme-resistant strains of the bacterial food 
poisoning agents Listeria monocytogenes and C. botulinum, rendering one 
of the body's main defense mechanisms useless against resistant 
strains. The comment likened the potential selection of lysozyme-
resistant strains of bacteria to the selection of penicillin-resistant 
bacteria as a result of its widespread use. The comment pointed out 
that the body could not readily substitute the lysozyme naturally 
present in secretions such as tears and saliva for another 
antimicrobial.
    The mechanism of action of lysozyme involves hydrolysis of the 
structural peptidoglycan present in cell walls of susceptible bacteria. 
Therefore, development of resistance to lysozyme would require that a 
bacterium develop a variant of peptidoglycan that is resistant to the 
action of lysozyme. Development of such a variant peptidoglycan is, in 
principle, possible. However, as already discussed, lysozyme activity 
has been shown to be present in bacteria, fungi, plants, and almost all 
animal tissues. If such relative ubiquity has not resulted in the 
clinically significant selection of lysozyme-resistant bacteria to 
date, the use of lysozyme in those cheeses that are susceptible to late 
blowing is unlikely to favor selection of lysozyme-resistant bacteria 
and adversely affect the public health. Moreover, FDA is not 
considering lysozyme for use as a widespread food preservative. Rather, 
FDA is considering the narrow question of whether the use of lysozyme 
in preventing late blowing in cheese is generally recognized as safe. 
FDA disagrees that this limited use in cheese is analogous to the 
widespread use of antibiotics such as penicillin and the subsequent# 
selection of antibiotic-resistant bacterial strains. Therefore, FDA 
concludes that the use of lysozyme in preventing late blowing in cheese 
does not raise concerns about the selection of lysozyme-resistant 
strains of L. monocytogenes or C. botulinum.

V. Specifications

    The agency finds that, because the potential impurities in the egg 
white lysozyme preparation that may originate from the source or 
manufacturing process do not raise any basis for concern about the safe 
use of the preparation, the general requirements and additional 
requirements for enzyme preparations in the monograph on Enzyme 
Preparations in the Food Chemicals Codex, 4th ed. (1996), which are 
being incorporated by reference in accordance with 5 U.S.C. 552(a) and 
1 CFR part 51, are adequate as minimum criteria for food-grade egg 
white lysozyme enzyme preparation. Lysozyme assay can be performed 
using a method entitled ``Lysozyme hydrochloride, Microbiological 
Determination,'' which is included in the petition (Ref. 21) or by 
using any appropriate validated method.

VI. Conclusions

    The agency has evaluated all available information and finds, based 
upon the published information about the manufacturing methods used in 
the preparation of egg white lysozyme enzyme preparation, and published 
data and information about the identity and characteristic properties 
of egg white lysozyme, that the enzyme component of egg white lysozyme 
enzyme preparation is unaltered from the lysozyme found in the commonly 
consumed food, eggs. The agency also finds, based upon generally 
available and accepted information, that when the preparation is 
manufactured in accordance with Sec. 184.1550(c), the source, egg 
whites, and the manufacturing process will not introduce impurities 
into the preparation that may render its use unsafe. Further, the 
agency finds, based upon published information, that egg white lysozyme 
enzyme preparation will achieve its intended technical effect of 
preventing late blowing in cheese contaminated with C. tyrobutyricum. 
Therefore, the agency tentatively concludes, based upon the evaluation 
of published data and information, corroborated by unpublished data and 
information, that the egg white lysozyme enzyme preparation described 
in the regulation set out below is GRAS for use by the general 
population in preventing late blowing in cheese.
    To give interested persons an opportunity to comment on the 
proposed label declaration that is a condition of use required for GRAS 
status, FDA is issuing this tentative final rule under 21 CFR 
10.40(f)(6). FDA will review any comments that are relevant to this 
condition of use and that are received within the 75 day comment period 
and will respond accordingly to these comments in the Federal Register.

VII. Environmental Considerations

    The agency has carefully considered the potential environmental 
effects of this action. FDA has concluded that the action will not have 
a significant impact on the human environment, and that an 
environmental impact statement is not required. The agency's finding of 
no significant impact and the evidence supporting that finding, 
contained in an environmental assessment, may be seen in the Dockets 
Management Branch

[[Page 12425]]

(address above) between 9 a.m. and 4 p.m., Monday through Friday.

