[From the U.S. Government Printing Office, www.gpo.gov]






                                                                   TASK  I.A.1
















                                     POTENTIAL  DETOXIFICATION
                                               OF
                                     SHEBOYGAN  HARBOR PCB'S
























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                                             POTENTIAL DETOXIFICATION
                                                         OF
                                              SHEBOYGAN HARBOR PCBS






                                                          COASTAL ZONE
                                                       INFORMATION CENTER






                                        This study was funded in part with
                                        financial assistance provided by
                                        the State of Wisconsin, Division of
                                        Energy and Intergovernmental
                                        Relations, Coastal Management
                                        Program through a grant under the
                                        Coastal Zone Management Improvement
                                        Act of 1980, as amended,
                                        administered by the Office of Ocean
                                        and Coastal Resource Management,
                                        National oceanic and Atmospheric
                                        Administration.

                                        The development of this project was
                                        under the supervision 'of William C.
                                        Sonzogni, University of Wisconsin,
                       "k,
         QQN3tP,,1fPEP V E              Madison, Wisconsin.
                 @ARJM


                                                 US Department of Commerce
                                             NOAA Coastal Services Ccnter Library
                                                  2234 South Hobson Avenue
                                                  Charleston, SC 29405-2413




















                                 EXECUTIVE SUMMARY






                            POTENTIAL DETOXIFICATION OF
                                SHEBOYGAN HARBOR PCBS



















                                                        William C. Sonzogni
                                                     Principal Investigator











               over the last two years research has been conducted on
          Wisconsin's Sheboygan River, a tributary to Lake Michigan located
          north of the City of Milwaukee. The Sheboygan River was studied

          because it is contaminated by polychlorinated biphenyls (PCBs), so

          much that it has been declared a Superfund Site by the U.S.

          Environmental Protection Agency and a Great Lakes Area of Concern

          by the U.S./Canadian International Joint Commission. The river is

          also a source of PCBs to Lake Michigan.      Further, the PCBs, at

          least some of more toxic PCB compounds, are a potential health risk

          to inhabitants of  the river or users of the river.     Thus, this

          research was designed to provide important information on a major

          environmental problem affecting the coastal zone of the Great

          Lakes.


               The focus of the research was to determine whether PCB

          dechlorination is occurring in the river's sediments.           High

          resolution gas chromatographic analyses were made on sediment,

          water, and fish collected from the river and harbor. In addition,

          several sail samples from a confined disposal facility holding

          dredged sediment were also analyzed. Laboratory experiments were

          also conducted to try and replicate some of the processes that

          occur in the natural environment.         information   from other

          investigators and the scientific literature was used when possible

          to supplement findings.

               Overall, the research was successful. All of the objectives

          were met. major findings of the research are summarized below.








               1. Data collected on PCBs in sediments indicate the
               distribution of PCB compounds (congeners) has changed relative

               to the distribution of PCBs that originally polluted the

               river.

               2. It appears, based on all the available evidence, that the

               changes in the PCB distribution are the result of

               dechlorination of the PCBs by anaerobic microorganisms in the

               sediment.

               3. Dechlorination was observed mainly in those section of the

               river with accumulation of muck (sedimentation areas) and

               where total PCB concentrations were high.

               4. The fact that many of the higher chlorinated PCB compounds

               (congeners) are being dechlorinated indicates that some

               natural detoxification is occurring, as the more highly

               chlorinated PCB compounds are generally considered to be more

               toxic.

               5. The results suggest that it may be possible to modify

               natural processes to speed up the dechlorination as a

               bioremediation measure. Complete degradation of the PCBs may

               even be possible.

               6. PCB concentrations in the harbor (about 20 km downstream

               from the source) were much lower than the source area and the

               distribution of PCB compounds did not show evidence of

               degradation.

               7. PCB concentrations in the water were much lower than in

               sediment, but concentrations increased downstream from the

               source. The distribution of PCB compounds varied between the








              2. One thesis was published as a result of this research, and
              two research papers are written so far and will be submitted

              to refereed scientific journals.
              3.   Several presentations have been made on the project at

              scientific meetings, including an international workshop on

              biological remediation of contaminated sediments sponsored by

              the U.S. Environmental Protection Agency. A summary of the

              presentation is to be published in the workshop proceedings.

              4. The multidimensional gas chromatograph, purchased for the

              Wisconsin Laboratory.of Hygiene partly through this project,

              will be a continuing resource for future PCB and other complex

              organic analysis. As a result of demonstrating the ability of

              this machine to measure "toxic" PCB compounds in this project,

              Fort Howar d Paper Company has decided to purchase one of these

              machines to help them in their efforts to free the lower Fox

              River and Green Bay from PCB pollution.

              5. The Wisconsin Department of Natural Resources has become

              increasingly concerned about the presence of certain PCB

              compounds (congeners) , and methods to analyze these compounds

              refined during this project will be used in the future by the

              State Laboratory of Hygiene in -doing analyses for the

              Department.

              6. Information on PCBs developed in this project will be

              useful to the Green Bay Mass Balance Study now ongoing on the

              Fox River and Green Bay.

              T. Results of this study will be useful to the current

              Superfund investigation and remedial action plan. In fact,








               dissolved and particulate PCBS.
               8. The types of PCBs found in fish from the Sheboygan River

               were different from the PCB pattern found in contaminated

               sediments.

               9. While experiments to induce anaerobic dechlorination of

               PCBs using bacteria extracted from Sheboygan River sediments

               were negative, further experiments would likely produce

               positive results.- Based on recent laboratory work by others
               (with different river sediments), it is clear dechlorination

               can be demonstrated in the laboratory although results are

               highly variable. Key factors controlling dechlorination are

               not fully understood.

               10. The multidimensional (heart cutting) gas chro*matogr aph is

               an effective tool for resolving PCB compounds (congeners) that

               otherwise can not be measured.

               11. Some of the "toxic" PCBs (having dioxin-like effects),

               that can't be resolved by conventional high resolution

               techniques, were quantified using the multidimensional gas

               chromatograph. Concentrations were always very low and there

               were indications that dechlorination of several of these

               compounds was occurring.

               This research has led to a number of benefits and products

         other than the research findings. The most important of these are

         listed below.

               1. Training was provided to students and technicians.         A

               Master of Science degree in Water Chemistry was received by

               one student working on  the project.








               this -study probably had an influence or helped stimulate

               current ef f orts to bioremediate dredged Sheboygan sediment

               through microbial processes.
               8. The results of this study will be Ugeftl to other coastal

               sites with in-place pollution due to PCBs.             Natural

               dechlorination may be a useful remedial strategy      in some

               situations.

               9. The Assessment and Remediation of Contaminated Sediments

               (ARCS) Program, set up under the 1987 amendments to the U.S.

               Clean Water Act, has set the Sheboygan River as one of its

               pilot study areas. Data from this study will be essential for

               this Program, and information has already been given to

               program officials.

               10. Last but not least, findings from this study will benefit

               the living near or using the S heboygan River. Knowing that

               some of the more highly chlorinated, and thus potentially most

               toxic, can be dechlorinated provides hope that the PCB problem

               can eventually be solved.

               The hard work, support and cooperation of many individuals

         made this study possible.     co-principal investigators included

         Professors David Armstrong and Anders Andren, and Dr. John Konrad

         of  the Wisconsin Department of Natural Resources' Bureau of

         Research. ., Laura Maack Brondyk served as Assistant Researcher

         throughout most of the proDect, and played a major role in

         initiating the project.     Margaret David served as the student

         researcher for the project, and the core findings of the project

         are described in her Master of Science       dissertation.    Robert








         Lawrence served as an Assistant researcher during the second half

         of the study. Many staff at the Wisconsin Laboratory of Hygiene

         contributed their expertise, particularly to the analytical
         chemistry part of the work. Notable contributors were David

         Degenhardt, Thomas Gibson, Carol Buelow William Krick and Thomas

         Dunnick.    Professor Ronald Schell and Terry Kurzynski provided

         bacteriological expertise.

               Finally, the generous financial support of the Wisconsin

         Coastal Management Program and Wisconsin Sea Grant Program are'

         gratefully acknowledged. In addition to the fiscal support, the

         administrative support and general encouragement given by the

         staffs of these organizations was really appreciated.              The

         Wisconsin State Laboratory of Hygiene and the Water Chemistry

         Program of the University of Wisconsin, as well as the Wisconsin

         Department of Natural Resources also provided support for this

         project.




















                                    ATTACHMENTS



         1. PCB Congener  istribution in Sheboygan River Sediment Fish and
              Water (M.S. Thesis by Margaret David).

         2. Concentrations of Toxic Polychlorinated Biphenyl Congeners in
              Sheboygan River (USA) Sediments (paper submitted to the
              Bulletin of. Environmental Contamination and Toxicology).

         3. PCB Dechlorination in the Sheboygan River, Wisconsin (summary
              to be published in proceedings of a workshop on biological
              remediation of contaminated sediments).

         4. Summary of Laboratory Experiments to Dechlorinate PCBs using
              Bacteria Extracted from Sheboygan River Sediments.

         5. An Evaluation of the Potential for PCB Dechlorination in
              Confined Disposal Facilities.




























                            PCB CONGENER DISTRIBUTION IN
                      SHEBOYGAN RIVER SEDIMENT, FISH AND WATER
















                                Margaret Meigs David



                  A thesis submitted in partial fulfillment of the
                          requirements for the degree of




                                  Master of Science
                               (Water Chemistry Program,
                  Department of Civil and Environmental Engineering)


                                   March 7, 1990













                                     Acknowledgements



                  I would like to thank my parents for their undying

               support and encouragement. They have been great parents and

               fantastic friends. Without my friend David, this project

               would be an abysmal skeleton of what it now is. David's

               ideas and criticisms were excellent and his drive to truly

               understand science, inspiring. I am very glad that my

               younger brother and I have been in graduate school at the

               same time. Andrew has been a source of humor and hope, as

               well as an amazingly fluid mechanics and differential

               equations tutor. Insightful comments have come from Peter

               who is omniscient and of course, my older brother. He

               constantly piques my curiosity about the world and desire to

               learn more.

                   I would especially like to extend my gratitude to all of

               those in the State Laboratory of Hygiene who worked with me.

               Carol Buelow and Tom Gibson have been extraordinarily

               helpful. Carol has saved me many a time from complete and

               total frustration and has been a mentor to me. Dave

               Degenhardt has made my life much simpler and for him I hope

               that National Dave Degenhardt day will become a state

               holiday with big band music and ice cream.

                  I would like to thank Bill Sonzogni for even considering

               me as a research assistant and then for getting involved in

               the dirty work of the science--sampling oozing, cold, wet











               river sediment.   Laura Maack and Debbie DeLuca were great

               colleagues with whom one could discuss detection limits and

               decaffeinated coffee. I would also like to thank Dr.

               Armstrong who inspite of hectic schedules has always made

               time to discuss this project and provide very thoughtful

               criticism.   And lastly, I would like to thank Dr. Andren

               whose spunky interesting lectures got me involved in water

               chemistry in the first place.














                                         TABLE OF CONTENTS

                 Chapter 1 Introduction  ..................................    1

                 Chapter 2 Objectives   ....................................   4

                 Chapter 3 Literature Review of Aerobic and Anaerobic
                            Bacteria  ......................................   6

                      3.1. Aerobic Degradation of PCBs     ..................  6
                      3.1.1. Introduction    ...............................   6
                      3.1.2. Enzyme Systems    .............................   7
                      3.1.3. Aerobic Microbial Degradation Studies
                               of PCBs  .................................... 10

                      3.2. Anaerobic Degradation of PCBs     ............... 15
                      3.2.1.   Introduction  ..............................  15
                      3.2.2.   Anaerobic Degradation of Structurally
                               Similar Compounds  .........................  17
                      3.2.3.   Anaerobic Microbial Degradation
                               Studies of PCBs  ...........................  21

                 Chapter 4 Site Description and Historical
                            Use of PCBs  ..................................  30

                 Chapter 5 Sample Collection,
                            Materials and Methods  ........................  38

                      5.1. Sample Collection    ...........................  38
                      5.1.1. Sediment   ..................................   38
                      5.1.2. Fish   ......................................   41
                      5.1.3. Water   .....................................   42


                      5.2. Materials   ...................................   43
                      5.2.1.   Solvents  ..................................  43
                      5.2.2.   Chromatographic Adsorbents   ................ 43
                      5.2.3.   Standards .................................   43
                      5.2.4.   Other Chemicals and Materials   ............. 44
                      5.2.5.   Equipment .................................   45
                      5.2.6.   Glassware .................................   45

                      5.3. Extraction and Cleanup Methods     .............. 45
                      5.3.1. Sediment   ..................................   45
                      5.3.2. Fish   ......................................   49
                      5.3.3.   Water  .....................................  52


                      5.4. Gas Chromatography    ..........................  57

                      5.5. Grain Size Analysis of Sediment     ............. 59














                 Chapter 6 Results    ......................................    64

                       6.1. Grain Size     ..................................   64

                       6.2. Sediment Samples from the Sheboygan
                       River .......................................     65
                       6.2.1.   Total Concentrations of PCBs in Sediment..65
                       6.2.2.   Homolog Distribution Patterns in Sediment.71
                       6.2.3.   Most Prominent Congeners in Sediment     ...... 76
                       6.2.4.   Accuracy, Precision and Confirmatory
                                Analysis for Sediment Samples     ............. 81

                       6.3. Sediment Samples from the Harbor       ............ 83
                       6.3.1.   Total PCB Concentrations
                                in Harbor Sediments    .......................  83
                       6.3.2.   Homolog Distribution in Harbor Sediment       ... 83
                       6.3.3.   Most Prominent Congeners
                                in Harbor Sediment    ........................  86
                       6.3.4.   Precision and Accuracy for Harbor Samples.88

                       6.4. Water Samples from the Sheboygan River       ...... 88
                       6.4.1.   Total PCB Concentrations in Water      ......... 88
                       6.4.2.   PCB Homolog Distribution in Water      ......... 90
                       6.4.3.   Most Prominent Congeners in Water      ......... 92
                       6.4.4.   Accuracy, Precision, and Confirmatory
                                Analysis for Water    ........................  93

                       6.5 Fish   .........................................     95
                       6.5.1. Total PCB Concentrations in Fish       .......... 95
                       6.5.2. Homolog PCB Distribution in Fish       .......... 95
                       6.5.3. Most Prominent Congeners in Fish       .......... 99

                 Chapter 7 Discussion of Results     .......................   101

                       7.1. Discussion of Sediment Results      .............. 101
                       7.1.1. Grain Size Correlations with
                                PCB Concentration   ........................   101
                       7.1.2. Total PCE Concentrations in Sediment       ..... 102

                       7.2. Discussion of Possible Mechanisms causing
                             the Congener Distribution in Sediment       ...... 105
                       7.2.1. Physical Processes Affecting the
                                Distribution of PcBs in Sediment     ......... 106
                       7.2.2. Biological Processes Affecting the
                                Distribution of PCBs in the Sediment     ..... 112


                       7.3. Discussion of Water Results     .................  123
                       7.3.1. Total PCB Concentrations in Water       ........ 123
                       7.3.2. Homolog and Congener Distribution
                                in water  .................................    125















                       7.4. Discussion of Fish Results     .................  126
                       7.4.1. Total PCB Concentrations in Fish       ......... 126
                       7.4.2. Homolog Patterns in Fish     .................  131
                       7.4.3. Most Prominent Congeners in Fish       ......... 132

                 Chapter 8   Conclusions   ................................   140

                       8.1.  Sediment  ...................................    140
                       8.2.  Water  ......................................    142
                       8.3.  Fish  .......................................    144
                       8.4.  Concluding Remarks    .........................  145

                 Bibliography   ..........................................    147

                 Appendices
                       Appendix A Limits of Detection
                       Appendix B PCB Congener Results















                  List of Tables

                  Chapter 3

                  Table 3-1   Homolog Distribution for Pattern AB and B         ...... 25
                  Table 3-2   Energy Derived from the Oxidation of
                              Glucose to Carbon Dioxide and Water
                              using various Oxidants     .........................    26

                  Chapter 4

                  Table 4-1   PCB Mixtures Used at Tecumseh Products Site         .... 36

                  Chapter 5

                  Table 5-1   operating Conditions for
                              the Gas Chromatograph     ..........................    58

                  Chapter 6

                  Table  6-1  Results of Sieved Sediment Samples       .............  66
                  Table  6-2  Total PCB Concentrations in River Sediment         ..... 69
                  Table  6-3  Most Prominent congeners in sediment        ........... 78
                  Table  6-4  Percent Recovery from Spiked Sediment        .......... 81
                  Table  6-5  Duplicate Sediment Samples      .....................   82
                  Table  6-6  Total PCB Concentrations in Harbor Sediment         .... 84
                  Table  6-7  Most Prominent Congeners
                              found in Harbor Samples     ........................    86
                  Table  6-8  Total PCB concentrations in Water      ..............   89
                  Table  6-9  Most Prominent Congeners in Water      ..............   93
                  Table  6-10 Total PCB Concentrations in Fish       ..............   96
                  Table  6-11 Most Prominent Congeners in Fish       ..............   99

                  Chapter 7
                  Table 7-1   PCB Concentrations in Sheboygan
                              River Sediment   ................................     103
                  Table  7-2  Changes in Sheboygan River Sediment
                              Relative to Aroclors (Sediment Samples
                              with concentrations greater than 50 ppm)        ...... 119
                  Table  7-3  Comparison of Congeners Predicted by
                              Brown et al. based on Anaerobic Degradation
                              to Congeners Observed in Study      ................  120
                  Table  7-4  Total PCB Concentrations in Water      .............  124
                  Table  7-5  Total PCB Concentrations in Fish
                              from the Sheboygan River     ......................   127
                  Table  7-6  A Comparison of Most Prominent
                              congeners in Fish    .............................    133
                  Table  7-7  Clearance Rates for Specific Congeners        ........ 136
                  Table  7-8  BCF Values Reported in the Literature        ......... 140












                 List of Figures

                 Chapter 3

                 Figure 3-1 Major Aerobic Degradation Pathway for PCBs        ..... 9
                 Figure 3-2 Anaerobic Degradation of Chlorobenzoates         ...... 19

                 Chapter 4

                 Figure 4-1 The Sheboygan River and Tributaries        ........... 31

                 Chapter 5

                 Figure 5-1 Map of Sample Sites      ...........................  39
                 Figure 5-2 Analysis of PCBs in Sediment       .................. 47
                 Figure 5-3   Analysis of PCBs in Fish   ......................   51
                 Figure 5-4   Analysis of PCBs in Water    .....................  53

                 Chapter 6

                 Figure 6-1   Map of Sheboygan River illustrating Average
                              Sediment Core   .................................   67
                 Figure 6-2   Homolog Distribution in    Sediment
                              (Concentration >50 ppm     Sites B, C, D, E)    ..... 73
                 Figure 6-3   Homolog Distribution in    Sediment
                              (Concentration >50 ppm.    Site T)  ............... 74
                 Figure 6-4   Homolog Distribution in    Sediment
                              (Concentration <50 ppm,    Sites B,C,D,E)   ........ 75
                 Figure 6-5   Homoloa Distribution in    Sediment
                              (Concentration <50 ppm,    Site T)  ............... 77
                 Figure  6-.6 Most Prominent Congeners in Sediment
                            . (Sites B,C,D,E)   ...............................   79
                 Figure  6-7  Most Prominent Congeners in Sediment
                              (Site T)  ......................................    80
                 Figure  6-8  Homolog Distribution in Harbor Sediment      ....... 85
                 Figure  6-9  Most Prominent Congeners in Harbor Sediment       ... 87
                 Figure  6-10 Homolog Distribution in Water      ................ 91
                 Figure  6-11 Most Prominent Congeners in Water      ............ 94
                 Figure  6-12 Homolog Distribution in Fish      ................. 97
                 Figure  6-13 Homolog Distribution in Fish and Sediments        ... 98
                 Figure  6-14 Most Prominent Congeners in Fish      ............ 100



                 Chapter 7

                 Figure 7-1 Ortho vs Meta/Para Congeners in Sediment        ..... 115
                 Figure 7-2 Ortho vs Meta/Para Congeners in
                              Spiked Sediment .....................   I ......... 117
                 Figure 7-3   Correlation between Fish Fat
                              and PCB Concentration   ........................   129













               Figure 7-4 Correlation between Packed
                          and Capillary Column Results ................. 130
               Figure 7-5 Predicted BCF from Mackay
                          versus Calculated BCF from Field data ........ 138















                                   Chapter I introduction

                    Polychlorinated biphenyls are a class of 209 compounds

                comprised of two benzene rings with one to ten chlorines.

                The physical and chemical properties of the 209 compounds,

                referred to as congeners, vary dramatically. Some congeners

                are oily liquids at room temperature, while others are white

                powdery solids.  Vapor pressures vary orders of magnitudes

                among the congeners; boiling points span almost 200 degrees

                (Erickson, 1986).

                   Because of their wide range of physical and chemical

                properties, thermal stability, and chemical resistance to

                acids and bases, PCBs have been used in many industries for

                a wide variety of purposes.     PCBs were commonly used in

                transformer oils, dielectric fluids, hydraulic fluids,

                plasticizers, and calbonless copy paper     (Erickson, 1986;

                National Acaedemy of Science, 1979).

                   PCBs were first manufactured in the late 1920s by direct

                chlorination of biphenyl with chlorine gas. Monsanto Chemical

                Corporation, one of the largest producers of PCBs, marketed

                PCB mixtures under the tradename of Aroclor. Each Aroclor mix

                was sold as a weight percent of chlorine. The last two digits

                of the Aroclor name signified the weight percent.     Aroclor

                1242 contains 42 percent chlorine by weight and consists

                predominantly of the trichlorinated congeners (Furukawa,


















                                                                                 2

                1982)

                    In the early 1930s, PCBs were found to have delitrious

                health effects and workplace threshold limit values were set.

                It was not until 1977, however, that PCBs were banned from

                commercial use because of possible health effects (Erickson,

                1986).   The scientific community is uncertain about the exact

                risks posed by PCBs.       When humans are exposed to PCBs in

                large doses, they are thought to elicit one or more of the

                following responses: wasting syndrome, skin disorders such as

                chloroacne, thymic and splenic atrophy, liver damage,

                endocrine and reproductive dysfunction, tetratogenesis and

                carcinoqenesis (hepatocarcinoma) (Parkinson and Safe, 1987).

                Several congeners are thought to be more toxic than others.

                These "toxic congeners" have a planar configuration similar

                to 2,3,7,8-tetrachlorodibenzo-p-dioxin, one of the most toxic

                environmental contaminants known to man.       Like dioxin, the

                toxic congeners are thought to bind to the direboxynucleic

                acid (DNA) at sites which are coded for enzyme synthesis

                (Chantry, 1989).

                   Because PCBs were used extensively in a variety of

                industries before their toxicological hazards were identified

                and because they are relatively inert, their presence in the

                environment is ubiquitous. PCBs have been measured in air,

                water, sediment, soil and biota around the world. Many sites



















                                                                              3

                in the United States contain high concentrations of PCBs

                (above 50 ppm).   Under federal regulation, these-sites are

                required to be cleaned up to background or limits set by the

                Environmental Protection Agency (EPA). The cost of cleaning

                up such sites is tremendous and the success of cleaning up

                such sites is often marginal.

                    The fate of PCBs in the environment is not completely

                understood, in part because it has been difficult to quantify

                all the physical and chemical properties associated with the

                different congeners.   Aerobic microbial degradation of the

                lower chlorinated PCBs has been shown to occur under certain

                conditions. Recently, there has been evidence that anaerobic

                degradation of PCBs occurs as well (Brown et al., 1987b; Chen

                et al., 1988; Rhee et al., 1989; Quensen et al., 1988). This

                finding is significant because PCBs and PCB-contaminated waste

                is often stored in anaerobic environments such as landfills,

                and sediments.   Determining if anaerobic degradation occurs

                and how it occurs has substantial implications for the

                treatment and disposal of PCBs. Significant economic gains

                could be achieved if sediments could be treated in situ. In

                addition, the risks of exposure would be considerably

                diminished if PCBs and PCB-contaminated material were treated

                in place.



















                                                                             4

                                      Chapter 2   objectives

                    The overall purpose of this study is to examine PCB-

                contaminated sediment from the Sheboygan River to. determine

                if anaerobic dechlorination is occuring.        The observed

                congener distribution will be compared to the predicted

                distributing based on existing knowledge of partitioning

                relationships, diffusion equations, Henry's Law constants, and

                observed biodegradation patterns.

                    The study objectives are fourfold.    The first objective

                is to determine the distribution of PCB congeners in the

                Sheboygan River sediments and to determine the variation with

                depth and distance.   The second objective is to determine if

                there is a significant difference between the congener

                distribution found in the river sediment versus the congener

                distribution found in the Aroclors originally used at the

                industrial site.

                    The third objective is to evaluate processes such as

                partitioning and biodegradation that could account for the

                change in the distribution of the congeners.     The work of

                Burkhard et al. (1985a, 1985b) will be used to evaluate the

                effects of partitioning and volatilization on the congener

                distribution. The work of Brown et al. (1987), Rhee et al.

                (1989) and Quensen et al.(1988) will be used to evaluate the

                effects  of anaerobic biodegradation       on  the    congener















               distribution. The last objective was to examine the congener

               distribution in other matrixes such as fish and water and to

               compare these distributions to that in the sediment.


















                                                                                6

                Chapter 3 Literature Review of Aerobic and Anaerobic Bacteria

                     one of the goals of this research was to determine whether

                 sediment from the Sheboygan River had PCB congener patterns

                 characteristic of anaerobic dechlorination as described by

                 Brown et al. (1984), Quensen et al. (1988), Chen et al. (1988)

                 and Rhee et al. (1989). This chapter presents a synopsis of

                 existing knowledge of aerobic microbial degradation as well

                 as anaerobic microbial degradation.

                     An understanding of aerobic processes is important since

                 rivers are dynamic systems that vary temporally and spatially,

                 scouring and depositing sediments in a somewhat random

                 fashion.   As a result of these processes, it is likely that

                 sediment that was anaerobic becomes aerobic and vice versa.

                 Therefore, sediment samples may have been exposed to both

                 types of environments.

                     This chapter contains a brief overview of the following

                 subjects; aerobic microbial degradation, aerobic bacteria,

                 PCB-degrading enzymesand the environment necessary for PCB-

                 degradation.    A similar overview of anaerobic degradation

                 follows the discussion of aerobic microbial degradation.



                 3.1. Aerobic Degradation of PCBs

                 3.1.1. Introduction

                     Since the first discovery that aerobic bacteria degrade


















                                                                               7

                PCBs (Ahmed and Focht, 1973) , many aerobic, PCB-degrading

                bacteria have been identified. These bacteria are apparently

                widely distributed in the environment. For example, Chantry

                (1989) cites sixteen PCB-metabolizing genera in a review of

                the literature.    Several of these genera have been found in

                the environment, notably Pseudomonas, Vibrio, Aeromonas,

                Microccus,   Acinetobacter,    Bacillus,    and    Streptomyces

                (Furukawa, 1982).    In sediments from the Hudson River, which

                contain PCBs, twenty isolates from five PCB-degrading genera

                have been identified: Acetobacter, Acinetobacter, Alcaligenes,

                Klebsiella and Pseudomonas (Furukawa, 1982).



                3.1.2. Enzyme Systems

                    Two major aerobic enzyme systems are thought to enable

                bacteria to degrade PCBs. All PCB-degrading bacteria employ

                a dioxygenase enzyme, in which both atoms of the oxygen

                molecule are added to the product (Stryer, 1981).             In

                contrast, mammals use a monooxygenase system for PCB

                metabolism.  A monooxygenase system, also referred to as a

                mixed function oxygenase system, is one in which one atom of

                oxygen is added to the product and one atom goes to f orm water

                (Stryer, 1981).

                   The most common enzyme system used by aerobic PCB-

                degrading bacteria is the 2,3 dioxygenase enzyme. Serving as


















                                                                               a

                an electron acceptor, molecular oxygen is attached to an

                vacant 2,3 site (or 5,6 site) on the aromatic ring to produce

                a cyclic peroxide intermedlate and then a cis-dihydrodriol

                (see figure 3-1).     The cis-dihydrodriol then forms a 2,3

                dihydroxy compound which undergoes oxidative meta ring

                cleavage between the first and second carbon atom to form a

                benzoic acid (Safe, 1984). Although the chlorobenzoates can

                be mineralized by bacteria, there are no known bacteria which

                can both degrade PCBs and mineralize the chlorobenzoates.

                    A second pathway for the degradation of PCBs, a 3,4

                dioxygenase, has been proposed by Bedard and coworkers (Bedard

                et al. , 1987a; Bedard et al., 1987b) based on experiments with

                the bacteria, Alcaligenes, and Aroclor 1242. Three of Bedard

                and coworker's experimental results suggest an alternative

                enzyme to the 2,3 dioxygenase.         First, PCBs that were

                sterically hindered for the 2,3 dioxygenase enzyme (i.e

                chlorinated at the 2,3 sites) were readily metabolized by

                Alcaligenes H850.     Second, Alcaligenes was not able to

                metabolize para chlorinated biphenyls; however, it easily

                metabolized ortho chlorinated biphenyls. These results are

                in striking contrast to the conclusions drawn from previous

                experiments which indicated that ortho chlorinated congeners

                are extremely resistant to degradation by bacteria using the

                2,3 dioxygenase system (Furukawa, 1982; Furukawa et al.,









                                   CIX                          Bi.phenyi
                                       02           NADH2       Biphenyl. 2,3-dioxygenase


                                                    NAD +
                                               OH OH
                                              H       H
                                                                cis-2,3-Siphenyl-dihydradial
                                  CIX
                                                    NAD +
                                                                Biphenyl-dihydrodiol dehydrogenase
                                                    NAOH2


                                                                2,3-Dihydroxybiphenyl
                                  CIX
                                      0 2                       2,3-Biphenyl catechol oxygenase

                                            H 01,   OH
                                               0C
                                                                Ring fission product (yellow)
                                  Ox
                                     H20                        Hydrase

                                                    0    0
                                        C"0     H 0 @ C"  ffH
                            CIX @& '@O H          H2CD-K H

                              Figure 3-1       Major Aerobic Degradation Pathway
                                                For PCBs

                         (Source: General Electric Research and Development, 1987)


















                                                                                 10



                 1978).   Lastly, there is some evidence that the expected

                 metabolites from a 3,4 dicxygenase pathway are formed (Bedard

                 et al., 1987b).



                 3.1.3. Aerobic Microbial Degradation Studies of PCBs

                     Many different bacteria have been found to oxidize PCBs.

                 Although they use similar enzyme systems, the extent and the

                 rate of degradation among bacterial strains differ depending

                 an the number and the position of chlorines on the biphenyl

                 molecule. Three experimental results are important.        First,

                 as the chlorine content of the PCB increases, the ability of

                 bacteria to metabolize PCB aerobically decreases.         Second,

                 mixed   cultures   of   bacteria    appear   to   increase     the

                 biodegradation rates of PCBs. This finding is important since

                 natural environments often contain mixed cultures of bacteria.

                 Thirdly, many bacteria appear to require an additional

                 substrate for growth, a process referred to as cometabolism.

                     Tucker and coworkers (1972) examined the ability of

                 microorganisms in activated sewage sludge to degrade Aroclor

                 mixtures at concentrations of 2.5 and 5.0 ppm. Most of the

                 degradation of the mono and di chlorobiphenyls in Aroclor 1221

                 took place in less than a day.         There was a 26 percent

                 decrease in Aroclor 1242 in two days and no degradation of the















                more highly chlorinated Aroclor 1254.      From these results,

                Tucker and coworkers concluded that as the degree of

                chlorination    increased,   the   degree   of    biodegradation

                decreased.

                   Experiments performed by Wong and Kaiser (1975) also

                support this conclusion. Microorganisms (Achromobacter and

                Pseudomonas) from lake water were placed in 0.05 percent

                solution of Aroclor 1221, 1242, and 1254.         The bacteria

                metabolized Aroclor 1221 and 1242 but not Aroclor 1254.

                Aroclor 1221 was completely degraded in one month. Rates for

                Aroclor 1242 and 1254 were not established.

                    Furukawa, Tomizuka and'Kamibayashi (1983) also found that

                as the degree of chlorination increased, the ability of the

                microorganisms to degrade PCBs decreased.            Using PCB

                contaminated soil (50 ppm) and the, bacteria, Acinetobacter,

                these researchers found that Kaneclor 200 (predominantly

                dichlorobiphenyls) was rapidly degraded in four hours.

                Kaneclor 300 (primarily trichlorobiphenyls) and Kaneclor 400

                (primarily tetrachlorobiphenyls) were also susceptible to

                biodegradation but to a lesser degree.            Kaneclor 500

                (primarily pentachlorobiphenyls) was recalcitrant. Similarly,

                microbes from activated sewage sludge were unable to degrade

                Kaneclor 500 at concentrations of 0.1, 5, and 10 ppb (Kaneko

                et al., 1976).

















                                                                                 12

                    Mixed bacterial cultures may be better able to degrade

                PCBs than pure strains.     A mixed strain was found to degrade

                Aroclor 1242 in five days, a rate considerably faster than

                values previously reported in the literature (Clark et al.,

                1979).    The increase in the rate may be because only readily

                water soluble PCBs were used (a solution was saturated with

                Aroclor 1242). This may have biased the experiment because

                the lower chlorinated congeners are more soluble and also more

                easily degraded.      In addition, the PCBs may have been more

                available to bacteria since they were in solution and not

                adsorbed to particulates.         Again the lower chlorinated
                congeners were most'easily degraded.

                           Furukawa and coworkers (1983) extensively studied

                two bacterial strains, Alcaligenes and Acinetobacter, both of

                which degrade   PCBs using a 2,3 dioxygenase enzyme.           The

                following generalizations about aerobic PCBs degradation

                result from their work and      summarize much of the aerobic


                research to date.

                     1) As the number of chlorines increased on,the biphenyl

                molecule, the ability of the bacteria.to degrade the compound

                decreased.     This decrease may be due to increased steric

                hinderance as  additional chlorines are added to the molecule.

                Congeners with greater than four chlorines were recalcitrant.

                     2) PCBs containing chlorines in the ortho position of the

















                                                                                   13

                 ring were also resistant to degradation. These congeners may

                 be sterically hindered; the 2,3 dioxygenase system requires

                 vacant sites at either 2,3 or 5,6 position.         If both ortho

                 positions are filled on the opposite ring to the ring that the

                 enzyme is attacking, the attack by the enzyme will be

                 hindered.

                       3) PCBs containing all the chlorine atoms on a single

                 ring were generally degraded faster than isomers with the same

                 total number of chlorines distributed on both rings.

                       4) Ring cleavage occured more often with congeners that

                 had fewer chlorines.

                       5) PCBs having adjacent unchlorinated sites, particularly

                 at the 2,3 position, were more readily degraded than those

                 congeners chlorinated at the 2,3 position.

                      In addition to these factors, it appears that a substrate

                 for growth may be important since PCB-degrading bacteria are

                 unable to completely mineralize PCBs.

                      Cometabolism, as defined by Horvath (1972), is "any

                 oxidation of substances without the utilization of the energy

                 derived from the oxidation to support microbial growth. 11

                 There is evidence that the higher chlorinated congeners cannot

                 be degraded without additional carbon sources.            "Although

                 complete mineralization of monochlorobiphenyls has been

                 reported, no culture with the ability to completely mineralize


















                                                                                 14

                 more highly chlorinated biphenyls or to grow on any congener

                 that has one or more chlorine on each ring has unequivocally

                 been described" (Kohler et al., 1988).            A variety of

                 substrates have been used.       Biphenyl is by far the most

                 common; however, acetate has also been used (Clark et al.,

                 1979).

                     Brunner and coworkers (1985) found the most important

                 factor affecting PCB degradation in soil was enrichment of the

                 soil with biphenyl.      After incubating soil for 210 days,

                 Brunner and coworkers noted that little if any degradation

                 took place in soil that was not inoculated with a growth

                 medium (such as biphenyl) or with bacteria (such as

                 Acinetobacter).     In contrast, when the soil was inoculated

                 with a growth medium such as biphenyl, degradation increased

                 tenfold. Inoculation with bacteria increased biodegradation,

                 but not as dramatically. With the exception of 4-

                 chlorobiphenyl, Acinetobacter cannot grow upon any of the PCBs

                 because it is unable to dehalogenate the ring fission products

                 (Brunner et al., 1985).
                      Kohler also noted that the addition of biphenyl

                 substantially increased the ability of bacteria to metabolize

                 PCBs.   When bacteria, Acinetobacter and Arthobacter, were

                 grown in the presence of biphenyl, degradation rates of

                 Aroclor,1254 increased substantially (Kohler et al, 1988).

















                                                                             15

                      Interestingly it has been found that adding Aroclor 1221

                increased the ability of the Pseudomonas to degrade Aroclor

                1254 (Lui, 1980).        This phenomenon is suggestive of

                cometabolism. Aroclor 1254 (300 ppm) was degraded in 18 days

                to 0.59 ppm. In contrast, when Aroclor 1221 was added, the

                bacteria degraded the same concentration of Aroclor 1254 in

                8 days.    Sodium ligninsulfonate was also added to these

                cultures resulting in the formation of an emulsion.      It is

                hypothesized that the emulsion increased the surface area of

                PCBs available to the bacteria and therefore expedited the

                biodegradation of these compounds (Lui, 1980).

                     Another example of cometabolism is a study of four

                congeners by Parson and Sijm (1988). Grown on biphenyl and

                4-chlorobiphenyl in a continuous culture system, Pseudomonas

               -could metabolize 4-chlorobiphenyl, 2,213,31 (congener 40),

                2,215,51 (congener 52), 2,3,1415 (congener 70) and 3,314,41

                (congener 77).    If, however, the Pseudomonas were cultured

                on nutrient broth or benzoates they eventually lost the

                ability to metabolize these four congeners (IUPAC # 40, 52,

                70, 77).



                3.2. Anaerobic Degradation of PCBs

                3.2.1. Introduction

                    Although the fate of PCBs in aerobic sediments has been

















                                                                              16

                well studied, very little is known about the fate of PCBs in

                anaerobic sediments such as landfills, and confined disposal

                facilities.    Since anaerobic environments such as river

                bottoms and lake bottoms represent the final sink for PCBs

                (National Academy of Science, 1979), it is important to

                determine what, if any, processes PCBs undergo in anaerobic

                environments.

                    In contrast to the plethora of information that exists

                about aerobic microbial degradation, there is surprisingly

                little information about anaerobic microbial degradation.

                Anaerobic microbial Inetabolic pathways are not well understood

                and anaerobic bacteria which can degrade PCBs have not been

                isolated.

                    The first studies presented little if any experimental

                evidence of anaerobic degradation.        Work by Kaneko and

                coworkers (1976) showed that anaerobic incubation of Kaneclor

                500 (equivalent to Aroclor 1254) in the presence of sewage

                sludge for twenty days resulted in no degradation.

                Similarly, when Aroclor 1242 was incubated anaerobically in

                soil enriched with biphenyl and Acinetobacter, no evidence for

                degradation was seen (Brunner et al., 1985).

                    Although these studies were useful, they do not

                conclusively demonstrate that these compounds do not degrade

                in an anaerobic environment.            Alternatively, these

















                                                                              17

                experiments may indicate that the environment present in the

                experiment was not suitable for anaerobic degradation.         A

                variety of environmental conditions can affect the type of

                microbial degradation and the organisms present. Some of the

                more common variables are pH, temperature, nutrients (such as

                yeastextract), reduction potential, substrate, andenrichment

                techniques.



                3.2.2. Anaerobic Degradation of Structurally Similar Compounds

                   Although initial studies of anaerobic degradation of PCBs

                presented   little    evidence   of   anaerobic    degradation,

                structurally similar compounds such as the chlorophenols

                (Woods et al., 1989; Mikesell and Boyd, 1988; Krumme and Boyd,

                1988), 4-chlororesorcinol (Fathepure et al., 1987) and

                chlorobenzoates (Horowitz et al., 1983; Suf lita et al., 1982,

                Palmer et al., 1989) have been shown to be degraded by

                anaerobic organisms.     Chlorobenzoates are perhaps the most

                relevant of the three compounds mentioned above because they

                are intermediate products in the microbial metabolism of PCBs.

                     Mono, di, and trihalobenzoates have been found to be

                completely mineralized to carbon dioxide and methane using

                bacteria from lake sediments and sewage sludge as well as

                enriched cultures grown on 3-chlorobenzoate (Suflita et al.,

                1982; Horowitz et al., 1983).      Dechlorination was always


















                                                                             18

                accompanied by methane production.    The proposed reductive

                dechlorination pathway is shown in figure 3-2. In contrast

                to aerobic degradation, removal of the chlorine was necessary

                before the chlorobenzoate could be mineralized to carbon


                dioxide and methane.

                     The dechlorination was thought to be a biologically

                mediated reaction for two reasons. First, the reaction was

                inhibited if oxygen was present or if the sediments had been

                sterilized (e.g., autoclaved, radiated, or poisoned with

                formaldehyde).   Obligate anaerobes, for obvious reasons, do

                not function in the presence of oxygen.             Autoclaving

                terminates biological degradation because most    enzymes are

                denatured above 39 degrees C (Keeton, 1980). Zeikus and

                Winfrey (1976) also found similar temperature limitations in

                their study of methanogenic organisms in aquatic environments.

                     Second, a lag period was observed that lasted 1 week to

                52 weeks depending on the position and number of chlorines on

                the benzoate.    once the microbes were acclimated, no lag

                period was seen.      A lag period is commonly found in

                biologically mediated degradation.   Time is needed to select

                for a population with the necessary enzymes for metabolizing

                the chemical.   There was a preferential loss of the chlorines

                in the meta position indicating that position of the chlorine

                was important.










                  COOH    C1         COOH    CC        COOH             CH -4

           C I         C I                 C I                          C02





                  Figure 3-2 Anaerobic Degradation of Chlorobenzoates

                   (Source: Suflita et al., 1982)


















                                                                                   20





                        The lag period was found to be correlated to the amount

                 of   substrate     present.       For    example,    4-amino     3,5

                 dichlorobenzoate at low concentrations (13 to 26 gM) was not

                 degraded after 21 weeks whereas higher concentrations (820 gM)

                 were degraded within four weeks (Horowitz et al., 1983).

                     In experiments with chlorophenol, the position and the

                 number of chlorines also influenced the degree of degradation.

                 Ortho chlorinated congeners had the shortest lag period (7-21

                 days).    Meta chlorinated isomers followed with a lag period
                 of 31-49 days and para chlorinated congeners were not degrad'ed

                 (Woods    et   al.,   1989).       No   dechlorination     of    the

                 monochlorophenols or chlorines from the position para to the

                 hydroxyl group (a metabolite) was seen in the entire 7 months

                 of the experiment.

                     In studies with 4-chlororescorinol, a product of dye and

                 pharmaceutical industries, it was found that the lag period

                 could be decreased from three weeks to two days by adding

                 yeast extract and trypticase. In addition, the rate of

                 dechlorination increased (Fathepure et al., 1987).

                     These studies suggest three generalizations.             First,

                 position and number of chlorines , af f ect the ability of

                 microorganism to degrade the compound.               Second, the


















                                                                             21

                degradation pathway may be a function of the concentration of

                the compound.   High concentration may induce degradation to

                occur whereas low concentrations may not. Third, additional

                substrates such as yeast extract may be necessary for

                dechlorination to occur.




                3.2.3 Anaerobic Microbial Degradation Studies of PCBs

                    Few articles in the existing literature describe

                anaerobic degradation of PCBs. Much of the current knowledge

                of anaerobic degradation has come from the General Electric

                Research and Development Center in New York.     This section

                describes their study of anaerobic dechlorination in sediments

                from the Hudson River and two other research studies of


                anaerobic dechlorination.

                    Between 1952-1973, an estimated 362,000 kg of Aroclor 1242

                was released into the Hudson River in New York (Brown et al.,

               .1984) contaminating over 134 metric tons of sediment (Brown

                et al., 1987b). Hudson River sediments were analyzed by the

                General Electric Research Group to determine the total

                concentration of PCBs in the sediment and the distribution of

                congeners. The concentrations ranged from 1 ppm to 2700 ppm

                (Brown et al., 1984).      When the chromatograms from the

                sediment samples were compared to a standard chromatogram of

                the original source material, Aroclor 1242, a loss of the


















                                                                              22

                higher chlorinated congeners was seen.    A corresponding gain

                in the lower chlorinated congeners was observed.              In

                particular, there was a relative decrease in the levels of

                tri, tetra and penta chlorobiphenyls and increase in the

                levels of mono and di chlorobiphenyls.      These results were

                attributed to anaerobic microbial degradation.