VIII. Analysis of Economic Impacts

A. Benefit-Cost Analysis

    FDA has examined the impacts of this tentative final rule under 
Executive Order 12866. Executive Order 12866 directs Federal agencies 
to assess the costs and benefits of available regulatory alternatives, 
and, when regulation is necessary, to select regulatory approaches that 
maximize net benefits (including potential economic, environmental, 
public health and safety effects; distributive impacts; and equity). 
According to Executive Order 12866, a regulatory action is 
``significant'' if it meets any one of a number of specified 
conditions, including having an annual effect on the economy of $100 
million, adversely affecting in a material way a sector of the economy, 
competition, or jobs, or if it raises novel legal or policy issues. FDA 
finds that this tentative final rule is not a significant regulatory 
action, as defined by Executive Order 12866. In addition, it has been 
determined that this final rule is not a major rule for the purpose of 
congressional review.
    The primary benefit of this action is to remove uncertainty about 
the regulatory status of the petitioned substance. FDA is tentatively 
affirming the GRAS status of egg white lysozyme in cheese only when the 
ingredient statement of the bulk and packaged food that contains the 
cheese includes the common or usual name of the substance, i.e., ``egg 
white lysozyme.'' The labeling requirement will add a small cost to the 
future use of the petitioned substance, and therefore, is not a 
significant action under the Executive Order 12866.
    FDA has examined the impacts of this tentative final rule under the 
Unfunded Mandates Reform Act of 1995 (UMRA) (Pub. L. 104-4). A written 
statement under section 202(a) of the UMRA is not required for this 
rule because the rule does not impose a mandate that results in an 
expenditure of $100 million or more by State, local, and tribal 
governments in the aggregate, or by the private sector, in any 1 year.

B. Regulatory Flexibility Act

    FDA has evaluated this tentative final rule under the Regulatory 
Flexibility Act. The Regulatory Flexibility Act (5 U.S.C. 601-612) 
requires Federal agencies to consider alternatives that would minimize 
the economic impact of their regulations on small entities.
    FDA believes that this tentative final rule is not likely to have a 
significant economic impact on a substantial number of small entities. 
However, the agency seeks comment on this tentative conclusion. First, 
FDA is tentatively affirming the GRAS status of egg white lysozyme in 
cheese only when the ingredient statement of the bulk and packaged food 
that contains the cheese includes the common or usual name of the 
substance, i.e., ``egg white lysozyme.'' This labeling requirement will 
impose only minimal costs to the future use of the petitioned 
substance. Second, FDA has information that the petitioner does not 
currently sell egg white lysozyme in the United States (Refs. 22 and 
23). Moreover, FDA is not aware of any manufacture or use of cheese 
containing egg white lysozyme in the United States. If no small 
entities are currently manufacturing or using cheese containing egg 
white lysozyme, the proposed labeling requirements would not impose any 
cost to small entities. However, because FDA does not have any 
information on whether other entities in the United States are 
manufacturing or using cheese containing egg white lysozyme, FDA is 
unable to conclude, in this tentative final rule, that there will be no 
significant economic impact on a substantial number of small entities. 
Therefore, the agency seeks comment on the manufacture or use, by any 
small entity, of cheese containing egg white lysozyme. In its final 
rule, the agency will, based on any relevant comments received, 
determine whether there is a significant economic impact on a 
substantial number of small entities.

IX. References

    The following references have been placed on display in the Dockets 
Management Branch (address above) and may be seen by interested persons 
between 9 a.m. and 4 p.m., Monday through Friday.
    1. Carini, S., G. Mucchetti, and E. Neviani, ``Lysozyme: 
Activity Against Clostridia and Use in Cheese Production--A 
Review,'' Microbiologie Aliments Nutrition, 3:299-320, 1985.
    2. Procter, V. A., and F. E. Cunningham, ``The Chemistry of 
Lysozyme and Its Use as a Food Preservative and a Pharmaceutical,'' 
CRC Critical Reviews in Food Science and Nutrition, 26:359-395, 
1988.
    3. Fleming, A., ``On a Remarkable Bacteriolytic Element Found in 
Tissues and Secretions,'' Proceedings of the Royal Society, 93:306-
317, 1922.
    4. Blake, C. C. F., D. F. Koenig, G. A. Mair, A. C. T. North, D. 
C. Phillips, and V. R. Sarma, ``Structure of Hen Egg-White 
Lysozyme,'' Nature, 206:757-783, 1965.
    5. Kuhara, S., E. Ezaki, T. Fukamizo, and K. Hayashi, 
``Estimation of the Free Energy Change of Substrate Binding in 
Lysozyme-Catalyzed Reactions,'' Journal of Biochemistry, 92:121-127, 
1982.
    6. Malcolm, B., K. Wilson, B. Matthews, J. Kirsch, and A. 
Wilson, ``Ancestral Lysozymes Reconstructed, Neutrality Tested, and 
Thermostability Linked to Hydrocarbon Packing,'' Nature, 345:86-89, 
1990.
    7. ``The Use of Lysozyme in the Prevention of Late Blowing in 
Cheese,'' Bulletin of the International Dairy Foundation, No. 216, 
1987.
    8. Chaplin, M. F., and C. Bucke, Enzyme Technology, Cambridge 
University Press, New York, NY, pp. 66-72, 1990.
    9. Denmark Ministry of Agriculture Circular on Approved Enzymes 
for Cheese, Journal nr. 10.1.5-18/83, January 13, 1984.
    10. French Republic Ministry of Agriculture Service Note 5236, 
April 21, 1987.
    11. Federal Republic of Germany, Bundesgesetzblatt Nr. 56, pp. 
2103-2107, November 22, 1985.
    12. Official Gazette of Italian Republic, General Series n. 23, 
October 4, 1986.
    13. Spanish Ministry of Health and Consumption, General 
Direction of Public Health, General Sanitary Register, File n. 8826, 
June 20, 1983.
    14. Monograph on ``Enzyme Preparations,'' Food Chemicals Codex, 
National Academy Press, Washington, DC, 4th ed., pp. 133-134, 1996.
    15. Pariza, M. W. and E. M. Foster, ``Determining the Safety of 
Enzymes Used in Food Processing,'' Journal of Food Protection, 
46:453-468, 1983.
    16. Memorandum dated March 20, 1990, from Food and Color 
Additives Review Section, FDA, to Direct Additives Branch, FDA, 
``Use of Lysozyme to Prevent the `Late Blowing' of Cheese.''
    17. Memorandum dated August 5, 1996, from Chemistry Review 
Branch, FDA, to Biotechnology Policy Branch, FDA.
    18. Kessler, D. A., M. R. Taylor, J. H. Maryanski, E. L. Flamm, 
and L. S. Kahl, ``The Safety of Foods Developed by Biotechnology,'' 
Science, 256:1747-1749 and 1832, 1992.
    19. Crespo, J. F., C. Pasqual, A. Ferrer, A. W. Burke, J. M. 
Diaz Pena, and M. M. Esteban, ``Egg White-specific IgE Level as a 
Tolerance Marker in the Follow-up of Egg Allergy,'' Allergy 
Proceedings, 15:73-76, 1994.
    20. Anet, J., J. F. Back, R. S. Baker, D. Barnett, R. W. Burley, 
and M. E. H. Howden, ``Allergens in the White and Yolk of Hen's Egg; 
a Study of IgE Binding by Egg Proteins,'' International Archives of 
Allergy and Applied Immunology, 77:364-371, 1985.
    21. Lysozyme Hydrochloride, Microbiological Determination.
    22. Letter dated November 25, 1996, from John B. Dubeck, Keller 
and Heckman, to Linda Kahl, FDA.
    23. Letter dated February 28, 1997, from John F. Foley, Keller 
and Heckman, to Linda Kahl, FDA.