                    This finding is particularly significant for at least

                three reasons. First, all prior aerobic studies had suggested

                that the more highly chlorinated congeners (notably those

                containing more than four chlorines) were very resistant to

                degradation.

                    Second, this degradation process appears to dechlorinate
                the more highly chlorinated congeners such as the "toxic

                congeners" and thus reduce the risk that these congeners

                present.      The "toxic congeners" have a 4,41 chlorine

                substitution with at least two chlorines in the meta position

                and no more than one ortho chlorine which is adjacent to the

                meta chlorine.    Although the overall concentration of PCBs

                in the sample remains the same, the reduction in the number

                of chlorines increases the potential for aerobic degradation

                to occur and decreases potential risk to health and biota.

                The lower chlorinated congeners are generally thought to be

                less toxic than higher chlorinated congeners.

                    Lastly, typical PCB residues in the sediment today are

















                                                                              23

                the more highly chlorinated biphenyls, even though they

                represent only about 35 percent of the total PCBs manufactured

                (Furukawa, 1982).    It is important to identify bacteria cr

                processes which can degrade the more highly chlorinated

                congeners since they are a significant proportion of the PCBs

                in the environment.

                    Brown and coworkers (1984) have identified several

                chromatographic patterns which they attribute to different

                bacterial strains. Within the Hudson River, five patterns A,

                B, C, E, and X (Brown et al., 1984) occur. They are outlined

                briefly below and discussed in more detail in chapter seven.

                Each of the patterns shows a'loss of meta and para congeners.

                    The congener distribution of pattern A is similar to

                Aroclor 1242.   Relative to Aroclor 1242, concentrations of

                mono and di chlorinated congeners and tri and tetra

                chlorinated congeners are reduced (e.g., congeners 18, 17, 28,

                20/21/33, 22, 44, 37, 70/76, 66 and 56/60).            Relative

                concentrat.ions of the penta and hexa chlorinated congeners are

                slightly enhanced.     This pattern was typically seen in

                sediment samples from surface-water interfaces.

                    Pattern B was found in many of the Hudson River samples.

                 Again in comparison to Aroclor 1242, pattern B is

                characterized by a decrease in the concentrations of congeners

                eluting after congener 52/49/47 as well as a decr      ease in

















                                                                             24

                congeners 28/31 and 22. In general, there was an increase in

                the mono, di and tri chlorinated congeners.        Within the

                dichlorinated congeners, congener 6 and congener 8 increase.

                Within the trichlorinated congeners, congeners 17, 16, 32 and

                27 are substantially increased.    Pattern BI was the same as

                B with the exception that congener 6 was not present. Table

                3-1 shows the homolog weight percents for typical A and A/B

                patterns.

                   In comparison to Pattern B, pattern C showed less removal

                of the higher chlorinated congeners with the exception of

                congeners 52/49/47, which were readily removed. The di and

                tri chlorinated groups decreased, most notably congeners 6,

                5/8, 17 16/32 and 27.         Congeners 1, 3,        and 4/10

                correspondingly increased.

                    Pattern E shows a selective loss of the higher chlorinated

                congeners and in general was found in combinations with

                pattern B and C. Pattern X was similar to pattern B; however,

                levels of congeners 1, 6, and 5/8 were below pattern B; levels

                of 19, 24, and 28 were above pattern B.

                    Brown and coworkers found a significant decrease in the

                meta and para chlorines in all of the above patterns.

                Thermodynamical analysis shows that the meta and para

                dechlorination is favored over ortho dechlorination.        The

                standard reduction potentials (E) of congeners with chlorines

















                                                                                25

                 in the ortho'position, are more negative than isomers





                    Table 3-1 Homolog Distributions for Pattern AB and B

                 Pattern   Conc       of Samples)     Wt % of Homolog Group

                            (ppm)                      (Number of Chlorines)

                                                  1       2      3        4      5


                 Pattern A+B 162.1      (8)      6.6   26.6   39.6     18.1    8.1

                 Pattern B     659      (13)    13.0   41.6   30       13.7    4.0

                 Aroclor 1242                   0.7    13.8 46.2       30.5    8.5




                 (Source: Brown et a l., 1984)



                 containing the same number of chlorines (Rusling and Miaw,

                 1989). For example, the toxic congener 3, 3', 4, 41 does not

                 contain any ortho    chlorines and it has a more, positive

                 standard potential. It is also one of the more easily reduced

                 compounds.   A more  positive standard potential indicates a

                 greater tendency for the reaction to occur as indicated by the

                 Nernst equation: delta G = -n F (delta E).

                    Bacteria which are able to anaerobically degrade PCBs may

                 have a thermodynamic advantage over those that do not (Brown

                 et al., 1987b). Since the PCB nucleus is not destroyed when

                 reductive dechlorination occurs, it is possible that the
















                                                                                  26

                 microbes use the PCBs as electron acceptors rather than a

                 carbon source (Brown et al., 1987b)          Brown and coworkers

                 compared the calculated values of Gibbs free energies for the

                 oxidation of glucose for a variety of reactants (table 3-2).

                  The second highest energy level of any acceptor is

                 hexachlorobenzene, which is similar in structure to PCBs.

                 Note the energy value is four times higher than



                 Table 3-2 Energy Derived from the Oxidation of Glucose
                            to Carbon Dioxide and Water using Various
                            Oxidants
                                          Reduced            Gibbs Free
                 Oxidant                  Eroduct            Energy (kcal/mole)

                 Oxygen                   Water              -676.1
                 Hexachlorobenzene        Benzene            -410.16
                 Monochlorobenzene        Benzene            -369.5
                 Sulfate                  -Sulfur            -131.78
                 Carbon Dioxide           Methane            - 95.63

                 (Source: Brown   et al., 1987)


                 carbon dioxide.

                     Two other research groups have examined the ability of

                 anaerobes to degrade PCBs.          Both groups concluded that

                 dechlorination occurs.      Their experiments were carried out

                 under different conditions and subsequently their conclusions

                 are different although not necessarily contradictory.

                      Quensen et al. (1988) extracted anaerobes from the Hudson

                 River at a site which ranged in concentration of PCBs from 60@

                 to 562 ppm. After introducing the anaerobic culture into a

















                                                                             27

                series of sterilized, "clean" Hudson River sediments, PCBs

                were added at concentrations of 14 ppm, 140 ppm and 700 ppm.

                to different reaction vessels.     Controls were established

                using sterilized samples and no inoculation.

                     Reductive dechlorination occured most rapidly in the more

                highly contaminated sediments (Quensen et al., 1988). In the

                700 ppm sample, 53 percent of the chlorine was removed in four

                months with an increase in the mono and di chlorinated

                biphenyls of 71 percent. No dechlorination was observed in

                the least concentrated sample (14 ppm).        The sterilized

                control sediments and sediment containing microorganisms from

                a PCB-free site in the Hudson River showed no evidence of

                dechlorination.

                     Dechlorination appeared to occur predominantly from meta

                and para positions.   This finding supports the thermodynamic

                advantage of the meta and para positions postulated by Brown

                et al. (1987) and Rusling and Miaw (1989). It is interesting

                to note that Quensen and coworkers observed a stepwise

                decrease in the chlorine composition of the 700 ppm sample

                with time.

                          The Wadsworth Center for Laboratory and Research,

                Department of Health, in New York also preformed several

                anaerobic experiments. An enriched bacterial culture from the

                Hudson River was introduced into a medium containing 20 ppm


















                                                                             28

                of Aroclor 1221 ( a lower chlorinated Aroclor

                consisting mainly of mono and di chlorinated biphenyls) and

                a 90.6 percent decrease was observed in the monochlorinated

                biphenyls and a 65.5 percent decrease in the dichlorinated

                biphenyls (Chen et al., 1988).

                     When bacteria were incubated anaerobically with pure

                monochlorinated biphenyls (0.75 ppm) for 80 days, complete

                degradation of the chlorinated biphenyls was seen.           In

                contrast, w hen a higher chlorinated congener, 2,4,21,41

                tetrachlorobiphenyl was used, no anaerobic degradation was


                seen.


                      Absence of dechlorination in the higher chlorinated

                congener does not conclusively demonstrate the inability of

                bacteria to anaerobically degrade the higher chlorinated

                congeners.    Insufficient incubation period or insufficient

                quantities of substrate could inhibit degradation.          The

                anaerobic bacteria were incubated for 40 days with the higher

                chlorinated congeners and 80 days with the lower congeners.

                In contrast, Quensen and coworkers used an incubation period

                of 112 days.    It is possible that a lag period may occur

                before significant dehalogenation takes place.      A lag was

                observed in Quensen's work where very little dechlorination

                occured in the first month.        In addition, Quensen and

                coworkers found dechlorination to occur only in the more



















                                                                              29

                highly concentrated samples (above 700 ppm) and not in the

                less concentrated samples (14 ppm)       The concentration of

                tetrachlorobiphenyl in the experiment by Wadsworth Center was

                considerably lower (0.25 ppm (Chen et al., 1988)).     This may

                be an insufficient quantity of substrate to support microbial

                growth.

                    Recently, this same research group found significant

                anaerobic degradation to occur in untreated sediments,

                biphenyl -amended sediment, and biphenyl -amended and bacteria-

                inoculated sediment from the Hudson River and Moreau sediment

                (Rhee et al., 1989).     The concentrations of PCBs were much

                higher than previous experiments, 707 ppm and 936 ppm

                respectively.      These sediments were under a nitrogen

                atmosphere.    No degradation was observed under an anaerobic

                carbon dioxide/hydrogen gas environment with similarly treated

                sediments.   It is unclear why. Quensen and coworkers (1988)

                found bacterialidegradation to occur in an environment which

                was 80 % nitrogen and 20 % carbon dioxide.

                     The biphenyl-amended sediments showed a significant

                decrease in the higher chlorinated. congeners. This suggests

                that biphenyl may be needed for the degradation of the higher

                chlorinated congeners, a finding which has also been suggested

                for the aerobic cometabolism of the higher chlorinated

                congeners.



















                                                                          30

                  Chapter 4 Site Description and Historical Use of PCBs


                  This chapter describes the hydrology and geology of the

               study area. The history of PCB usage at the site is

               presented as well as a btief summary of the regulatory

               precautions that were enacted to protect human health and

               the environment.

                   The Sheboygan River flows westward and drains into Lake

               Michigan at the city of Sheboygan. Sheboygan is

               approximately 89 km.directly north of Milwaukee. The river

               is a total of 220 stream km long and has a drainage basin of

               approximately 272 square km (Wisconsin Department of Natural

               Resources, 1989). Figure 4-1 shows a map of the Sheboygan

               River and drainage basin as well as its two tributaries, the

               Onion and the Mullet rivers. Beginning at the Sheboygan

               Falls dam and ending at the harbor in Sheboygan, the 22.4 km

               river segment studied is a small portion of the total

               length. This portion of the river drains approximately 57

               square km (Wisconsin Department of Natural Resources, 1989).

               The river is slowed by two small dams located near the

               village of Kohler.

                   The Sheboygan River has a mean annual discharge of

               approximately 7.3 cubic meters per second ( 258 cfs) based

               on data collected from 1942 to 1986. The lowest flow for

               this century 0.0283 cubic meters per second (1 cfs) was

















                                                                                                                                     31



                               Figure 4-1 The Sheboygan River and Tributaries




                                            -N-




                                       0               1 Miles

                                                                                           Ole
                                                                                        .0e


                                                                                       Sheboygan






                                                          Kohler
                                                                                    River


                           Sheboygan
                             Falls



















                                                                                                         lrwwpotmtd Area



                                                                                                         Dam



                                                                                                        Waumbed boundary










                            (Source: Wisconsin Department of Natural Resources, 1989)

















                                                                                 32



                 recorded in August, 1922.     The highest flow was 217 cubic

                 meters.per second (7,680 cfs) in March, 1975. Average

                 annual rainfallduring 1978 to 1986 was 762 cm per year

                 (Wisconsin Department of Natural Resources, 1989).

                 Although, the Wisconsin Department of Natural Resources

                 (WDNR) concluded that the river bottom from Sheboygan Falls

                 to Sheboygan was generally scoured in 1980, major areas of

                 deposition were noted, above both of the dams, near Kiwanis

                 Park in Sheboygan and near the Eight street island in

                 Sheboygan. The average rate of deposition was calculated to

                 be 10 cm/yr (Wisconsin Department of Natural Resources,

                 1989).    The river deposits approximately 22,923 cubic

                 meters (30,000 cubic yards) of sediment a year into.the

                 harbor (Wisconsin Department of Natural Resources, 1989).

                    The Sheboygan River lies in the geologic province of the

                 Eastern ridges and lowlands. The dominant feature is a

                 plain interspersed with north-south ridges, paralleling       Lake

                 Michigan. 'The sedimentary bedrock is comprised

                 alternatingly of weather resistant dolomites which form the

                 ridges, and erodible sandstones and shales which form the

                 lowlands (Paull and Paull, 1976). Glacial features such as

                 kames, eskers, drumlins and kettle holes are abundant.

                 Within the river, the sediment is predominantly fine-grained


















                                                                            33

                sand interspersed with silt and occasionally clay.

                    The Sheboygan River and harbor have been extensively

                studied since the area was first identified as an "Area of

                Concern" by the International Joint Commission and as a

                I'Superfund site" by the Environmental Protection Agency

                (EPA). An "Area of Concern" is defined as a locality where

                water quality objectives, standards, criteria or guidelines

                are not being met and remedial actions are necessary to

                restore the water quality.    A I'Superfund site" is an area

                which, according to EPA's national ranking system, poses a

                substantial risk to human health and the environment. once

                an area is designated as a Superfund site,- parties

                responsible for cleaning up the site are identified, where

                possible, and remediation efforts are initiated under the

                supervision of government agencies.     If no responsible

                party can be identified, money from the national hazardous

                waste trust fund can be used to clean up the site.     A

                lengthy study of the extent of contamination and other

                relevant information such as groundwater flow, potential

                routes of exposure, and possible treatment alternatives is

                necessary before.any remediation can begin.

                     Contamination of the river sedimehts by PCBs was first

                identified in 1978,.when the WDNR found sediment containing
 0              190 ppm of PCBs downstream from Tecumseh Products, a die

















                                                                          34

              casting plant located on the river at Sheboygan Falls.

              Further sampling at the Tecumseh Products site revealed soil

              samples with concentration of PCBs as high as 120,000 ppm.

              The samples were found to contain oil-soaked soil as well

              as oil soaked rags, pressure hoses and other refuse

              (Wisconsin Department of Natural Resources, 1989).

                  When the Sheboygan River and harbor was identified as a

              Superfund site, Tecumseh Products was named as potentially

              responsible party. Tecumseh Products and the previous

              company at the industrial site, the Die Cast Corporation,

              used hydraulic fluids in the manufacture of outdoor motors.

              Use of hydraulic fluids containing PCBs commenced after a

              large fire occurred at the Die Cast plant. The cause of the

              fire was attributed to the combustion of hydraulic fluids.

              Subsequently the Die Cast Corporation switched to a

              hydraulic fluid which was fire resistant.

                   Monsanto Company began to manufacture fire resistant

              hydraulic fluids containing PCBs in the early 1950's. The

              first formulation that they marketed was Pydrol F9, the

              formulation believed to be used at Tecumseh Products and the

              Die Cast Corporation. Table 4-1 contains the chronology of

              PCB use at these two companies. The major formulations used

              by Tecumseh contained Aroclor 1248 and 1254. Tecumseh

              Products acquired the Die Cast Corporation in 1966 and used


















                                                                             35

                the same equipment as the Die cast corporation.      Based on

                interviews and available records, it is thought that the Die

                Cast Corporation also used Pydrol F9.

                    Tecumseh Products ceased using hydraulic fluids

                containing PCBs in 1971 because the major producer of PCBs

                restricted sales to closed systems such as transformers.

                The government did not prohibit the 'manufacture of PCBs and

                use of PCBs until July 1, 1977 (Federal Register 42 part

                6531 and Federal Register 44 part 315141).     Before PCBs were

                regulated, Tecumseh Products used material     from the plant

                and soil from around the plant to construct    a low dike at

                the river's edge. Since the dike sloped at     45 degree angle

                into the river, it was relatively easy for     sediment to be
                introduced into the river during high water    events

                (Wisconsin Department of Natural Resources,    1989).

                        In 1979, Tecumseh Products voluntarily removed 2,046

                cubic meters (72,300 cubic feet) of PCB-contaminated

                material from the dike and surrounding area. This material

                was sent to a federally-licensed disposal area outside of

                Cincinnati, Ohio.

                         Since that time the area has been designated an

                "Area of Concern" by the IJC as well as "Superfund site".

                The EPA has extensively sampled the river and some of the

                harbor.     The U.S. Army Corps of Engineers has completed

















                                                                            36






                 Table 4-1 PCB Mixtures used at the Tecumseh Products Site




               Company                Date          Formulation used

               Die Cast Corp       1959-1966       Thought to have used

                                                   Pydrol F9,(52.5% 1248)

               Tecumseh Products   1966-1969       Pydrol F9 (52.5% 1248 a)

                                   1970-1971       Chemtrend HF30 (65% 1254,
                                                      4.6% 1248 a)

               a -- The remaining percentage is a non-PCB product

               (Source: Wisconsin Department of N  atural Resources, 1989;
               Mark Thimke, Lawyer for Tecumseh Products, Lauderin and
               Associates,,Milwaukee, WI, personal communication; Paul
               Micheal, Analytical Chemist, Monsanto, Chicago, IL, personal
               communication.)



               several studies of the harbor to assess the nature and

               extent of contamination. Contaminated sediments in the

               harbor have prevented dredging of the harbor since 1969.
               This has resulted in a subs@antial loss to the shipping

               industry because barges must travel with less than a full

               cargo to get in and out of the harbor.

                    A fish consumption advisory has been in effect since

               1978 when it was found that fish from the Sheboygan area

               exceeded the'Food and Drug Administration limit of 5 ppm,


















                                                                         37

               which was subsequently lowered to 2 ppm in 1984 (Federal

               Register 38 part 18096, Federal Register 49, part 21514).

               A waterfowl consumption advisory was issued in 1987 when the

               waterfowl were found to exceed the 3 ppm fat based limit set

               by the Food and Drug Administration (Wisconsin Department of

               Natural Resources, 1989).

                   Most recently, the EPA has begun the removal this fall

               of 1,910 cubic meters (2,500 cubic yards) of sediment from

               the river in the vicinity of Tecumseh Products and Rochester

               Park. These highly contaminated sediments will be placed in

               a.confined disposal facility and monitored to determine.if

               anaerobic and aerobic biodegradation is occurring. In

               addition, 1,854 square meters (20,000 square feet) of

               sediment will be amoured in place by covering the sediments

               with a fabric liner, followed by sand and gravel. The

               sediments will remain amoured until final treatment

               decisions are made in 1991 (United States Environmental

               Protection Agency, 1989).











                    chapter 5 Sample Collection, Materials and Methods

                    This chapter delineates how sediment, fish and water

               samples were collected and prepared for high resolution gas

               chromatography analysis. A summary of operating conditions

               of the gas chromatographs is included as well as a

               discussion of the quality control measures used. Lastly,

               the procedure used for analyzing grain size is explained.



               5.1. Sample Collection

               5.1.1. Sediment

                    Two sediment sampling trips were made to the Sheboygan

               River (December 2, 1988 and April 28, 1989). On the first

               trip, five sites were sampled: above the Sheboygan Falls dam

               (Sheboygan Falls, site A), at Rochester Park (Sheboygan

               Falls, site B), above the upper Kohler dam (Kohler, site C),

               above the lower Kohler dam (Kohler, site D) and near Kiwanis

               Park (Sheboygan, site E). Figure 5-1 shows the exact

               locations. on the second trip, samples were only taken at

               Rochester Park (Site T).

                    Based on information from previous studie s, these five

               sampling sites were selected because it was thought they

               would represent concentrations varying from high (Rochester

               Park) to low (above Sheboygan Falls dam). At each site, an

               effort was made to sample areas of deposition, since there


















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                                                                           41

                is evidence that all things equal that finer grained

                material contains higher concentrations of PCBs than coarser

                grained material (Karickhoff, 1979).

                     Sediment cores were taken with a hand-held cylindrical

                coring device which was plunged into the soft sediment. The

                coring device was then slowly removed and a cap placed on

                the bottom of the cylinder while it was still under water.

                Since the cylinders, approximately 8 cm in diameter and 1.5

                meters in length, were made out of clear plexiglass, it was

                relatively easy to examine the core. In all cores , the

                sediment-water interface remained undisturbed.and intact

                with distinct boundaries between different sediment

                compositions. The cores were extruded out of the cylinders

                using a hydraulic pump. Each core was divided into 15 cm

                segments which were put into glass mason jars with teflon

                caps and stored at 4 degrees C until time of analysis.




                5.1.2. Fish

                     Fish samples were collected by the Wisconsin Department

                of Natural Resources in the summer of 1988. Ten of the fish

                were collected in the Kiwanis Park area; the remaining

                eleven fish were caught in the harbor area.   The fish

                tissue was frozen until time of analysis.



















                                                                           42


                5.1.3 Water

                    Water samples were collected from the Sheboygan River

                at five locations corresponding to the five locations of the

                sediment cores. Two sampling trips were made. One on

                August 3 to collect water from Kiwanis park, above Sheboygan

                Falls and Rochester Park; and a second on August 17 to

                collect water from Kiwanis park, above the upper Kohler dam,

                and below the lower Kohler dam.

                   At each site, samples were taken from a boat in the

                middle of the river. A narrow-neck one liter bottle

                enclosed in a steel cage was dropped gently over the edge of

                the boat and lowered to the river bottom, usually no more

                than one to two meters. The bottle was then pulled at a

                rate such that the bottle was just filled when it was a few

                centimeters from the water's surface. This method, referred

                to as the depth integrated profile method, is thought to

                give a more representative sample than a single point-sample

                (personal communication, Leo Haus, United States Geologic

                Survey, 1989). The one-liter bottle was cleaned according

                to State Laboratory of Hygiene's procedure for water

                sampling bottles (1989) and was rinsed with river water

                before sampling. A clean bottle was used at each site.

                    The narrow-neck bottle was emptied into a nineteen-

                liter beverage canister. The bottle was lowered several


















                                                                          43

               more times until the canister was filled. The canisters

               were rinsed with hexane three times before leaving the lab.

               The canisters were brought back to the lab and filtered

               within three days.



               5.2. Materials


               5.2.1. Solvents


                   Solvents: Acetone, Cyclohexane, Ethyl Ether, Hexane,

               Iso-octane, Methylene Chloride. High purity (Baxter,

               Burdick and Jackson).

               5.2.2. Chromatographic Adsorbents

                   Florisil: Pesticide residue grade, 60/100 mesh,

               activated at 130 degrees C for 12 hours and stored until use

               in an air tight container at room temperature (Floridin

               Co.).

                   Silica gel: 100-200 mesh, grade 923, activated at 130

               degrees and stored until use at 130 degrees C until use.

               Deactivated with 5 % distilled water at least one hour

               prior to use (Davison Co.).

               5.2.3. Standards

                    Aroclor standards 1232, 1242, 1254, 1260 in methanol

               were obtained from the EPA-EMSL Quality Assurance Branch in

               Cincinnati, OH and diluted to appropriate levels. Internal

               standards, congener 30 and 204, were ordered from Ultra-


















                                                                          44


               scientific.


               5.2.4  Other Chemicals and Materials

                   Sodium Sulfate: granular, heated at 130 degrees

               overnight to activate (Mallinckrodt).

                   Copper: granular, 20-30 mesh (J.T. Baker Inc.)

                   Glass wool: soxhlet-extracted in 50/50 acetone hexane

               for four hours (Corning glassworks).

                   Boiling Chips: soxhlet-extracted in 50/50 acetone/hexane

               for four hours (Cargille Scientific Co).

                   Glass Filters: 29.3 cm diameter GC 50 grade, wrapped in

               aluminum foil and ashed for ten hours at 400 degrees C and

               stored in foil packets until used (Micro Filtration

               Systems).

                   XAD-2 Resin: (Sigma Chemical Co.) Resin was extracted

               fdr 24 hours each in methanol, acetone, hexane, methylene

               chloride and then extracted four hours in acetone, hexane

               and acetone. The resin was then rinsed with distilled water

               and stored in glass bottles filled with distilled water

               until needed.

                   ASTM Soil Sieves for grain size analysis

                   19-liter stainless steel canisters (Beverage Industry).

                   Bio-beads Gel Resin (SX-3): for the Gel Permeation

               Column (Biorad).


















                                                                          45

               5.2.5. Equipment

                   Hewlett Packard 5880 Gas Chromatograph with DB-5 column

               and ECD detector.

                   Hewlett Packard 5890 Gas Chromatograph with DB-1 column

               and ECD detector.

                   Hewlett Packard 5790 Gas Chromatograph with a Packed

               Column.

                   Low Pressure Gel Permeation Column Autoprep 1001

               Chromatograph (ABC Labs)

                   Rotovap and air blowdown apparatus

                   Filtration apparatus: 293 mm millipore filtration head

               5.2.6. Glassware

                   Cleaning protocol: Glassware was washed with laboratory

               detergent, rinsed with hot municipal tap water and two

               rinses of distilled water, and dried at 180 C. 'For water

               analysis, all glassware was rinsed three times with hexane.

                   General glassware as well as chromatographic columns (2

               cm id and 1 cm id), resin columns (3 cm by 17 cm), 500 ml

               separatory funnels, glass soxhlets and flasks were needed.



               5.3. Extraction and Cleanup Methods

               5.3.1. Sediment

                    The method used to cleanup sediments was based on the

               State Laboratory of Hygiene procedure for congener specific


















                                                                            46

               PCB analysis (State Laboratory of Hygiene, 1989). A

               schematic of this process is shown in figure 5-2 and a

               summary of the method used is given below.

                   The sample was removed from the mason jars, placed on

               aluminum foil and allowed to air dr
                                                  ,y until the sample had

               less than 30 percent moisture but at least 10 percent

               moisture. To insure that a homogenized representative

               sample was used, the sediment was sieved through a number 10

               sieve and the residuals, predominantly twigs and shell

               fragments, discarded. Ten grams of the sediment was dried

               overnight at 120 degree C to determine the moisture content

               of the soil.

                    Fifty grams of the sediment was placed in an acetone-

               washed filter thimble which was placed into an acetone-

               washed soxhlet.    Three hundred ml of a 50/50 hexane/acetone

               solution was poured into a soxhlet flask. Before the

               soxhlet, flask, and distillation column were assembled,

               granular copper was added to the soxhlet flask to remove any

               sulfur interference and to insure uniform boiling. The

               sediment was soxhlet-extracted for 8 hours. The temperature

               was set so that the solvent would cycle about five to eight

               times an hour.

                     Once the extraction apparatus had cooled, the sediment

               was discarded and the acetone-hexane solution containing the









               Figure 5-2 Analysis of PCBs in Sediment


                         Air dry sediment on
                              benchto                  Greater
                                     I                   than
                                Deter mi ne             30 %
                                moisture
                                     I
                        Soxhlet extract 50 g in
                     60/50 Hexane/Acetone - 9 hrs


                           Add Iso-oct and conc
                              Remove water



                            Florisil Fractionation
                                     I

                                Concentrate




                          Silica gel Fractionation


                      Analyze by GC-ECD for PCBS







  0

                                                                                                            47












  9














  is




















                                                                          48

                PCBs was placed in a warm water bath and concentrated to

                approximately two ml under a gentle stream of filtered air.

                The sample was diluted with approximately ten ml of iso-

                octane and again concentrated to remove any remaining

                acetone. Enough sodium sulfate was added to the sample to

                adsorb any water remaining in the beaker after the blowdown

                step.

                    Florisil columns were prepared by pouring hexane into a

                twenty ml column until it reached the base of the bulb.

                Next the column was filled with two grams of sodium sulfate,

                followed by 22 grams of Florisil and then another two grams

                of sodium sulfate. The hexane was drained to the top of the

                first sodium sulfate layer and the eluant discarded.    The

                sample was pourbd into the column and eluted at a rate of

                five ml/minute. When the sample reached the top of the

                sodium sulfate layer, the beaker that had contained the

                sample was rinsed with a small amount of.hexane and added to

                the column. After the first rinse reached the sodium

                sulfate layer., a second rinse was added to the column. The

                sample was eluted with 200 ml of a 96/4 percent hexane/ethyl

                ether solution.

                    When all of the solution was eluted, the sample was

                concentrated in a warm water bath under a stream of air to

                approximately five ml before silica gel fractionation. The



















                                                                           49

                silica gel columns were prepared by filling a column (10 mm)

                to the base of the bulb with hexane, adding two grams of

                sodium sulfate, five grams of deactivated silica gel and

                then another two grams of sodium sulfate. Best results were

                found when the silica gel was deactivated with five percent

                distilled water and allowed to equilibrate for an hour in a

                sealed Erlenmeyer flask.

                     The sample was eluted by the same procedure used for

                the florisil fractionation with the following two

                exceptions: the flow rate was 1-2 ml/min and the sample was

                eluted with 50 ml of hexane.   When all of the hexane had

                eluted through the column, the sample was concentrated and

                the solvent switched from hexane to iso-octane. Next the

                sample was diluted to 50 ml and screened on the packed

                column to determine if further dilution (or concentration)

                of the sample was necessary before performing high

                resolution gas chromatography.



                5.3.2. Fish

                     The procedure for extracting PCBs from fish was similar

                to that used for sediment and for water with the exception

                that a gel permeation column was necessary to remove fat

                which would interfere with high resolution gas

                chromatography. The procedure used was that of the State



















                                                                          50

                Laboratory of Hygiene (State Laboratory of Hygiene, 1989)

                and is shown schematically in figure 5-3.

                     The entire sample, weighing approximately 200 grams was

                ground up in a high speed blender with dry ice to homogenize

                the sample. The dry ice was allowed to sublime overnight.

                Ten grams of the sample was mixed with 60 grams of sodium

                sulfate to remove any moisture in the sample and then the

                sample was poured into a 20 mm i.d. chromatographic column

                containing two centimeters of sodium sulfate.

                Dichloromethane (230 ml) was used to elute the sample at a

                rate of 5 ml/min. once the dichloromethane was through the

                column, the sample was concentrated to five ml in a warm

                water bath under a gentle strdam of filtered air.

                Cyclohexane was added to the sample until a.final volume of

                ten ml was reached. A two ml aliquot was taken from the ten

                ml sample and was weighed. The solvent was evaporated from

                the sample and reweighed to determine the percent fat in the

                sample.

                     To remove the fat from the sample, a gel permeation

                column (GPC) was used. GPC  is based on the principle of

                size exclusion. Larger molecules, such as fats move

                relatively quickly through the column; however, smaller

                molecules such as PCBs, DDE and toxaphene, move more slowly

                because they permeate the multitude of smaller channels in









                 Figure 5-3 Analysis of PCBs Jn Fish


                            Homogenize fish sample
                            Grind sample w/dry ice
                                    I
                         Mix sample w/sodiurn sulfate
                         Extract with dichloromethane
                                Concentrate



                                               Determine
                                                 %  fat




                              Gel Permeation
                                 Cleanup


                            Silica Gel Fractionation
                                    I
                          Analyze by GC-ECD for PCBS


















                                                                            52

                the gel which are not available to the larger fat molecules.

                Five ml of each sample was injected onto the GPC. The first

                100 ml of eluate to come off the column was discarded

                because it contained the larger fat molecules. The second

                160 ml was collected as this fraction contained the PCBs and


                was concentrated to five ml.

                    Silica gel fractionation was performed using the same

                procedure as that for sediment: five grams of silica gel (5%

                deactivated with water) and 50 ml of hexane for the eluant.

                After silica gel fractionation, the sample was concentrated

                to five ml in iso-octane and diluted if necessary before

                analyzing on the gas chromatograph.




                5.3.3. Water

                     PCBs were extracted from the water using the

                filterhead-resin bed method. Although a brief summary of

                this procedure is given below, a more detailed discussion

                can be found by Marti (1984), Crane (1986) and the State

                Laboratory of Hygiene (1989).   Figure 5-4 shows a schematic

                of the procedure used.

                    After the samples were brought back to the lab, the 19-

                liter beverage canister containing the water sample was

                connected to a tank of ultra pure nitrogen gas by a teflon

                hose and also connected to the filterhead by copper tubing.









                 Figure 5-4 Analysis of PCBs in Water


                               Pressurize cannister
                              Force sample through
                              Glass filter. and resin



                                 Place filter and
                                  resin in soxhlets
                                  Soxhlet-extract
                                     for 16 hrs


                       Liquid-liquid extraction w/hexane



                                        conc



                               Florisil Fractionation

                                          I
                              Silica Gel Fractionation

                                          I

                           Analyze by GC-ECD for PCBs







  9

                                                                                                            53













  0














  0



















                                                                            54

                An ashed glass filter was placed on the filterhead and

                wetted with distilled water before the filterhead was

                fastened securely in place. An XAD resin column was

                attached to the bottom of the filterhead. The canister was

                pressurized with approximately 5 psi of gas; causing the

                water to flow from the canister through the filterhead, the

                resin and into a carboy located below the filterhead. To

                maintain a flow rate of 200 ml/minute, it was necessary to

                have the gas pressure at 1 to 2 psi.

                    The resin columns were prepared by fitting a short bit

                of tubing with a clamp over the end of a glass column (3 cm

                by 17 cm).    The tubing was attached to a water aspirator to

                help pack the column. The column was then filled with

                distilled water. A small portion of glass wool was pushed

                into the column and any air bubbles driven out by tapping

                the wool with a glass stirring rod. Enough XAD-resin was

                poured into the column so that.the column had a resin bed

                depth of 14 cm. A small portion of glass wool was placed on

                the top. The packed resin remained in distilled water until

                just after the canister was pressurized whereupon the tubing

                and clamp at the end of the column were removed and sample

                water began to flow through the resin.

                    Once all of the water had been filtered, the volume of

                the filtrate in the carboy was measured and the filtrate


















                                                                          55

                discarded. Next the resin column was uncoupled from the

                filterhead, and inverted over a hexane-rinsed soxhlet.

                Acetone was added until the resin would flow into the

                soxhlet. Both glass wool plugs were added to the soxhlet.

                The column was then rinsed with 60 ml of acetone and 40 ml

                of hexane. Enough of the 60/40 acetone/hexane solution was

                added to insure a final volume in the soxhlet and flask of

                350 ml. After boiling chips were dropped into the flask,

                the flask and soxhlet were placed on a hot plate and

                soxhlet-extracted for 16 hours.

                   The filter was peeled off the filterhead, wrapped

                tightly and placed into a soxhlet. Then the beverage

                canister was wiped with a small bit of glass wool to remove

                any particles. The canister was rinsed three times with 50

                ml of a 60/40 acetone/hexane solution. The filterhead was

                also wiped with glass wool to remove particulates and both

                pieces of glass wool were placed in the soxhlet containing

                the filter. Three consecutive rinses of 50 ml of

                hexane/acetone were siphoned from the canister, through the

                filterhead and caught in a beaker. These rinses were added

                to the other rinses in the soxhlet containing the filter.

                Again enough 60/40 acetone/hexane solution was added to the

                soxhlet and flask until a final volume of 350 ml was

                reached. Boiling chips were added and the filter was


















                                                                            56

                soxhlet-extracted for 16 hours.

                    Since the filter, resin and glass wool contained water,

                all of the samples needed to be liquid-liquid extracted

                after soxhlet-extraction. The sample was poured into a 500

                ml separatory funnel and extracted three times with 75 ml of

                hexane. Next the sample was placed on to a rotovap to

                concentrate the sample to approximately 30 ml whereupon it

                was transferred to a water bath and concentrated under a

                gentle stream of air to ten milliliters. About 20 ml of

                iso-octane was added and the sample was concentrated again.

                This step was repeated for all of the samples.

                    Once the samples were concentrated to approximately five

                ml, they were ready for florisil fractionation and silica

                gel fractionation.   The general procedure for preparing and

                eluting the compounds was the same as that for sediments;

                however, for florisil fractionation, only eight grams of

                florisil were needed for the column and only 50 ml of 94/6

                hexane-ethyl ether was needed for the eluate. The same

                procedure was used for silica gel fractionation.

                    All samples were concentrated to five ml after

                fractionation and screened on the packed column to determine

                if further concentration of the sample was necessary before

                analyzing the sample on the high resolution gas

                chromatograph.

















                                                                          57



               5.4. Gas Chromatography

                   High resolution gas chromatography was used for the

               analysis of PCBs in all matrixes. Table 5-1 contains the

               operating conditions for the three machines used. All of

               the samples were first screened on the packed column, and

               then run on the Hewlett Packard (HP) 5880. Confirmatory

               analysis of congeners was done on the HP 5890.

                   The HP 5880 was calibrated before each run using a 0.61

               gg/g standard containing Aroclor 1232, 1248, 1262 at 0.25

               mg/l, 0.18 mg/l, 0.18 mg/l respectively; the HP 5890 was

               calibrated with a 1.22 gg/g standard of similar composition.

               No more than ten samples were run in a single batch and

               periodically a standard was run at the end of the batch to

               assure that later samples were correctly identified. Each

               sample was spiked with internal standards 30 (12.5 ng) and

               204 (13.8 ng) to correct for variability in injection sample

               size and to correct for drift in the rention times of the

               congeners. Ten percent of the samples were run in duplicate

               to assess the precision of the analysis. A matrix spike for

               each batch of samples was run (a single batch typically

               contained 10 samples) to assess the accuracy of the

               analysis.


















                                                                            58





                                       Table 5-1
                  operating conditions for the Gas Chromatographs


                Hewlett-Packard 5880  Gas Chromatograph
                    60 m DB5 column,  0.2 mm ID
                    Hydrogen carrier  gas, Nitrogen makeup gas
                    Electron Capture  Detector at 320 degrees C
                    Oven temperature  program
                        @90 C -- 0.5  min
                         90   150 C at 10 C/min;
                         150   220 C at 1 C/min;
                         220   270 C at 3 C/min, hold for 2 min
                    Injection size of 3A1

                Hewlett-Packard 5890 gas chromatograph
                    60 m DBl column, 0.2 mm ID
                    Hydrogen carrier gas, Nitrogen makeup gas.
 0"                 Electron Capture Detector at 320 degrees C
                    Oven temperature program, same as HP 5880
                    Injection size of 2A1

                Hewlett-Packard 5790 gas chromatograph
                    6 ft x 4 mm column
                    Pacied with 4% SE 30/ 6% OV 210
                    Oven temperature of 235 degrees C
                    Injection size 2gl


                   Confirmatory analysis was done for approximately ten

                percent of the samples using the HP 5890. Additional

                confirmatory analysis was done by State Lab of Hygiene

                personnel on a mass spectrometer for specific congeners.

                    Limits of dectection were available for each method from

                the State Laboratory of Hygiene (State Laboratory of

                Hygiene, 1989). Limit of dectection is defined as the

                response that is three times the standard deviation of the


















                                                                            59

                background noise.   Congener values which fell below the

                limit of dectection were not reported.   Limits of

                dectection for each method are included in Appendix A.    In

                addition, congeners which deviated from the standard

                retention time by more than a tenth of a minute were not

                reported. For congeners which coelute, this value was

                relaxed to a twentieth of a minute.




                5.5. Grain Size Analysis of Sediment,

                     Grain size analysis determines the percent of sand,

                silt and clay in a sediment sample. The larger particles

                such as pebbles and sand can be physically separated and

                classified using sieves having different sized apertures;

                however, the smaller particles such as clay and silt cannot

                be separated using sieves and therefore must be separated by

                gravitational methods (such as a sedimentation test).

                     For sedimentation tests, most often a slurry containing

                fine particles and distilled water is made up and a

                hydrometer which measures the density of a solution is

                placed in the slurry. At given time intervals, readings are

                taken. As the time of the analysis increases, the heavier

                particles settle out and the solution becomes less dense.

                     Sedimentation tests are based on Stoke's Law which

                describes the velocity of spherical particles of a known


















                                                                         60

               density in a solution of known density and viscosity
                      V    -4 - 'YW 6L
                            16TI
               where


                    v - velocity (cm/sec)

                      - the specific weight of a solid (g/cm)

                      - the specific weight of water (g/cii)

                      - the viscosity (g-sec/cm2)

                    D - the diameter of the particle

               The equation can be rewritten to solve for the diameter at

               any time since all other variables are known. (The velocity

               can be calculated indirectly by knowing the depth of the

               hydrometer and the time.)

                    Grain size analysis was preformed on 18 of the

               sediment samples. Because a large amount of fine material

               was recommended for the sedimentation test (Das, 1986 and

               Head, 1980), a small subset of the total samples (ten

               samples) was used for the sedimentation test because other

               samples contained insufficient quantities of sediment to do

               the test.

                    The method used for sieving was based on the American

               Society for Testing and Materials (ASTM), Standard Method

               for Particle Size Analysis, Method D422-63 (American Society

               of Testing and Materials, 1987). Slight modifications to

               this procedure were suggested by Norm Severson (personal


















                                                                           61

                communication, Department of civil Engineering, Madison, WI

                1989).

                     A summary of the method follows (for further detail,

                see ASTM D422). A sediment sample weighing between 100 to

                200 grams was placed in a No. 200 mesh ASTM sieve over a

                large basin. None of the sample was pretreated with

                hydrogen peroxide or hydrochloric acid as it was desired to

                determine the particle size distribution in an unaltered

                environmental sample. The hydrogen peroxide removes organic

                matter and the hydrochloric acid removes carbonate cement

                between particles.

                    The sample was rinsed with a steady stream of tap water

                (referred to as "wet-washing") until the water running out

                from the sieve was clear. Wet-washing is necessary because

                there is a tendency for the finer clay and silt particles to

                adhere to the larger particles when a sample is dry sieved.

                The turbid, murky water from wet-washing was collected in

                the basin and allowed to settle overnight. The water was

                siphoned off and the sample was dried, crushed to break up

                any aggregation of particles, and weighed.

                    Fifty grams of the fine sediment was used for the

                sedimentation test. A 150 ml of a deflocculating agent was

                added to the sediment to disperse large flocs of aggregated

                particles which may settle out in the absence of a


















                                                                           62

                deflocculating agent. The deflocculating agent replaces the

                divalent and trivalent cation with sodium and causes the

                clay micelles to repel (Hillel, 1982). The dispersing agent

                was 35.7 grams of sodium hexametaphosphate (commercially

                known as Calgon) and 7.94 grams of sodium carbonate in one

                liter of distilled water.

                    The sample was then mixed using a high speed mechanical

                stirrer for two minutes.   The slurry was transferred into a

                one-liter sedimentation cylinder and diluted with distilled

                water to the one-liter mark. After a rubber stopper was

                placed in the end of the cylinder, the cylinder was inverted

                sixty times in one minute. Immediately after the last

                inversion, a hydrometer (ASTM 152 H) was placed in the

                slurry and readings were taken at the following intervals: 0

                seconds, 15 seconds, 30 seconds, 1 minute, 2 minutes, 4

                minutes, 8 minutes, 15 minutes, 30 minutes, 60 minutes, I

                hour, 2 hours, 4 hours, 8 hours, and 24 hours. Corrections

                for temperature, meniscus level and density of the

                deflocculating agent were made to the hydrometer readings

                (see Das, 1986 for further detail).

                    The sample remaining on the sieve after being wet-

                washed was dried in an oven (120 degrees C) and sieved

                through a number 10 sieve and a number 40 sieve.   The

                former separates the large sand grains from the medium sand


















                                                                         63

               grains while the latter separates the medium sand grains

               from the fine sand grains (American Society for Testing and

               Materials, 1987).   A number 10 sieve is routinely used in

               preparation of sediment samples for gas chromatography

               analysis. Unfortunately, sediment samples were prepared for

               gas chromatography analysis before it was decided that the

               samples would be analyzed for grain size and subsequently

               the material that remained on the number 10 sieve was

               discarded before it was weighed. Since the sediment is a

               fine grained fluvial deposit, almost all of the sediment

               passed through.a number 10 sieve. In addition, most of the

               matter not passing was organic matter such as twigs,

               material which would not ordinarily be used in a grain size

               test.


