List of Subjects in 21 CFR Part 184

    Food ingredients, Incorporation by reference.
    Therefore, under the Federal Food, Drug, and Cosmetic Act and under

[[Page 12426]]

authority delegated to the Commissioner of Food and Drugs and 
redelegated to the Director, Center for Food Safety and Applied 
Nutrition, it is proposed that 21 CFR part 184 be amended as follows:

PART 184--DIRECT FOOD SUBSTANCES AFFIRMED AS GENERALLY RECOGNIZED 
AS SAFE

    1. The authority citation for 21 CFR part 184 continues to read as 
follows:

    Authority: 21 U.S.C. 321, 342, 348, 371.

    2. Section 184.1550 is added to subpart B to read as follows:

Sec. 184.1550  Egg white lysozyme.

    (a) Egg white lysozyme (CAS Reg. No. 9001-63-2) is the enzyme 
peptidoglycan N-acetylmuramoylhydrolase (EC No. 3.2.1.17) obtained by 
extraction from egg whites. The enzyme catalyzes the hydrolysis of 
peptidoglycan in the cell walls of certain bacteria including 
Clostridium tyrobutyricum.
    (b) The ingredient meets the general requirements and additional 
requirements for enzyme preparations in the monograph on Enzyme 
Preparations in the Food Chemicals Codex, 4th ed. (1996), which is 
incorporated by reference in accordance with 5 U.S.C. 552(a) and 1 CFR 
part 51. Copies are available from the National Academy Press, 2101 
Constitution Ave. NW., Washington, DC 20418, and may be examined at the 
Center for Food Safety and Applied Nutrition's Library, 200 C St. SW., 
rm. 3321, Washington DC, or at the Office of the Federal Register, 800 
North Capitol St. NW., suite 700, Washington, DC.
    (c)(l) The ingredient is used in cheeses, as defined in 
Sec. 170.3(n)(5) of this chapter, in accordance with Sec. 184.1(b)(3) 
at levels not to exceed current good manufacturing practice.
    (2) The affirmation of the use of this ingredient as generally 
recognized as safe (GRAS) as a direct human food ingredient is based 
upon the following conditions of use:
    (i) The ingredient is used as an enzyme as defined in 
Sec. 170.3(o)(9) of this chapter.
    (ii) Current good manufacturing practice utilizes a level of the 
ingredient sufficient to prevent the late blowing of cheeses caused by 
the bacterium Clostridium tyrobutyricum during cheese production.
    (iii) The ingredient statement for both bulk and packaged food that 
contains cheese manufactured using egg white lysozyme shall include the 
common or usual name ``egg white lysozyme'' to identify the source of 
the protein.

    Dated: March 3, 1998.
L. Robert Lake,
Director, Office of Policy, Planning and Strategic Initiatives, Center 
for Food Safety and Applied Nutrition.
[FR Doc. 98-6571 Filed 3-12-98; 8:45 am]
BILLING CODE 4160-01-F