                                                                             64

                                      Chapter 6 Results

                    This chapter contains the results of grain size analyses

                as well as the concentration and distribution of the PCB

                congeners in sediment, water and fish samples collected from

                the Sheboygan River.



                6.1. Grain Size

                    In general, the sediment is comprised of fine-grained sand

                and silt interspersed with clay and organic debris. The grain

                size test results determining the sand, silt, and clay

                fractions are also given in table 6-1.

                    The sediment samples are composed predominantly of fine

                sand, although a few samples are comprised of almost equal

                percentages of fine sand, and silt and clay.   In general, the

                proportions of fine sand and fines (silt and clay) remain the

                same for different locations, reflecting the relative

                homogenity of the fluvial deposits.

                     The medium sand fraction is probably overestimated since

                this fraction was in part composed of organics such as leaf

                fragments, and grass.   Organic matter can be removed using

                hydrogen peroxide.   Hydrogen peroxide was not used in the

                pretreatment of samples since it was desired to measure the

                grain size in an unaltered state representative of field

                conditions.


















                                                                             65

                    In cases where there was sufficient sample, sedimentation

                tests were performed to separate silt from clay.          These

                results are summarized in table 6-1.   As can be seen from the

                table, the silt fraction is higher than the clay fraction in

                all but one of the samples and in many of the samples the

                silt fraction is almost twice the clay fraction. Again the

                variation of the percentages of silt and clay with location

                is small.



                6.2. Sediment Samples from the Sheboygan River

                6.2.1. Total Concentrations of PCBs in Sediment

                     The total PCB concentrations ranged from a high of 1590

                ppm found above the Upper Kohler dam to a low of 0.04 ppm

                found   above    the   Sheboygan    Falls   dam     (background

                concentrations).   Although there is considerable variation

                among the samples, in general total PCB concentrations

                decreased as one moves downstream away from the source.     The

                average value for each core is shown on a map in figure 6-1.

                Values for each individual 15 cm section of core are given in

                table 6-2.

                     As seen from the samples (A samples) collected above

                Sheboygan Falls Dam, the background concentrations are very

                low (an average of 0.05 ppm).    For the remaining cores, the

                top 15 cm. of the core, containing the sediment-water



















                                                                                 66






                          Table 6-1 Results of Sieved Sediment Samvles
                                Based an ASTM Definitionsa

                                       Sand Fraction           Fine Fraction

                 Sample    PCB Conc     Lfedi     Fine          Silt       Clay
                        (ppm, dry wt)
                                  (2.Omm-4254m) (425Am-75Am) (74Am-5Am) (<SAm)

                 Al-2       0.04        4.9         62.3          20.4       12.6
                 A2-2       0.02         8.2        66.9                        9.7


                 B1-1       8.5         6.4        64.8         18.9         9.9
                 B1-2     345.9         8.0        53.9         23.9        14.2
                 B1-3     334.2         8.8        59.0         19.3        13.1


                 Cl-3       9.8         5.3        47.3         28.4        18.4


                 D2-3     300.3         15.9       43.8                41.1*
                 D2-4       4.2         6.4        61.6                31.7*


                 E3-1       7.7         8.0        62.9                29.3*


                 TlA       50.3         8.9        69.4         19.8         9.5
                 T2B       76.9         5.4        76.4                18.4*
                 T2C       97.8         4.6        62.3         14.8        18.4
                 T2D        3.5         5.2        65.4         21.0         8.7
                 T3B       60.9         5.5        63.8         15.7        15.0
                 T3C        0.5         2.4        90.5                  6.8*
                 T3D        0.4         3.4        80.7                15.9*
                 T4B       12.5         5.4        85.0                  9.6*





                      Indicates combined silt and clay fractions
                 aAmerican Society of Testing and Materials, 1987








                                                                              .,Ooor



                                                                  kohler
                                                                             :7    EM
                                                           Above L.
                          Above Sheboygan                                     I    A
                          Falls Dam                               DI   88.9
                                                                 D2=   95.
                               = 0.044
                          Al
                                                    Above U. Kohle     Da
                                                   C1 =40.2
                           2   = 0.049                  =534.1
                                                      2

                                _Rochester Park
                                    =173.1




                                                                           J@3




                                                   12
                                             B2
                                                                                                                      E it
                                             83  = J62.3
                                             TI  =  24.1
                                                             Rochester Park
                                             T 2 =  49.3
                          --.2n Falls
                                             T3  = 27,
                                                      3.9
                                             T4  =  36.0






                                                      Figure 6-1 Map of Sheboygan River illustrating Average
                                                                   Sediment cor,e Concentrations
















                                                                                                                                 0               2n
                                                                                                                                                       N7


                                                                                                                                           r
                                                                                                                  it
                                                                                                                                              ll'- TIOt-All
                                                      .4-



                                                                                                                                                    -17  CJ3




                                                                  ,It






                                                                                                                           4L
                                                                                         Kiwanis Park

                                                             21
                                                                                                                                                         44@-t
                                                                                         El     1.3
                                                                                         E2=    2ol        E3
                                                                                                       LAID-

                                                                                                            I  at

                                                                                         21                               63


                                                                              jc
                                                                                                                                            I A-4 Z-    V


                                                                                                                                                I   j


                                                                                                                                                               J



                                                  a I
                                                            8      28,                                                 2?.
                                                                                          oll

                                                                                                                                all


                                                                                                           _4
                                                                                       a0i                                                                            K.r
                                                -61
                                                                                  IT.
                                                                                                           ;A
                                                                                                                                    IL,41
                                                                                                                           CP
                                                                                                                                         Lit
                                                                                                                                                                    A
                                                                                                                                               Xj

                                                                                                                                                   ev 'if i
                                                                                                                                                               sy "I


                                                            Figure 6-1 Map         of Sheboygan        River     (Continued)




















                                                                                 69


                     Table 6-2 Total PCB Concentrations in River Sediment
                                             (ppm, dry wt)

                 Sample                                   Concentration

                 Above Sheboygan Falls
                        Dam


                 Al-1                                          0.0354
                 A1-2                                          0.0176
                 Al-3                                          0.0381
                 A1-4                                          0.0866
                 Al-5                                          0.0916
                                                                        Ave. 0.044
                 A2-1                                          0.033
                 A2-2                                          0.0235
                 A2-3                                          0.0663
                 A2-4                                          0.1611
                 A2-5                                          0.0581
                                                                        Ave. 0.049


                 Rochester Park


                 B1-1                                          8.5
                 B1-2                                         345.9
                 B1-3                                         334.2
                 B1-4                                          3.6
                                                                        Ave. 173.1
                 B2-1                                         113.9
                 B2-2                                         143.1
                 B3 (composite)                               162.3
                                                                        Ave. 139.8
                 Above Upper Kohler Dam                                         -

                 C1-1                                          28.7
                 C1-2                                           8.6
                 C1-3                                           9.8
                 CI-4                                         113.6
                                                                        Ave. 40.2
                 C2-1                                           3.9
                 C2-2                                          12.4
                 C2-3                                         1585.8
                                                                        Ave. 534.0
                 Above Lower Kohler Dam


                 Dl-1                                           4.4
                 D1-2                                         117.9
                 D1-3                                         144.5
                                                                        Ave. 88.9


















                                                                                70



                 Table 6-2 Total Concentration of PCBs in River Sediment
                 (cont.




                 Sample                                   Concentration

                 Above Lower Kohler Dam (cont.)

                 D2-1                                         8.2
                 D2-2                                         70.5
                 D2-3                                       300.3
                 D2-4                                         4.2
                                                                      Ave. 95.8
                 Kiwanis Park


                 El-l                                          1.9
                 E1-2                                          0.7


                 E2-1                                          2.1


                 E3-1                                          7.7

                 Rochester Park (Second Sampling Trip)

                 T1A                                          50.3
                 TlB                                          21.3
                 TlC                                           0.8
                                                                    Ave. 24.1
                 T2A                                          18.8
                 T2B                                          76.9
                 T2C                                          97.8
                 T2D                                           3.5
                                                                    Ave. 49.3
                 T3A                                        1053.7
                 T3B                                          60.9
                 T3C                                           0.5
                 T3D                                           0.4
                                                                    Ave. 278.9
                 T4A                                           8.6
                 T4B                                          12.5
                 T4C                                          26.8
                                                                    Ave. 16.0


















                                                                                 71

                 interface, had relatively low concentrations of PCBs (an

                 average of 21.4 ppm excluding sample T3A). The next two core

                 segments (15-30 cm and 30-45 cm) typically had the highest

                 concentrations (an average of 174.4 ppm), while the bottom

                 core segment     (45-60   cm)   had on average the          lowest

                 concentrations (an average of 2.4 ppm excluding sample C1-4).



                 6.2.2. Hamolog Distribution Patterns in Sediment

                      Samples from the first sampling trip (in which all sites

                 were sampled) could be divided into two distinct groups based

                 on their homolog patterns.     One pattern had an enrichment in

                 the lighter chlorinated congeners relative to the original

                 Aroclor patterns while the other pattern had a distribution

                 similar to the original Aroclor patterns.       This distinction

                 appehred to be a function of concentration.                Samples

                 containing PCB concentrations greater than 50 ppm. appeared to

                 be enriched with the lighter chlorinated congeners, whereas

                 those with less than 50 ppm PCBs were not.

                      Samples containing 50 ppm. or more PCBs had significantly

                 higher concentrations of mono and dichlorinated congeners

                 compared to the original Aroclors introduced into the river.

                 Using a multivariate t test, these samples were found to be

                 statistically different in composition from Aroclor 1248,

                 Aroclor 1254,- and a 50/50 mixture of Aroclor 1248 and 1254


















                                                                             72

                All p values were less than 0.05. A p value smaller than 0.05

                means that the samples are significantly different at the

                ninety fifth percentile confidence interval. Thare is less

                than a f ive percent chance that the sample came from a

                distribution that contained the Aroclor mixes.      Figure 6-2

                and 6-3 show the homolog distribution of sediment samples with

                PCB concentrations above 50 ppm from the first and second

                sampling trips.     Although the two sets of data are f rom

                different sampling  trips, there is no statistical difference

                between these samples (p=0.8870, using a test of two means).

                     In samples containing PCBs concentrations less than 50

                ppm from the first sampling trip (sites B,C,D,E), the homolog

                patterns slightly resembled the original Aroclors introduced

                into the river; however, they were statistically different

                from Aroclor  1248, Aroclor 1254 and a 50/50 mix 6f Aroclor

                1248 and 1254 (using a multivariate t test p values were found

                to be less than 0.05 for all comparisons).            This is

                illustrated graphically in figure 6-4.

                     There were too few samples with concentrations less than

                50 ppm from the second sampling trip (Rochester Park site, T

                samples) to.use a multivariate t test.     These samples were

                found to be statistically different from the di,   tri, tetra,

                hepta and octa homolog groups of the original Aroclors.using

                a univariate t test ( p < 0.05).    This is illustrated in










                                                                                       Figure 6-2 Homolog Distribution in
                                                                                 Sediment (Conc.> 60 ppm, Sites B,C,D,E)
                                                             60  Wt. percent


                                                             50 -                                                                                          .... . .. .. .. .. . . .. ........




                                                                                                                                 .......................   .................
                                                             40   . ......        .. . ... ...                  . .....


                                                             30-                                        . . .....                              ............. .................. ...............




                                                                                                          .... .......             ..........
                                                             20--                    . . ........                                                        ...................... ..............



                                                             10   . ................... . ........ ........... ....                                          .................. .... ..............


                                                               0
                                                                   Mono            Di          Tri       Tetra Penta Hexa Hepta                            Octa         Nona
                                                                                                              Homolog Group

                                                                                            Sample           EM Std 1248                       Std 1254

                                                       (Ave. of 11 samples)









                                                                                    Figure 6-3 Honnolog Distribution in
                                                                                    Sediment (Conc. > 50 ppm, Site T)

                                                               Wt. percent
                                                          60



                                                          50   -                 . .... ...... .




                                                                                                                                               ..............................
                                                          40                     . . . .... ... . ......... ..



                                                                                                            ...........                 . ......... .....................
                                                          30   -


                                                          20   . .. .. ... ............. .                   ........... ..... ....... ....................  .......................................... ... ........ ... .....................


                                                          10          .............
                                                                                                                                           ...........  ........................ ..............



                                                            0                                                ...
                                                                Mono            DI          TO        Tetra        Penta        Hexa Hepta Octa                       Nona
                                                                                                             Conc. (ppm)

                                                                                         Sample          M Std 1248                  ED Std 1254

                                                    (Ave. of 5 samples)











                                                                                 Figure 6-4 Homolog Distributio                                 n in
                                                                           Sediment (Conc.< 60 ppm, Sites B,C,D,E)

                                                           Wt. percent
                                                       60


                                                       50-




                                                                                                                           ................-.................. ..........-..................................
                                                       40                          . .. .. . ........ .. ....



                                                       30--                                                    ... . ........   .... ..... .. .......... ..... .......... ...................... .......... ... .............



                                                       20              .. ....                            .......... ..... .......... .................. ........... ........................................ ...............




                                                                                                                                            ................ ........................ ........... . ...-
                                                       10                                                                  ............



                                                         0
                                                             Mono            DI          Tri       Tetra Penta               Hexa Hepta Octa                      Nona
                                                                                                        Homolog Group

                                                                                      Sample           M Std 1248                 EEI Std 1254

                                                 (Ave. of 14 samples)

















                                                                                 76

                 figure 6-5. Interestingly, the samples from the second

                 sampling trip containing less than 50 ppm (Rochester Park, T

                 samples) were very similar to the distribution of the more

                 highly concentrated samples from the same area (p= 0.8875).



                 6.2.3. Most Prominent Congeners in Sediment

                     The most prominent congeners are listed in table 6-3.

                 These results are also presented graphically in figure 6-6 and

                 figure 6-7.    Relative to the original Aroclors, particularly

                 high concentrations of congeners 5/8 (congeners 5 and 8

                 coelute) were seen. The highest concentrations were found in

                 samples containing more than 50 ppm PCBs, 29.2 percent on

                 average. In contrast, samples containing less than 50 ppm of

                 PCBs, contained only 11.4 percent on average. Areas of high

                 concentrations of PCBs, such as Rochester Park, had high

                 concentrations    of congener     5/8   throughout    the    core,

                 independent of concentration in each individual segment of

                 core. Samples from Rochester Park had on average 24 percent

                 congener 5/8 whereas all other sites had on average 11.5

                 percent. Congeners 4/10, 16/32, 17, 24/27, 47/48 and 26 were

                 also abundant in comparison to the original percentages in the

                 Aroclors.      Although 28/31, 52, 66/95, and 77/110 were

                 prominent congeners, they were present in diminished

                 quantities in the sample relative to the original quantities










                                                                                          Figure 6-6 Homolog Distribution in
                                                                                          Sediment (Conc. ( 60 ppm, Site T)

                                                                   Wt. percent
                                                              60


                                                              so-                                                                                   .....................



                                                                                                                   .............................                 ................
                                                              40


                                                              30    -                                      ...... ....   ............                           ................


                                                              20    -      . .. ....     ..... .. ..      .....     .......... .............           .............



                                                                                       ...........
                                                                                                                                                             .............. ..........
                                                              10


                                                                                                                                                           1771
                                                                0
                                                                     Mono            DI           TO         TO tra,      Penta Hexa Hepta Octa                                 Nona
                                                                                                                     Conc. (ppm)

                                                                                              Sample             EM Std 1248                 E:DStd V54

                                                       (Ave. of 9 samples)


















                                                                                   78






                             Table 6-3 Most Prominent congeners in sediment
                                        (Weight Percent of Total)

                  Congener        1st trip        2nd trip          Aroclor Aroclor
                             samples >50 ppm (R.Park only)            124,a     1254a
                                                samples >50 ppm

                  4/10             2.1                                1.6       0.03
                  5/8             29.9                 27.4           2.1       0.06
                  17               9.2                 10             1.2       <0.1
                  16/32           10.1                  9.8            1.9       <0.1
                  47/48            7.7                 7.2            4.0       0.4
                  24/27            2.8                 3.4            0.18      <0.1
                  28/31            6.3                 6.6            17.7      0.6
                  66/95            3.7                 4.1            9.0       7.6
                  77/110           2.5                                2.5       9.2
                  26               2.0                 2.9            0.4       <0.1
                  52                                   2.4            4.4       4.2
                  49                                    2.0            3.0       1.0





                  Congener 1st trip           2nd trip            Aroclor    Aroclor
                          samples <50 ppm (R. Park only)           1248a      1254a
                                             samples <50 ppm


                  5/8             6.3                  17.3         2.1       0.06
                  17                                   10.3         1.2       <0.1
                  16/32           4.1                  11.3         1.9       <0.1
                  47/48           6.3                   8.3         4.0       0.4
                  28/31           8.6                   9.9        17.7       0.6
                  66/95           7.9                   4.0         9.0       7.6
                  77/110          4.1                   3.0         2.5       9.2
                  26              3.1                               0.4       <0.1
                  52              4.3                   1.8         4.4       4.2
                  49              4.1                   2.2         3.0       1.0
                  41/64/71                              2.9         0.7       <0.1

                  a Manchester,   1988









                                                                          Figure 6-6 Most Prominent Congeners
                                                                                      in Sediment (Sites B,C,D,E)

                                                       Wt Percent
                                                 35-


                                                 30   . ................ .... ..-.......... ........................ ..... ........ ........... . .... .. . . .... . .. ....... ... ... . .. .... . ..... .. . ...... . ... . .-.... ...... ..I...... .. . ....... ... ... .... .. ............


                                                 25   . ...... .............. ............ ............                                   . .. . ... ... .................. ..... .. ..... .. I - -- ... - ... - -


                                                 20   . ....................... ............................................ . ..... .. ...... .. . . . .. . . .. .. . .......... . ....... ... .. . ..... .I... ..... .. .... ... .. .................... .. .. .. . . ... ......... ........ . ..


                                                 is   . ............  .......... ............. ...... ....I........... . ... ...... .. . .... ........ . ... ... .. . . ........ .I - - .. - .. .1 .. . .......... .. .. ............ .. . ............................... .. . .... ..... .. ... .


                                                 10   . ...........  .......... . ............. ................. . ............. . ...... .... . .... .. ............... ... ........... ........... ............. .. ..... ... . .......... ................. ..................... . ......... ....  ...........


                                                      . . ....... ............ ...............  ............ .................. ................... ......................................................



                                                   0-
                                                         4/10       6/8 16/32            17     24/27 26 28/31               47/48        49       62      66196 77/110
                                                                                                           Congener


                                                               Samples         60 ppm                Samples < 60 ppm                      60/60 Std       1248/1264


                                             (Ave of 11 Samples, <50 ppm
                                             Ave of 14 Samples, <50 pprn)









                                                                             Figure 6-7 Most Promin                            ent Congeners
                                                                                                in Sediment (Site T)


                                                          Wt Percent
                                                     30


                                                     25   . ... .............................. ......-........ ...I........ . .. ............. . ... ... .... .. ... . .... .. .. ........ .. ... ........... ............... ...... .......... ..... ............... .. ......... ........ .................... .....


                                                     20   . ........ I........................................................  ........................................................................................................................................................................................



                                                          . ......................................  ..................................................................................................I..........I.................................... I.  ................................... ....................................



                                                     10   . ................ ......................  ................................................................................................................................................................................ ....................................



                                                                  ......... ...... ... .................................... ................. ..................................................................... ................ . . . ..




                                                        0
                                                             5/8        17     16/32 24/27 26               28/31 47/48           49 41/64/71 62             66/96 77/110
                                                                                                        Homolog Group


                                                                            50 pprn                   > 50 ppm                     50/50 Std 1248/1254


                                               (Ave of 6 samples >60 ppm
                                               Ave of 9 samples <60 ppm)


















                                                                                81


                in the Aroclor mixtures.




                6.2.4. Accuracy, Precision and Confirmatory Analysis for

                        Sediment Samples

                     A duplicate sample and a spiked sample were included in

                every batch of samples run. Sediment was spiked with an

                Aroclor mixture of Aroclor 1232, 1248 and 1262 at a level of

                0.61 ppm.    The recoveries for the spikes* are listed in the

                table below and are calcluated by summing the individual

                congener concentration.      The recoveries varied from a high

                of 108.1 percent to a low of 81.8 percent with an average

                recovery of 92.5 percent.



                     Table 6-4 Percent PCB Recovery from Spiked Sediment

                Run  1     81.8
                .Run 2     108.3
                Run  3     81.9
                Run  4     95.4
                Run  5     94.4
                Run  6     94.5
                Run  7     89.2
                Run  8     94.4




                     Values for duplicate samples are given in table' 6-5.

                Confirmatory analysis was performed on approximately 25

                percent of the samples (15 samples out of a total of 59

                samples) using a DB-1 column. Good correlation was found

                between the two data sets (correlation coefficient of 0.983).















                                                                                82

                     Since many of the samples had very high concentrations of

                congener 5/8, additional confirmatory analysis of this

                particular congener was sought. Six of the samples,

                containing high concentrations of PCBs (342.8 ppm on average)

                and high concentrations of congener 5/8 (43.7 percent on

                average), were analyzed using an Electron Impact Gas

                Chromatography Mass Spectrometer. All of the spectra





                             Table 6-5 Duplicate Sediment Samples

                Duplicate        Results           Mean     Percent Agreement


                1                345.9, 414.8    380.4        90.9
                3                4.6, 5.2        4.9          93.9
                2                7.0, 8.2        7.6          92.1
                7                76.9, 69.3      73.1         94.8
                6                0.53, 0.49      0.51         92.2
                4                97.8, 134.8     116.3        84.1
                5                4.4, 5.1        4.75         92.6
                8                20.74, 20.92    20.83        99.6




                confirmed that the previously identified congener 5/8 peak

                was indeed a dichlorinated biphenyl. The sample spectra were

                compared to a library of known spectra compiled by the

                National Bureau of Standards.     A computer algorithm designed

                to match the most significant peaks determined the agreement

                between the sample and known standard.        Good agreement was

                found for all spectra as indicated by the percent match which



















                                                                                   83

                 varied between 83.2 and 96.6 percent with an average match of

                 91.8 percent.



                 6.3. Sediment Samples from the Harbor

                        Three cores from the Sheboygan Harbor were obtained from

                 the EPA and analyzed to determine the PCB congener

                 distribution.     The cores were located near the mouth of the


                 harbor.       Two cores were taken at a sediment depth of

                 approximately 3 0. 5 cm (1 ft)     The remaining core ranged from

                 the surface-water interface to a depth of 50.8 cm (20 inches)



                 6.3.1. Total PCB Concentrations in Harbor Sediments

                        Table 6-6 contains boring depths of the core and the

                 concentration of PCBs found in each segment of the three


                 cores.



                 6.3.2. Homolog Distribution in Harbor Sediment


                       The homolog distribution are similar in distribution to

                 the original Aroclor mixtures (see figure 6-8).           They are

                 however statistically different (p < 0.05).       In addition, the

                 patterns appear to be similar to the homolog patterns seen for

                 sediment samples containing less than 50 ppm PCBs.


















                                                                               84








                    Table 6-6 Total PCB Concentrations in Harbor Sediment

                Core                                 PCB Conc. (Ppm, dry wt.)


                H18
                     0-1.3 cm                                   36.3-
                     1.3-5 cm                                   15.7
                     5-10 cm                                    83.6
                     10-15 cm                                   30.7
                     15-20 cm                                    3.5
                     20-31 cm                                    1.4
                     31-41 cm                                    0.3
                     41-51 cm                                    0.9


                Hll
                     31-41 cm                                    3.5
                     41-51 cm                                   27.7


                H15
                     20-31 cm                                    91.2










                                                      Figure 6-8 Homolog Distribution
                                                              in Harbor Sediment

                                       Wt. percent
                                     60


                                     50 -                                 ...... ......... .. ...


                                     40-


                                     30 . ......


                                     20-




                                                                           . . . ........ . . ... .................. ................... ........... . ......
                                     10-        . . .. ......


                                      0
                                         Mono     DI      TO    Tetra  Penta Hexa     Hepta Octa      Nona
                                                                  Hornolog Group

                                                    sarnpie   20 Arocior 1248          Aroclor 1254

                                 (Ave. of 11 samples)




















                                                                                 86

                 6.3.3. Most Prominent Congeners in Harbor Sediment

                       The most prominent congeners are listed in table 6-7.

                 Most of these congeners are also found in high concentrations

                 in the original Aroclor mixtures 1248 and 1254 as noted in the

                 table below.    The most prominent congeners are also presented

                 graphically in figure 6-9.



                  Table 6-7 Most Prominent Congeners found in Harbor Samples
                             (Weight percents)

                 Congener        Samnle     Aroclor 1248         Aroclor  12544

                 28/31            13.9                17.7                0.6
                 47/48            7.4                 4.0                 0.4
                 66/95            7.2                 9.0                 7.6
                 17/110            @6.6               2.5                 9.2
                 49               5.5                 3.0                 1.0
                 52               5.6                 4.4                 4.2
                 26               4.9                 0.4                 <.1
                 41/64/71         4.3                 0.7                 0.4.
                 44               3.6                 5.2                 3.0
                 16/32            3.7                 1.9                 <.l

                 a Manchester, 1988



                      Congener 28/31 is the most abundant congener in the

                 harbor samples.     It is also the most abundant congener in

                 Aroclor 1248.    Congeners 47/48, 49, 52, 26, 41/64/71, and

                 16/32 are enriched in the harbor sediment relative to their

                 concentrations in the Aroclors. The only congener found in

                 diminished quantities relative to. the Aroclor mixtures was

                 congener 66/95.








                                                                           Figure 6-9 Most Prominent Congeners
                                                                                               in Harbor Sediment


                                                        Wt Percent
                                                   16


                                                   14   . ... .................... ... ........ ... . .......... .......... . .......... .... . ... ............ ... .. ........ ........ ... . .. . . . ................ . ...... . ...... .. .... ... . .


                                                   12   . ........................ ...... ......... .. ...... ...... .... . ... ......


                                                   10   . ........................ . ........................... ............ ............. ..........................................  .................... ....................................................................


                                                                                                                          ............... . ........
                                                    8   . ........................ ... ........................ .......... ............. ...........................  ............................. ...................................................................................
                                                                                       .....                                          .....


                                                                                       .....                                          .....
                                                                        ................             .................................  ............
                                                    6   . ........................ ... .....                                                      .............. .................................. ............
                                                                                       ....                                           ....        ....
                                                                                       .....                                          .....       .....
                                                                                       ....                                           ....        ....
                                                                                       .....                                          .....       .....
                                                                                       ....                                           ....        ....
                                                                                       ...                                            ....        ....
                                                                                       .....                                          .....       .....
                                                                                       ....                                           ....        ....
                                                                                       .....                                          .....       .....
                                                                                       ....                                           ....        ....
                                                                           ...............        .............. ...............                                  ....   ............
                                                    4   . .. ..................... .
                                                                                                                          .....       ....        ...
                                                                                       .....                              ....          ...
                                                                                       .....                              ....        .....
                                                                                       ....                               .....       ....          ...      .....
                                                                                                                          ....        ....        .....      ....
                                                                                                                                      ....        .....      .....
                                                                                       ....                               ....        .....       .....      ....
                                                                                                                          .....       ....          ...      .....
                                                                                                                          ....        .....       .....      ....
                                                              .............. ................                                                                     .. ..........
                                                    2
                                                                                                                          ....        .....       .....


                                                    0
                                                         16/32          26       28/31 47/48               49          52       66/95       77/110         44       41/71
                                                                                                           Congener


                                                                                  Harbor Sediment                            50/50 1248/1254


                                             (Ave of 11 Samples)


















                                                                             88

                6.3.4. Precision and Accuracy for Harbor Samples

                    The recovery from spiked sediment was 101.6 percent and

                was calculated by summing the individual congeners.         The

                results of the duplicate analysis preformed on the samples

                were 1.4 ppm and 1.5 ppm.



                6.4.   Water Samples from the Sheboygan River

                6.4.1. Total PCB Concentrations in Water

                     The water samples are divided into particulate and

                dissolved concentrations based on an operational definition.

                The portion of the sample that was able to pass through the

                0.7 micron filter was defined as the dissolved fraction and

                the portion that remained on the filter was the particulate

                fraction. The particulate and dissolved concentrations are

                given in table 6-8.

                   The background concentrations were low, 0.007 and 0.003

                ppb for the particulate and dissolved fraction, respectively.

                The particulate concentrations in the non-background samples

                varied from an average low of 0.162 to an average high of

                0.502 ppb. The lowest particulate concentration in the water

                occured at Rochester Park, the site of the highest average

                sediment PCB concentrations, 173.1 and 139.8 ppm. This would

                be expected since the river water has been in relatively short

                contact with the contaminated sediment. The highest


















                                                                               89

                      Table 6-8 Total PCB Concentrations in Water (Pnb)

                Site                                   Concentration


                Above Sheboygan Falls Dam
                      (Background)

                Dissolved
                     Water sample 1                      0.003
                Particulate
                     Water sample 1                      0.007

                Rochester Park

                Dissolved
                     Water sample 1                      0.028
                     Water sample 2                      0.047
                Particulate
                     Water sample 1                      0.162
                     Water sample 2                      0.301

                Above Upper Kohler Dam


                Dissolved
                     Water sample 1                      0.190
                     Water sample 2                      0.204
                Particulate
                     Water sample 1                      0.464
                     Water sample 2                      0.432

                Above Lower Kohler Dam


                Dissolved
                     Water sample 1                      0.152
                     Water sample 2                      0.179
                Particulate
                     Water sample 1                      0.508
                     Water sample 2                      0.452

                Kiwanis Park


                Dissolved
                     Water sample 1                      0.071
                     Water sample 2                      0.069
                Particulate
                     Water sample 1                      0.275
                     Water sample 2                      0.327
















                                                                                 90

                 particulate concentration in the water was above the Lower

                 Kohler dam which had average sediment core concentrations of

                 88.9 and 95.8 ppm.

                     The dissolved concentrations varied between 0.028 and

                 0.204 ppb.     The lowest concentrations were again found at

                 Rochester    Park,   the   site    of   highest   PCB     sediment

                 concentrations.    The highest dissolved concentrations were

                 found above the Upper Kohler dam which had sediment

                 concentrations of  40.2 and 534 ppm, (the average of 534 ppm was

                 disceptively high   because the highest single 15 cm sediment

                 concentration was   found in this core, the remainder of the

                 core had low concentrations of PCBs).



                 6.4.2. PCB Homolog Distribution in water

                     The homolog groups for the dissolved and particulate

                 fractions are illustrated in figure 6-10.       In the dissolved

                 fraction, there is a dramatic increase in the lighter

                 chlorinated congeners relative to the original Aroclor

                 mixture, particularly the mono, di and trichlorinated

                 congeners.    Correspondingly, there is a decrease of the

                 heavier chlorinated congeners, notably the penta, hexa, hepta

                 and octachlorinated homolog groups.

                      In comparison to the dissolved fraction, the particulate

                 fraction contains decreased quantities of the mono, di and










                                                                                                   Figure 6-10 Homolog Distribution
                                                                                                                                in Water


                                                                         Wt. percent
                                                                   60




                                                                                                                                                                                      ...................-
                                                                   so -



                                                                                                                                 ................ ...                       ...........  ............
                                                                   40




                                                                                                                . .............. . ................ . .....           ..........
                                                                   30-




                                                                                                                            . ...............                         ..........       .............
                                                                   20    . ........... ..... ...



                                                                              ...... .....                                                                              ........... ... ............. ...............
                                                                   10
                                                                      0  IL
                                                                           Mono             DI            TO          Tetra Penta Hexa Hepta Octa                                            Nona
                                                                                                                           Homolog Group


                                                                                   Dissolved                       Particulate                       Std 1248                       Std 1254


                                                             (Ave. of 8 samples)

















                                                                             92

                trichlorinated congeners; however, there is still an

                enrichment of these lower chlorinated congeners relative to

                the original Aroclors. In general, the distribution is much

                more similar to the original Aroclors than the dissolved

                fraction.

                     Not surprisingly, the homolog distribution for the

                dissolved fraction was not found to be statistically similar

                to the original Aroclors or a 50/50 mix of the original

                Aroclors ( p < 0.05 for univariate t tests of each homolog

                group). The particulate fraction was also not statistically

                similar to the original Aroclors with the exception of the

                monochlorinated congeners and the nanochlorinated congeners

                which are in very small concentrations in both the sample and

                in the Aroclors ( p < 0. 05 for univariate t tests of each

                homolog group).



                6.4.3. Most Prominent Congeners in Water

                     The most prominent congeners found in water are listed

                in table 6-9 and shown graphically in figure 6-11.

                 Congeners 1, 4/10, 5/8, 17, 16/32, and 41/64/71 were in

                higher concentrations in the water samples than in the

                original  Aroclors.         Congener 70/76    was   in    lower

                concentrations than the Aroclors but still remained one of the

                most prominent congeners in the' sample. The remaining


















                                                                                  93




                         Table 6-9     Most Prominent Congeners in Water
                                     (Dissolved and Particulate Fraction)

                 Congene     Diss.        Part. Aroclor 1248   a  Aroclor 12548

                 1            3.4                          <0.1           <0.1
                 4/10         2.9                          1.59           0.03
                 5/8          26.0           7.6           2.1            0.06
                 17           7.9            3.6           1.2            <0.1
                 16/32        3.6                          1.9            <0.1
                 28/31        10.7           9.7           17.74          0.6
                 47/48        3.4            6.1           4.0            0.4
                 41/64/71     2.8            4.7           0.74           0.4
                 52           4.0            3.5           4.4            4.2
                 66/95        3.3            7.8           9.0            7.6
                 70/76                       4.2           7.1            4.4
                 77/110                      5.9           2.5            9.2
                 44                          3.1           5.2            3.0

                 a Manchester, 1988




                 congeners were of intermediate value..



                 6.4.4. Accuracy, Precision and Confirmatory Analysis for Water

                      The recoveries from spiking water with an Aroclor mixture

                 of 1232/1248/1262 in acetone were 83.1 and 91.7.      All samples

                 with the exception of the background sample were run in

                 duplicate and are shown in table 6-8.

                     Confirmatory Analysis was performed on approximately

                 thirty percent of the samples (six samples) using a DB-1

                 column.   The correlation coefficient for the two data sets

                 indicate good agreement (Correlation coefficient = 0.977).










                                                          Figure 6-11 Most Prominent Congeners
                                                                                in Water


                                             Wt Percent
                                         30-


                                        '25 -      ......... ..........      ..........    ................... .


                                         20-


                                         15-                             .............. ...........-         ... .............. ..............


                                         10-         . .. .......                                           . . ... .. ..... . .. . ........



                                                                                    ....................... ............................-F-1
                                          5  . ........ .. . ............... .. ... . . . ......
                                           0-                                              Ifl                   w] N:@:,
                                                1    4/10 5/8     17 16/32 28/31    41/71 44    47/48 62 66/9570/7677/110
                                                                                 Congener

                                                 =Dissolved                Particulate           50/50 Std 1248/1254


                                      (Ave of 8 samples)















                                                                             95


                6.5.   Fish


                6.5.1. Total PCB Concentrations in Fish

                     The fish concentrations varied from a low of 1.3 ppm to

                a high of 22.0 ppm. These results are summarized in table 6-

                10.    The highest values were found in carp and channel

                catfish; both of which are bottom feeding fish and have high

                concentrations of fat.



                6.5.2. Homolog PCB Distribution Patterns in Fish

                     The average homolog distribution pattern is shown in

                figure 6-12 with the Aroclor 1248 and 1254 and in figure 6-13

                with the homolog distribution for the river sediment and the

                harbor sediment. Relative to the Aroclors, there was a slight

                increase in the mono and dichlorinated congeners in the fish

                samples, probably due to the significant concentrations of

                the mono and di chlorinated congeners in the water and

                sediment.

                     Although the fish samples were enriched in the higher

                chlorinated congeners and appeared to be more similar to the

                Aroclors than the water or sediment samples, the samples were

                not found to be statistically similar to Aroclor 1248, Aroclor

                1254 or a 50/50 mix of Aroclor 1248 and 1254 using a

                multivariate t test (p < 0.0001).


















                                                                                96








                         Table 6-10 Total PCB Concentrations in     Fish

                 species      Capillary Column     Packed Column    % Fat


                 Small Mouth Bass      3.4           3.7            0.6
                     it                4.3           4.2            0.7
                     if                3.9           3.5            0.4
                     If                4.3           4.2            0.8
                     It                5.8           4.4            1.2
                                       6.8           9.0            3.0
                                       2.3           2.6            0.8
                                       2.0           2.3            0.5
                 Bluegill              2.7           2.2            1.4
                 Walleye             10.1            6.7            1.8
                 Northern Pike         3.6           6.6            0.5
                     If                4.6           3.9            1.0
                     if                6.9           6.2            1.0
                     it                6.4           3.8            1.6
                 Rock Bass             5.1           6.0            2.1
                     It                1.3           1.4            0.4
                 Channel Catfish       9.5          13.0            5.1
                     11              22.0           32.0            3.4
                     of              13.9           26.0            6.5
                 Carp                22.0           21.0            4.1
                                     18.9           15.0            13.0











                                                                                              Figure 6-12 Homolog Distribution
                                                                                                                             in Fish


                                                                  Wt. percent
                                                             60


                                                             so                                                         . .... .. ..... .


                                                             40                                                                                  ...........  ........................ .............



                                                                                                                          ........... .... .                                  .............
                                                             30--




                                                                                                                                                           ..................      .................... - -



                                                             10        ..........
                                                                                                                                                              ......................- ..................




                                                                0
                                                                     Mono              Di            TO          Tetra Penta                  Hexa Hepta                    Octa          Nona
                                                                                                                      Homolog Group


                                                                                                 Sample                       Std 1246                       Std 1254


                                                       (Ave. of 21 samples)










                                        Figure 6-13 Homolog Distribution
                                              in Fish and Sediments


                             Wt. percent
                           40




                           30 -                         ..........  ......................





                                      ..............................
                           20 . . .. .. ... ...... ..





                                                                       ...................
                           10 . .. .... . .. ..... ......... . . ... ....




                             0
                              Mono    Di   TO    Te tra Penta Hexa Hepta Octa  Nona
                                                   Homolog Group


                                        Sample      Harbor     River Sediment


                        (Ave. of 21 samples)


















                                                                                99

                 6.5.3. Most Prominent Congeners in Fish

                      The most prominent congeners are listed in table 6-11

                 and also presented in figure 6-14.



                          Table 6-11 Most Prominent   Congeners in Fish

                 Congene         Sample      Aroclor  124 a  Aroclor 1254a

                 28/31              5.7            17.7            <0.1
                 47/48              6.8            4.0             0.4
                 49                 4.0            3.0             1.0
                 41/64/71           4.0            0.7             <0.1
                 66/95              8.5            9.0             7.6
                 77/110             7.7            2.5             9.5
                 101                4.2            1.7             7.1
                 118                4.5            0.8             3.0
                 132/153/105        4.9            1.5             11.0
                 138/163            4.8            1.8             5.6

                 aManchester, 1988









                                                  Figure 6-14 Most Prominent Congener
                                                                     in Fish


                                       Wt. Percent
                                    20




                                    15                                     . ... . ....         . . . . . ........ ......




                                    10 -                               ................. ........... ........... ......





                                                                ..........    . . ..... ...........






                                     0
                                        28/31 47/48    49    41/71 66/95  77/110   101    118 132/163138/163
                                                                    Congener

                                                    Fish        Aroclor 1248    ED Aroclor 1254


















                                                                              101

                                Chapter 7 Discussion of Results

                     In this section results f rom the sediment, water and f ish

                analyses are compared to results from the literature.

                Explanations for the congener patterns are evaluated by

                comparing predicted patterns to observed patterns.



                7.1. Discussion of Sediment Results

                7.1.1. Grain Size Correlations with PCB Concentration

                    Since sediment samples composed of smaller particles have

                greater surf ace area f or adsorption relative to samples of the

                same volume consisting of larger particles, it is predicted

                that smaller particles will have higher concentrations of PCBs

                relative to larger particles (Simmons et al., 1980).         This

                tendency is enhanced by the fact that smaller particles are

                often comprised of organic carbon which has been positively

                correlated with PCB content (Elzerman and Coates, 1987).       In

                this study, to examine the effect of grain size on PCB

                concentration, the two variables were correlated.

                     The correlation of grain size and concentration of PCBs

                is confounded by spatial effects such as distance from the

                source and the depth of the core.      To control for spatial

                variation, samples from only one site (Rochester Park, samples

                B and T) were correlated with the grain size. other samples

                were not correlated because there were too few samples at each

















                                                                            102

                site to give a meaningful correlation.      PCB concentration

                depends slightly on grain size; as the grain size decreased,
                the concentration of PCBs increased (R2 value of 0.52 for a

                log log plot). The outlying points were samples from either

                the surface of the core or from the bottom of the core.     One

                possible explanation for these outlying points is that

                sediment which is in the bottom of the core is older sediment

                and may not have been exposed to significant concentrations

                of PCBs. Conversely, sediment at the top of the core may be

                newer and cleaner sediment.     Both of these samples would

                therefore be likely to have low concentrations of PCBs because

                they were not exposed to the same amount of PCBs as the middle

                section of core.




                7.1.2. Total PCB concentrations in Sediment

                   Total sediment concentrations of PCBs ranged from 0.016

                ppm to 1586 ppm as seen in table 6-2.         Although higher

                concentrations  were   found by the Superfund          remedial

                investigation on the Sheboygan River, the range of

                concentrations are comparable (table 7-1).       The sediment

                results show a decreasing trend with distance from the source,

                similar to the results found in this study.

                    Although reported concentrations from other highly

                contaminated sites is comparable to Sheboygan River

















                                                                               103




                   Table 7-1 PCB Concentrations in Sheboygan River Sediment'
                This Studyb                            Remedial Investigationc
                Core   Concentration (ppm)            care Concentration (ppm)

                B1         173.1                       R4            4300
                                                       R5            59


                B2/B3      139.8                       R6            15
                Tl         24.1                        R7            4500
                T2         49.3                        R8            0.4
                T3         278.9                       R9            28.0
                T4         16


                Cl         40.2                        R19            250
                C2         534                         R20            2.8
                                                       R21            5.4
                                                       R22            93


                D1         88.9                        R34            110
                                                       R103           72
                                                       R104           5.4


                D2         95.8                        R37            <1.5
                                                       R35            2.0


                El         0.8                         R85            0.3
                E2         2.1                         R86            4.2
                E3         7.7                         R87            4.6

                  The two sets of cores are grouped according        to general
                 roximity
                  Samples are averages of cores which were 30 to 45 cm deep
                c Remedial investigation (EPA Superfund study) samples are
                composite core samples which were 30 to 45 cm deep (Blasland
                and Bouck Engineers, 1988)

















                                                                               104

                sediments, the PCB, concentrations in Sheboygan River sediments

                are considerably higher than other sites in the Great Lakes

                region. Brown and coworkers found concentrations as high as

                2600 ppm in Hudson River sediments with an average

                concentration in the hot spot areas of 75 ppm (Brown et al.,

                1984).    In the tidal Hudson, sediment PCB concentrations

                dropped to an average of 10 ppm (Bopp et al., 1981).           PCB

                concentrations in the Great Lakes are much lower. In Lake

                Ontario, sediment values were found in the range of 200-1200

                ppb (Oliver et al., 1989).

                    Within the cores, as mentioned in Chapter 6, the top

                section of the core generally contained low concentrations of

                PCBs.   The middle section of core usually contained the

                highest concentrations of PCBs and the bottom section of core

                typically had the lowest concentrations of PCBs. One possible

                explanation for this distribution is that the bottom section

                of sediment was deposited before significant contamination

                occured.whereas the top section of core was deposited after

                significant contamination took place and contains the newer

                cleaner sediment.

                    There was one exception to this general trend. In sample

                T3A the highest concentration was in the top segment of core.

                T cores were collected four months after all other cores.

                During these four months, federal and state remedial

















                                                                                 105

                 investigations of the site continued and it is possible that

                 the sediments were disturbed by other investigators.



                 7.2 Discussion of Possible Mechanisms Causing the Congener

                 Distribution in Sediment

                      The results from the sediment analysis clearly indicate

                 that the sediment PCB patterns are significantly different

                 from the Aroclor originally introduced into the river.          The

                 mechanisms which can alter the congener distribution in the

                 sediment can be divided into three processes-- chemical,

                 physical-chemical and biological.

                     Abiotic chemical reactions in the sediment affecting the

                 PCB congener distribution are unlikely.           Photolysis and

                 oxidation reactions are unlikely to occur in the subsurface

                 anaerobic. environment present in most of the sediments.

                 Although chlorinated methanes and ethanes have been found to

                 undergo reductive dechlorination (Wade et al., 1969; Klecka

                 and Gonsoir, 1984), reductive dechlorination of PCBs is

                 thought not to occur in the conditions present in anaerobic

                 sediments (Quensen et al., 1988; Brown et al., 1987b;

                 Hutzinger   et   al.,    1974).       For   example,     reductive

                 dechlorination of PCBs has been demonstrated to occur by Boyer

                 and colleagues (1985) at 50-90 degrees C using sodium

                 hypophosphite.     These temperatures are well above the

















                                                                              106

                temperatures f ound in the environment.        Furthermore,   the

                reduction of chlorinated ethanes and methanes by iron is      not

                strictly abiotic because iron is present as iron porphyrin

                which is a biological enzyme present in hemeglobin,

                cytochromes and chlorophyll (Fessenden and Fessenden, 1982).

                 In general, there are very few abiotic pathways which

                completely mineralize organic compounds (Alexander, 1981).



                7. 2. 1. Physical-Chemical Processes Af f ecting the Distribution

               .of PCBs in Sediment

                     Four   physical-chemical    processes    can   affect    the

                distribution of PCB congeners in sediment: sediment-water

                partitioning, diffusion, air-water partitioning, and colloidal

                partitioning.       A discussion of each follows with an

                evaluation of the predicted congener distribution based on the

                mechanism compared to the observed distribution.

                    Sediment-Water    Partitioning.       Probably,   the    most

                significant partitioning process affecting the distribution

                of congeners in the sediment is the partitioning between

                sediment and water (Bopp et al., 1981, Burkhard, 1985a) . The

                partitioning of hydrophobic compounds between sediment and

                water can be described by the following relationship

                (Karickhoff, 1981):
                           Kp=Se Pe

















                                                                            107

               where Kp is the equilibrium partition coefficient'.

                     S' is the equilibrium sorbed pollutant concentration,
                     P' is the equilibrium aqueous phase concentration.

               The partition coefficient is corrected for the organic carbon

               present in the sediment. The corrected partition coefficient

               is expressed as the organic carbon based partition

               coefficient, K,,, and is related to the partition coefficient

               by the following equation:

                       KOC =KP*oc
               where k.,--is-the---or anic carbon based partition coefficient,
                      KP is the partition coefficient,
                     oc is the fraction of organic carbon.

               The octanol-water partition coefficient (Kow) can be related

               to the organic carbon based partition coefficient ('Koc) by the

               following relationship (Karickhoff et al., 1979):
                   Log Koc=1.0 Log Kow -0-21               (R'= 1. 0 0)
               Using this relationship, the Kc can be estimated from the Kow.

               Since the octanol-water partition coefficient t       ends to

               increase with increasing chlorination (Shiu and Mackay, 1986),

               the partition coefficient for PCBs should also increase with

               increasing chlorination.

                   The above     relationships predict     that   the higher

               chlorinated congeners should remain in the sediment while the

               lower chlorinated congeners should partition into the water.


















                                                                           108

                Field studies at other sites show an -enrichment of the higher

                chlorinated congeners in sediment which has been attributed

                to partitioning (Furukawa, 1982; Hansen, 1987). For example,

                sediment from the Puget Sound in Washington shows an

                enrichment of the higher chlorinated congeners and loss of

                lower chlorinated congeners to the water (Pavlou and Dexter,

                1979).

                     Using the relationships of Karickhoff (1979) and

                predictions of Henry's Law constants and octanol-water

                partition coefficients, Burkhard (1984) and Burkhard et al.

                (1985) modelled the partitioning of Aroclor mixtures in a

                three phase system containing air, water and suspended

                particulate matter. Beginning with an equimolar distribution

                of congeners in all phases, Burkhard and colleagues examined

                the effects of partitioning into different phases on the

                congener distribution in each phase.

                      The air phase was found to be depleted in the higher

                chlorinated congeners above IUPAC 160.    In the water phase

                there was some enrichment of the lower chlorinated congeners

                below IUPAC 40 and some depletion of congeners above IUPAC

                120.  The most dramatic change was found in the distribution

                of the congeners in the suspended particulate matter phase.-

                Congeners below IUPAC 80 were significantly depleted while

                congeners above IUPAC 80 were significantly enriched.      The
















                                                                            109

               results of Burkhard (1984) and Burkhard et al. (1985)

               illustrate that the distribution of congeners in sediment is

               dramatically affected by the sediment-water partition

               coefficients.    Equilibrium modelling using sediment-water

               partition   coefficients    predicts   increases   in    higher

               chlorinated congeners and decreases in lower chlorinated

               congeners in the sediment.

                   Diffusion. Diffusion of congeners from the sediment back

               to the water is slow relative to sedimentation rates and is

               inversely related to the partition coefficient (Fisher et al.,

               1983; DiToro et al., 1985).     The diffusion rate of PCBs in
               the presence of sediment (D*s) is function of the inverse'of
               the partition coefficient and is described by the following

               relationship (DiToro et al., 1985):



                        D S= DS/ (1+m*KP/@)
               where D*, is the diffusion rate in the presence of sediment
                           (CM2/day),

                     D. is the aqueous diffusion rate (no sediment present)
                             (CM2/day) ,

                     m   is the sediment solids concentration,
                     KP  is the partition coefficient,
                         is the porosity of the sediment.















               microns such as humic and fulvic acids, microemulsions and

               mineral precipitates (McCarthy and Zachara, 1989).           The

               overall effect of colloids in the partitioning of organic

               contaminants is to increase the "apparent" solubility of a

               compound in water and decrease the "apparent" partition

               coefficient.

                   The effect of colloidal partitioning on the sediments may

               be limited since PCBs were probably introduced into the

               Sheboygan River adsorbed to sediment (Wisconsin Department of

               Natural Resources, 1989).     Hydraulic fluid containing PCBs

               was spilled on to dirt and debris which were later used to

               build a flood dike. Since the PCBs were already adsorbed to

               dirt that later became river sediment, colloidal partitioning

               could only occur after the PCBs were desorbed from the

               sediment into the water and eventually adsorbed onto the

               colloids.   The first step of this process is governed by

               sediment-water partition coefficients which, as explained

               before, gives a distribution that is enriched in the higher

               chlorinated congeners and depleted in the lower chlorinated

               congeners.   Therefore, it appears unlikely that colloidal

               partitioning has a major influence on the distribution of

               congeners in the Sheboygan River sediments.

                   None of the processes described above adequately explain

               the distribution seen in the Sheboygan River sediments. Since


















                                                                           110

               This relationship illustrates that higher chlorinated

               congeners are more likely to remain in the sediment than the

               lower chlorinated congeners because the higher chlorinated

               congeners have higher partition coefficients and subsequently

               lower diffusion rates.

                   Air-Water Partitioning.     The distribution of congeners

               between water and air phase can be important because it can

               affect the distribution between the water and sediment.     The

               effect of air-water partitioning of congeners on the overall

               distribution of PCB congeners in sediment is likely to be

               small in comparison to sediment-water partitioning because the

               sediments are not in direct contact with the air. The vapor

               partitioning of PCBs from sediment in the Hudson River was

               calculated to have a small effect relative to sediment-water

               partitioning (Bopp et al., 1981).   Even if one were to assume

               that the transport of PCBs from the sediment to the water and

               then to air were significant, the congener patterns seen in

               the Sheboygan sediments still would not be predicted. Henry's

               Law constants, which describe the partitioning of PCBs between

               the air and water, show no particular trend with regard to

               molecular weight (Burkhard et al., 1985b).

                   Colloidal Partitioning. Currently, there is considerable

               debate about the effect colloids have on partitioning.

               Colloids are defined as particles with diameters less than 10


















                                                                              112

                sediment-water partition coefficients increase with molecular

                weight, an enrichment of the higher chlorinated congeners and

                a depletion of the lighter chlorinated congeners is predicted

                based on the partition coefficients.                 Equilibrium

                partitioning models such as Burkhard's do not predict the

                specific enrichment of a select small group of congeners such

                as 5/8, 17, and 24/27 which were found in this study.

                Instead, these models predict a gradual continuum of

                increasing concentrations of the higher chlorinated congeners

                in the sediments.

                    Henry's Law constants show no particular trend with

                molecular weight.    The most prominent congeners seen in the

                sediment do not exhibit particularly low Henry Is Law constants

                and therefore their presence cannot be explained in terms of

                these constants.

                    In summary, neither physical-chemical processes nor

                abiotic chemical reactions readily explain the distribution

                of congeners in Sheboygan River- sediments.   Consequently, one

                or more biological processes may better explain the measured

                distribution.   Possible biological processes are discussed

                next.




                7.2.2.   Biological Processes affecting the Distribution of

                PCBs in the Sediment

















                                                                            113

                    Aerobic and anaerobic microbial degradation are probably

                the most likely biological processes that could affect the PCB

                distribution in sediment. Aerobic degradation, if it occurs

                at all, occurs in the top few centimeters of sediment where

                oxygen is present. Because most of the samples analyzed were

                slices of the core located 45 to 60 cm below the sediment

                surface, very little of the sediment was thought to be

                aerobic. Aerobic PCB-degrading microbes metabolize the lower

                chlorinated congeners, those with less than four chlorines,

                but generally do not metabolize the higher chlorinated

                congeners (Furukawa, 1982).

                    The pattern seen in the sediments     is more typical of

                anaerobic degradation in which the        higher chlorinated

                congeners are metabolized to produce       lower chlorinated

                congeners.   Several results in this study suggest anaerobic

                microbial degradation.    First, there is a shift in the

                congener pattern from the higher chlorinated congeners to the

                lower chlorinated congeners. This cannot be accounted for by

                simple physical -chemical partitioning. Second, there appears

                to be some selectivity as to which congeners were lost that

                is characteristic of enzymatic microbial degradation.

                Lastly, the congener patterns appear to be a function of the

                concentration of PCBs in the sediment that is typical of

                biological degradation.

















                                                                            114

                     A shift in congener distribution from the higher

               chlorinated congeners to the lower chlorinated congeners was

               seen by Quensen and coworkers (1988) in their laboratory study

               of dechlorination by anaerobic bacteria.     Over a period of

               four months,       Quensen and coworkers saw a stepwise

               dechlorination of the higher chlorinated congeners to produce

               the lower chlorinated congeners. Brown and coworkers (1987a

               and 1987b) postulated such a stepwise dechlorination process

               based on their results from Hudson River sediment cores.

                   With regard to congener selectivity, there is a loss of

               meta and para chlorinated congeners compared to ortho

               chlorinated congeners.      Figure 7-1 contains the average

               number of meta and para chlorines and ortho chlorines

               calculated for each sediment sample along with averages

               calculated for the Aroclor mixtures (General Electric Research

               and Development, 1987).    When Aroclor mixture averages are

               plotted, a linear relationship can, be seen, indicating that

               any combination of Aroclors will also fall on this line.

               Therefore, any deviation from this linearity may be attributed

               to a physical or biological process and not a simple mixing

               of different Aroclors (General Electric Research and

               Development, 1987).     Brown and coworkers maintain that an

               upward deviation indicates evaporation, elution and oxidative

               biodegradation whereas a downward deviation indicates









                                                                  Figure 7-1 Ortho vs Meta/Para Congener
                                                                                      in Sediment Samples

                                                   Meta/Para CI
                                                 5



                                                 4-                                                                                                       ..... ..... .-






                                                 2  . . . . ........                                                                                  ......................
                                                                                                                     4-   ;V, X
                                                                                                                     x
                                                                                                                 X +X      D<X x 0
                                                                                                              X           0 -t-
                                                                                    ......... .     . . . . ........  ... ..... ............ ................... ..... ... .................. .. ...... ....



                                                 Ol                                                                                        I
                                                   0                   0. 5                                        1.5                    2                    2.5
                                                                                                    Ortho CI

                                                   - Aroclors                             4-   Env. Samples (13)                   Env.   Samples      (C)
                                                     13   Env. Samples (D)                x    Env. Samples (T)                0 Env.     Samples      (E)
                                                                                                                xx
                                                                                                                    4t3







0

                                                                          115











































                Fig 7-1



  40

















                                                                                116

                anaerobic dechlorination (General Electric Research and

                Development, 1987).

                      Most of the sediment samples from the Sheboygan River

                fall below the Aroclor mixture line indicating a depletion in

                the meta/para chlorines relative to the ortho chlorines.        For

                comparison, the average meta and para and ortho chlorines for

                the spiked sediment samples ("clean" sediments spiked with

                Aroclors) were calculated.      The spiked sediments could be

                thought of as a control since they do not undergo microbial

                degradation. These are shown in figure 7-2. Note that they

                all lie along the line as predicted (General Electric Research

                and Development, 1987).      In a laboratory study of anaerobic

                dechlorination a marked loss in average number of meta/para

                chlorines per biphenyl molecule was also seen (Quensen et al.,

                1988).   Brown and coworkers found similar results in their

                field study of Hudson River sediments (1987a).

                       Exactly why bacteria selectively dechlorinate meta and

                para chlorines is unclear. Rusling and Miaw (1989) have shown

                that reductive dechlorination of meta and para chlorines gives

                a   greater    Gibbs   free   energy    value   than     reductive

                dechlorination of ortho chlorines. It is possible that

                bacteria which selectively dechlorinate meta and para

                chlorines have a biological advantage over those that do not.

                     With regards to specific congeners, several congeners are







 0

                                                                            117














































               Fig 7-2
  0








                                                                       Figure 7-2 Ortho vs Meta/Para Congeners
                                                                                                       in Spiked Sediments

                                                                  Meta/Para Cl




                                                               4  . .. ...... ... . . .     . ..... .                                                                                      ..........



                                                               3-                                     ............                                                           .... .......



                                                               2 -                              . .. .. .......                                                               ...............







                                                               0
                                                                 0                        0.5                         1                       1.5                         2                       2.5
                                                                                                                          Ortho CI


                                                                                                        Spiked Samples                              Aroclors

















                                                                           118

               found in abundance which would not be expected based simply

               on physical partitioning or on the original weight percents

               present in the Aroclor mixtures.      Table 7-2 contains the

               average weight percent of each congener in the samples

               containing over 50 ppm, PCBs as well as the weight percents of

               congeners in Aroclor 1248, Aroclor 1254 and a 50/50

               combination of Aroclors 1248 and 1254.

                   In the sediment samples, there is a general trend of

               enrichment of the lighter chlorinated congeners and depletion

               of the heavier chlorinated congeners. In particular, several

               congeners were significantly enriched in comparison to the

               50/50 combination of Aroclor 1248 and Aroclor 1254. Congeners

               with a greater than two percentage point increase, relative

               to the 50/50 mixture, were congeners 5/8, 19, 17, 24/27,

               16/32, 26 and 47/48.     Congeners with a greater than two

               percentage point decrease were 18, 28/31, 52, 44, 70/76,

               66/95, 56/60, 101, 77/110, 132/153/105, and 138/163.

                   It is interesting to note that these results are

               comparable to the results of Brown and coworkers (1987b).

               Table 7-3 shows the congeners Brown and coworkers found to be

               significantly depleted and enriched in the sediments. With

               the exception of congener 16 which coelutes with congener 32

               in this study, and congeners 47 and 49, all of the congeners

















                                                                                   119



                       Table 7-2 Changes in congeners in Sheboygan River
                                    Sediment Relative to Aroclors (Sediment
                                    Samples with conc. greater than 50 ppm)

                 Congener                  Concentration (Wt. Percent)
                               Sample    Aroclor'           Aroclor  a        50/50
                               Average    1248                 1254           Mixture

                 1              0.6       <0.1                <0.1
                 3              b.d.      <0.1                <0.1
                 4/10           2.1        0.8                <0.1
                 7              0.1        0.1                <0.1
                 6              0.8        0.1                <0.1
                 5/8           29.1        2.1                 0.1                 ++
                 19             2.5        0.5                 0.1                 ++
                 18             0.1        5.0                 0.1
                 17             9.4        1.2                <0.1                 ++
                 24/27          3.0        0.2                <0.1                 ++
                 16/32         10.0        2.0                <0.1                 ++
                 26             2.3        0.4                <0.1                 ++
                 28/31          6.4       17.7                 0.6
                 33             0.1        2.9                <0.1
                 22             0.5        0.6                <0.1
                 45             0.4        0.9                <0.1
                 46             0.1        0.5                <0.1
                 52             2.1        4.4                 4.2
                 49             2.4        3.0                 1.0
                 47/48          7.5        4.0                 0.4                 ++
                 44             1.1        5.2                 3.0
                 37/42          1.4        5.2                 0.1
                 41/64/71       2.0        2.5                 0.4
                 40             0.1        1.1                .0.1
                 74             0.4        2.6                 0.9
                 70/76          0.3        7.1                 4.4
                 66/95          3.8        9.0                 7.7
                 91             0.9       <0.1                 0.9
                 56/60          0.13       5.9                 0.9
                 84/92          1.4        1.4                 4.0
                 101            0.8        1.7                 7.1
                 99             0.5        0.9                 2.5
                 97             0.2        0.9                 2.2
                 87             0.1        0.9                 3.1
                 85             0.1        0.9                 1.5
                 136            0.1        0.1                 1.0
                 77/110         2.4        2.5                 9.2
                 82             b.d.      <0.1                 0.9
                 151            0.2        0.1                 0.8


















                                                                               120

                                   Table 7-2 Continued

                Congener     Sample    Aroclor 1248      Aroclor 1254
                            Average

                135/144       .0.2        0.1               1.4
                149           0.8         0.4               6.0
                118           0.4         0.8               3.0
                146           0.3       <0.1                2.2
                132/153/105   1.1         1.5              11.0
                141           b.d.      <0.1                0.5
                137/176       b.d.      <0.1                0.5
                138/163       0.6         0.3               5.6
                182/187       0.2         0.1               0.2
                183           0.1       <0.1                0.3
                185           b.d.      <0.1                0.1
                174           b.d.        0.1               0.4
                177           0.1       <0.1                0.1
                171/202       b.d.      <0.1                0.1
                172/197       b.d.      <0.1                0.1
                180           0.1         0.8               1.0-
                170/190       0.2       <0.1                1.0
                201           0.1         0.1              <0.1
                196/203       b.d.        0.1              <0.1
                195/208       b.d.      <0.1               <0.1
                194           b.d.      <0.1               <0.1
                206           b.d.      <0.1               <0.1

                a -- data from Manchester,   1988
                b.d..-- indicates below detection

                -- indicates a decrease by at least two percentage points from
                50/50 ratio

                ++ indicates an increase by at least two percentage points
                from 50/50 ratio


















                                                                               121




                   Table 7-3 Comparison of Congeners Predicted by Brown et
                al.(1987b) based on Anaerobic Degradation to Congeners
                observed in this Study

                Diminished      This                 Enriched        This
                Congeners       Study a             Congeners        Studya


                33                                       2              +
                22                                       6              +
                74                                       8             ++
                70                                       4              +
                66                                      10              +
                56                                      17              ++
                60                                      27              ++
                118                                     32              ++
                105                                     19              ++
                18                                      53              n.m.
                16              ++
                44
                42             n.m.
                87
                85
                82
                101
                99
                97
                153
                138
                141
                52
                49               +
                47              ++

                a  indicates samples with concentration above 50 ppm
                - indicates present in diminished quantities relative to
                50/50 ratio
                -- indicates a decrease by at least two percentage points from
                   50/50 ratio
                +  indicates present in increased quantities relative to 50/50
                   ratio
                ++ indicates an increase by at least two percentage points
                   from 50/.50 ratio
                n.m. not measured in this study


















                                                                            122

               which were found to Pe depleted in Brown's study were also

               found to be depleted in this study.

                   The concentration of PCBs in the sediment appeared to be

               a significant variable. As mentioned before in areas of low

               PCB concentrations, the sediment chromatograms appeared to be

               similar to the original Aroclors. Chromatograms from areas

               of high concentration had much higher weight percents of the

               lower chlorinated congeners relative to the original Aroclors.

                other researchers have noted that anaerobic dechlorination

               appears to occur only when there are high concentrations of

               PCBs present (Brown et al., 1984, Brown et al., 1987b, Quensen

               et al., 1988, Rhee et al., 1989).

                   Microbial degradation is often restricted to areas of high

               substrate concentrations.    For example, toluene, xylene and

               naphthalene are metabolized by bacteria at high concentrations

               but not at low concentrations (Alexander, 1985). The minimum

               concentration of a chemical which is needed to support growth

               of a microbial population is referred to as the threshold

               concentration (Alexander, 1985). Above the threshold

               concentration organisms are able to obtain sufficient energy

               for both maintenance and growth. As the concentration of a

               chemical compound decreases,     the amount of the compound

               available to the organism by diffusion decreases until

               insufficient quantities are available for both growth and


















                                                                           123

               maintenance (Alexander, 1985).         Below this threshold

               concentration, additional energy sources must be f ound to

               support growth of the population, since the organism is no

               longer able to completely mineralize the substrate.

                   Cometabolism occurs below the threshold concentration to

               provide additional sources of nutrients for the organism.

               Anaerobic dechlorination of higher chlorinated congeners in

               some cases has been shown to occur by cometabolism with the

               addition of biphenyl (Rhee et al., 1989).        Based on the

               different chromatographic patterns seen for high and low

               concentrations of PCBs, it is quite possible that some

               threshold    concentration    exists    for   the     anaerobic

               dechlorination of PCBs.




               7.3. Discussion of Water Results

               7.3.1. Total PCB Concentrations in Water

                    The total PCB concentrations in water are similar to

               other values which have been reported for the Sheboygan River

               (see table 7-4).    In a survey of Wisconsin Rivers, Marti

               (1984) found the Sheboygan River contained 0.103 (s.d. 0.036)

               ppb total PCB in the river. The values reported in this study

               are considerably higher than the concentrations found by Crane

               (Crane, 1990) in the Fox River which ranged from 1.43 to 34.4

               ppt for the dissolved fraction and 2.11 to 103 ppt for the


















                                                                               124

                particulate fraction or by Swackhammer (1985) in Lake Michigan

                which ranged f rom 0. 3 to 3 ppt total PCBs.         Mudroch and

                coworkers (1989) found the Wheately Harbor in Lake Erie to

                have concentrations ranging from 0.086 ppb to 0.291 ppb total

                PCBs.   In Hamilton Harbor in Lake Ontario, concentrations

                ranged from 0.086 ppb to 0.213 ppb total PCBs (Mudroch et al.,

                1989).     However, the concentrations in sediment from the

                Great Lakes are lower than those from the Sheboygan River.



                             Table 7-4 Total PCB Concentrations in Water

                site                         Concentration (Dpb)

                                     WDNR   Remedial Investigation    This Study
                                     1978             1987
                Above Sheboygan
                Falls Dam            <0.3       <0.05  P               0.007 P
                                               <0.05   D              0.003 D


                Rochester Park       <0.4                              0.213 P
                                                                      0.038 D


                Above Upper
                Kohler Dam           <0.2      0.267 P                  0.448 P
                                              0.118 D                 0.197 D


                Above Lower
                Kohler Dam                     0.150 P                  0.480 P
                                              0.078 D                 0.166 D


                Sheboygan            0.3      0.159 P                  0.301 P
                                              0.059 D                 0.070 D


                P - Particulate Fraction
                D - Dissolved Fraction

                Source: Wisconsin Department,of Natural Resources, 1989

















                                                                             125



                7.3.2. Homolog and Congener Distribution in Water

                    The weight percents of each of the homolog groups did not

                vary greatly among the samples.   Higher concentrations of the

                lower chlorinated congeners were found in the dissolved phase

                than in the particulate phase. This would be expected since

                the lower chlorinated congeners are in general more soluble

                and less likely to partition onto the particulate fraction.

                    Congeners 1, 4/10, 5/8, 17, 16/32, 28/31, 41/64/71, 47/48,

                52 and 66/95 were the most prominent congeners in the

                dissolved fraction.      Congeners 5/8, 17, 28/31, 47/48,

                41/64/71, 52, 66/95, 70/76, 44 and 77/110 were the most

                prominent congeners in the'particulate fraction.     Since the

                solubility of PCB congeners generally decreases ' with

                increasing chlorination and since the most, prominent congeners

                listed in table 6-9 do not show particularly high solubilities

                (Shiu and MacKay, 1986), it is likely that their abundance in

                the water column is a function of their high concentrations

                in the sediment.

                   The distribution and relative percentages of the most

                prominent congeners in the water increasingly reflect the

                congener distribution of the sediments in the lower reaches.

                For example, at Rochester Park congener 5/8 constitutes 2.9

                percent of the particulate fraction of PCBs. In contrast, at


















                                                                           126

               the other sites, congener 5/8 is 9.2 percent of the

               particulate fraction.   The weight percents of congeners 47/48

               and congener 17 also increase at the other sites.       In the

               dissolved phase, this trend is more pronounced. At Rochester

               Park, congener 5/8 constitutes 14.6 percent of the dissolved

               PCB concentration.    The average for the other sites for

               congener 5/8 is 30 percent. The weight percents of congeners

               171, 16/32 and 47/48 in the dissolved phase also increase.

               Most likely, these increases in specific congeners, which are

               seen in abundance in the sediment, are due to the scouring of

               the river bottom.




               7.4. Discussion of Fish Results

               7.4.1 Total PCB concentrations in Fish


                    The total PCB concentrations found in the fish that were

               analyzed are similar to other reports in the literature.

               The Wisconsin Department of Natural Resources routinely has

               fish analyzed from the Sheboygan River.    Recent results are

               listed in table 7-5.         In the Kiwanis Park area, the

               Wisconsin Department of Natural Resources found the fish to

               contain an average total PCB concentration of 8.4 ppm which

               is comparable to 7.6 ppm found in this study (Wisconsin

               Department of Natural Resources, 1989). In the Harbor area,

               the average totals of 42.7 ppm are considerably higher than

















                                                                                127

                those found in this study.

                     Ten of the fish in this study were from the Kiwanis Park

                area and eleven were from the harbor. In a survey of fish in

                Wisconsin rivers (including the Sheboygan River) values ranged

                from 0.07 to 7.0 ppm (Maack and Sonzogni, 1988).         Carp and

                lake trout from the Wisconsin side of Lake Michigan were found

                to have PCB concentrations similar to those reported in carp

                and other fish in this study (16.3 ppm for carp and a range

                of 2.9- 33.8 ppm for trout (Kleinert, 1976 in Swain, 1983)).





                Table 7-5 Total PCB Concentration in Fish from the Sheboygan
                           River (ppm)

                Site           Year     of Fish    Ave Conc-(Ppm)     Species

                Above Sheb.
                    Falls      1987       6          <0.30          varied


                Below Sheb.
                    Falls      1987        7           24.9          Carp,
                                                                    Sm Bass


                Upper Kohler
                    Dam        1987       3          7.67           Sm Bass


                Lower Kohler
                    Dam        1985       9         9.65            Varied

                Kiwanis Park   1986       9         8.4             Varied

                Sheboygan      1983      17         42.7            varied


                (Source: Wisconsin   Department of  Natural Resources, 1989)


















                                                                           128

                In the more highly contaminated Hudson River, fish were found

               to have total PCB concentrations ranging f rom below detection

               to 7.07 ppm (Califano et al., 1982).

                    The highest total PCB concentrations were found in fish

               that had high concentrations of f at to body weight.       This

               finding suggests a positive correlation between percent fat

               and the level of PCBs in fish. To test this hypothesis, the

               percent fat was regressed against the total PCB concentration.

               The results are plotted in figure 7-3.         The regression

               equation for this line follows:
                   Log PCB = 1.061 log fat - 0.644       R2 value of 0.71.

               The correlation between percentage of fat in the fish and the

               PCB contamination is significant.

                   The fish samples were analyzed using both a capillary

               column and a packed column.    The two methods were compared

               to see how well they agreed.        Twenty one samples were

               analyzed with both types of columns.     The measurements are

               compared by regression:
               Log Cap Col = 1.05 log Packed Col + 0.0005    R2 value o.ss.

               These results are plotted in figure 7-4.       This result is

               important since it implies that packed column measurements

               made over the last twenty years (by comparing chromatographic

               patterns   of  a   sample   with   that   of   pure   Aroclars

               (Erickson,1986)) produce results similar to that










                                                                                                              Figure 7-3 Correlation between
                                                                                                             Fish Fat and PCB Concentration


                                                                               Log PCB Cone.
                                                                       1.4



                                                                       1.2-




                                                                                                                                                                                   ..........





                                                                                                                             ......... .-                             . .......                               . ........
                                                                       0.8-                             . ..... .



                                                                       0.6     . ... ............. .. ....... ..........................
                                                                                                                                                        ...........             . ......



                                                                       0.4                              . . . ... . .. ..... .... .. .. ... . ............. ..............-      ..........     ............................



                                                                       0.2     . .......



                                                                            0
                                                                           -0.6            -0.4            -0.2                0             0.2             0.4             0.6             0.8                1             1.2
                                                                                                                                       Log Percent Fat










                                                                                                                     Figure 7-4 Correlation between
                                                                                                             Packed and Capillary Column Results

                                                                                    Log Capillary
                                                                            1.4



                                                                            1.2     . .. ..... ...








                                                                                                                                                        ........ ...     .. ...............                        . ..................- ..........
                                                                            0.8 -



                                                                            0.6     . . ........... .. .. . . . . ..... ...........     . ..... ....................... . .....-...........         ................






                                                                            0.2     . . . ..... ...                .. . . .. .......


                                                                                01
                                                                                   0                0.2                0.4                 0.6                0.8                   1                 1.2                1.4                1.6
                                                                                                                                                     Log Packed

















                                                                           131



                obtained by summing individual congeners. Maack and Sonzogni

                (1988) also found good agreement for total PCBs measured by

                packed and capillary column chromatography.



                7.4.2. Homolog Patterns in Fish

                   The enrichment of the higher chlorinated congeners and

                depletions of the lower chlorinated congeners seen in the fish

                homolog patterns (figure 6-12) has been observed by others.

                In the heavily PCB contaminated Hudson River, Califano and

                coworkers (1982) found that fish congener patterns were more

                typical of Aroclor 1254 than of the material introduced into

                the river, Aroclor 1242.   Because fish generally are unable

                to metabolize the higher chlorinated congeners     (Niimi and

                Oliver, 1989; Dekock and Lord, 1988; Bruggeman et  al., 1981;

                Lech and Peterson, 1983), it is not surprising     to see an

                enrichment in the higher chlorinated congeners.      Work by

                Peterson and Lech (1983) suggests that fish are only able to

                metabolize mono, di and tri chlorinated biphenyls.     It is

                interesting to note that fish tend to metabolize PCBs much

                more slowly than mammals (Lech and Peterson, 1983).

                   The higher chlorinated congeners tend to bioaccumulate not

                only within the fish but also within the entire food chain.

                Since many of the fish in this study are carnivorous and are


















                                                                           132

               therefore higher on the food chain, it would be expected that

               higher total concentrations of PCBs should be f ound in the

               fish in comparison to other biological compartments (van der

               Oost et al., 1988; Oliver and Niimi, 1988). In particular,

               higher concentrations of the higher chlorinated congeners is

               expected (Oliver and Niimi, 1988).      The data collected in

               this study support these findings as the fish are

               significantly enriched in the higher chlorinated congeners

               relative to the water and overall are enriched in total PCB


               concentration relative to the water.




               7.4.3 Most Prominent Congeners in Fish

                    The most abundant congeners found by Maack and Sonzogni

               (1988), and Niimi and Oliver (1988) are listed in table 7-6.

                The standard deviations for the most abundant congeners found

               in this study were fairly small indicating that the

               percentages of each congener amongst the samples was

               relatively constant. This occured despite the variation in

               species, PCB content and weight of the fish.

                   As mentioned before several of the congeners seen in high

               concentrations in the fish in this study are prominent in

               commercial Aroclors.   For example, in Aroclor 1248 the top

               three congeners are 28/31, 70/76, and 66 in the following

               percentages, 17.7, 7.1 and 7.1 (Manchester, 1988).

















                                                                                       133





                       Table 7-6 A Comparison of Most Prominent Congeners
                                    (Weight Percent)
                           This Study     Niimj and Oliver         Maack and Sonzpgn    ib
                             Species        Br.     Lk.      Rainbow            Species
                             Variedc       Trout Trout Trout                    Varied
                  Congener                              Percent

                  28/31     5.7 (1.4)                                              7.9

                  47/48     6.8 (0.9)

                  49        4.0  (0.47)

                  41/64/71  4.0  (0.66)

                  66/95     8.5  (0.78)                                            7.4

                  70/76                                                            4.5

                  101       4.2  (0.3)        6.3       6.1    7.2                 6.0

                  118       4.5  (0.6)        5.5       6.3    5.6                 2.7

                  77/110    7.7  (0.9)        5.6      4.8     5.8                 6.8

                  132/153   4.9  (0.85)       7.3     10.8     9.1                 11.0d
                     /105                                                          2.6

                  138/163 e 4.8  (0.81)       5.4    5.9       5.7                 8.1

                  146                                                              3.4


                  149                         5.1    3.4       4.8                 4.3


                  180                         4.2    4.6       4.5                 3.7


                  'Oliver and Niimi, 1989
                  bMaack and Sonzogni, 1988
                  cParentheses indicate standard deviation
                  dCongener 105 was reported separately from congeners 132/153
                  e Congener 138/163 was reported together in this study;
                  however the other researchers reported congener 138 alone.

















                                                                          134


               In Aroclor 1254, the top three congeners 153/132/105, 110 and

               95 constitute the following percentages, 11.0, 9.2, and 7.6

               (Manchester, 1988).     Congeners 47/48, 49, 41/64/71 were

               prominent congeners in the sediment which may account f or

               their presence in the fish.

                    Oliver and Niimi (1988) found that twelve congeners, 153,

               101, 84, 138, 110, 180, 87/97, 149, 187/182 and 105,

               contributed to over half the total percentage of PCBs in fish.

                Maack and Sonzogni (1988) found the ten most prominent

               congeners contributed over half of the total percentage (66

               percent).   In this study, it was also f ound that the ten most

               abundant congeners contributed over half the total percentage

               of PCBs in fish (55 percent).

                    Several congeners have been identified as bei          ng

               significantly more toxic than other congeners (Safe et al.,

               1985). They are characterized as containing at most one ortho

               chlorine and two para and at least two meta chlorines.

               Congeners containing no ortho chlorines are considered the

               most toxic; their structure is similar to TCDD and elicits

               similar toxic responses (Safe et al., 1985). Of the nonortho

               chlorines, only congener 77/110 was detected using the methods

               described in this study (see table 6-11).  Although congeners

               77/110 were detected, it is unlikely that the contribution of
               congener 77' is significant (Hansen, 1987; Maack and Sonzogni,



















                                                                               135

                1988).   Of the mono ortho chlorinated congeners, 105 and 118

                were detected (see table 6-11).       The precise quantities of

                congener 105 are not known since this congener coelutes with

                153 and 132; however, Maack and Sonzogni (1988) found congener

                105 to contribute approximately twenty percent of the combined

                total of the three congeners in their study of Wisconsin fish.

                 Congener 118 was found in all fish (see table 6-11).

                     Maack and Sonzogni (1988) found only two of the

                monochlorinated biphenyls, congener 118 and 105, in their

                study. They were unable to determine the presence or absence

                of congener 77 si4ce it coeluted with congener 110 which was

                present in large concentrations in the sample.       Congener 77

                has been found in fish from the Waukegan Harbor in Lake

                Michigan in quantities slightly diminished from the original

                Aroclor introduced, Aroclor 1254 (Huckins et al., 1988).

                Congener 105 was also found in fish from the Waukegan Harbor

                in approximately the same quantities as that in Aroclor 1254.

                    The retention of the various PCB congeners is thought to

                be a function of selective elimination of congeners rather

                than selective uptake (Bruggeman et al., 1981).            Uptake

                efficiencies have been found to be independent of their

                partition coefficients (Oppenhuizen and Schrap, 1988; van der

                Oost et al., 1988). However, the elimination rates are found

                to be a function of hydrophobicity.             For the higher


















                                                                                136

                chlorinated congeners, elimination rates are very low (van der

                Oost et al., 1988; Bruggeman et al., 1981). Conversely, the

                lower chlorinated congeners have been found to have high rates

                of clearance.    In one study of dichlorobiphenyl, the compound

                was found to be completely excreted 14 days after the fish had

                been switched from a contaminated environment to a clean one

                (Bruggeman et al., 1984).        Table 7-7 gives a f ew of the

                clearance rates for PCBs as reported in the literature. Note

                that the clearance rates increase with      increasing molecular

                weight.



                      Table 7-7 Clearance Rates for Specific Congeners

                Congene                Halflife (days)          Source

                2,5                            10         Bruggeman et al., 1981
                2,31,41,5                      60               11
                2,21,5                         14.4             11

                3,3,4,4                        34.6      Huckins et al., 1988
                2,3,5                            2.5            11

                2,21,3,315,51                139* Oppenhuizen and Schrap, 1988
                2,21,3,31,4,41,6,61            139*             It


                     Based on elimination rates given in their paper





                    It is well known that PCBs can be bioaccumulated to rather

                high levels in aquatic organisms even though the ambient

                concentrations in the water may be considerably lower (Oliver

















                                                                            137

               and Niimi, 1988; Gooch and Hamdy, 1983).     The ability of an

               organism to accumulate a chemical from water is often

               characterized by the bioconcentration factor, i.e., the

               concentration of the chemical in the organism divided by the

               concentration in the surrounding water (MacKay, 1982).

                    Mackay    predicted    a    relationship    between     the

               bioconcentration factor and the octanol-water partition

               coefficient based on thermodynamics. Incorporating laboratory

               data from a Veith et al. (1979) into the thermodynamic

               prediction, Mackay developed the following equation:
                  Log BCF= Log KOW -1. 32 .     (R'=O. 9 5) .

               Figure 7-5 is a comparison between the BCF values calculated

               from this study and Mackay's predicted relationship.         The

               bioconcentration factors found in this study were obtained by

               dividing the average fish concentration of a particular

               congener by the concentration in water of that particular

               congener.     Then the octanol-water coefficient for the

               particular congener was found and a second BCF value

               calculated using the relationship of Mackay.

                     With the exception of the first two congeners and two

               other congeners, the BCF values calculated in this field study

               were lower than those predicted by Mackay (1982).      This is

               surprising because the BCF calculated from laboratory studies

               usually underestimates the amount actually accumulated by f ish













                                                    Figure 7-5 Predicted BCF from Mackay
                                                     versus Calculated BCF from Field data

                                                                I oc@
                                             6  Measured@BCF from.Sheboygan River


                                                  Line if there is a one to one
                                                . ................................................................................................................................................... ............  ..............................................
                                                correlation between predicted
                                                  and measured BCF values

                                             4  . .............................. ...... ............................................ .............................. ............. . ................................................................





                                                                                                           .... ..... ... .... ... .... ... .... . ... ....
                                             3  . .............................................................................. ..............................................B.. C..Fc.a..I.. c..u..I..a..t.. e..d..........
                                                                                                   from observed PCBs
                                             2  . .......................................................  ...................I................................ con ci i-n fish a-n-d  wate-1





                                                . .........................  .....................................................................................................................................................................................







                                             0.
                                                0           1            2            3            4              5          6             7
                                                                Predicted         BCF     from     Mackay         (1982)
                                                Predicted BCF is calulated from Log.BCF = Log                                   Kow -1.32
                                                                                   BGF (from data)

                                                BCF (from data) Is           (conc. POB In fish)/(conc. PCB In water,)







 0

                                                                                            138












  I*



                                           I .














                   fig 7-5

                                                                                       1


                                                                                          1
  0


















                                                                                 139

                in the environment, in part because it ignores diet as a

                contributipn of PCBs.      It has been shown that the diet is a

                major source of PCBs to fish particularly with regards to the

                higher chlorinated PCBs (Niimi and Oliver, 1988; Rubenstein

                et al., 1984; Bruggeman et al., 1984).           Although the BCF

                values in this study were slightly lower than the values

                suggested by Mackay, these values are within the range of

                values reported by Mackay (1982) in a survey of the literature

                (see table 7-8).

                     Above a log octanol-water partition coefficient of about

                six, the relationship between the octanol-water partition

                coefficient and bioconcentration factor is no longer a simple

                linear relationship.      This deviation can be seen in figure

                7-5 in which Mackay's predicted line continues upward whereas

                the BCF values based on field data start to decrease at a log

                BCF value of about five, corresponding to a log KOW of about

                six.      It    is   therefore    difficult    to   predict     the

                bioconcentration factor for chemicals having log octanol-water

                partition coefficients above six.        Other researchers also

                have reported that the relationship between bioconcentration

                factor and the octanol-water partition coefficient is no

                longer linear after there are six or more chlorines on the

                biphenyl (Bruggeman et al., 1984).


















                                                                           140




                      Table 7-8 BCF Values Reported in the Literature
                                  (from Mackay, 1982)
               Relationship                              Cite

               BCF = 0.907log Kow -0.361           Baughman et al., 1984
               BCF = 0.837log Kow -0.770                it
               BCF = 1.160log Kow -0.75            Metcalf et al. 1975
               BCF = 0.85log Kow -0.7              Veith et al. 1979
               BCF = 0.542log Kow +0.124           Neeley et al. 1974


               Log BCF =1.15 log Kow -0.4          This study




                    Several reasons have been suggested for this decline.

               The more highly chlorinated compounds may not be as readily

               available to the fish relative to the lower chlorinated

               congeners due to their low solubility in water. In addition,

               the higher chlorinated congeners are more likely to form

               colloids that can not permeate fish membranes and therefore

               will not be adsorbed (Bruggeman et al., 1984).     It is also

               thought that compounds with molecular weight (greater than

               600) or large volumes may not be adsorbed because they are too

               large to permeate membranes (Oliver and Niimi, 1986).


















                                                                            140

                                   Chapter 8 Conclusions

               8.1. Sediment

                   The overall finding from this study is that the PCB

               congener distributions from highly contaminated sediments

               (greater than 50 ppm) are considerably different from the PCB

               congener distributions of the Aroclors originally used at the

               industrial site.      The highly contaminated sediments are

               enriched in mono, di, and tri chlorinated congeners relative

               to the original Aroclors.            In sediments containing

               concentrations of PCBs less than 50 ppm, the congener

               distributions were similar to the original Aroclors.

                    The enrichment in the'highly contaminated sediments with

               lower chlorinated congeners is not easily explained by

               existing physical -chemical partitioning relationships or known

               abiotic chemical reactions.     This suggests that a biotic
               process Jrs@@may be responsible for the enrichment. Anaerobic

               dechlorination by bacteria has been shown to occur in other

               highly contaminated river sediments.     The similarity of the

               PCB congener distribution in highly contaminated areas of the

               Sheboygan River to the results from other anaerobic microbial

               degradation studies at highly contaminated sites suggests that

               the congener distribution is the result of microbial

               degradation. Microbial degradation is also suggested by the

               concentration dependence of the patterns seen in the

















                                                                             141

               sediments.    Other studies of anaerobic microbial degradation

               of organic compounds have found a similar concentration

               dependency.



                  Specific findings are listed below:

               1) The river sediment is largely composed of fine grain sand

               (40 to 90 percent of the sample). Grain size correlated with

               PCB content. Smaller particles were found to contain higher

               concentrations of PCBs relative to larger particles of the

               same volume.




               2) Total PCB concentrations varied spatially in the river

               sediment.   With depth, PCB concentrations in sediment cores

               were the lowest in surface sediments (top 15 cm of the

               sediment core) and the bottom sediments (greater than 45 cm

               from the sediment-water interface).     Highest concentrations

               were found in the center of the core (15-45 cm down from the

               sediment-water interface).



               3) The total concentration of PCBs diminished with distance

               from the source, presumably due to the transport of PCBs

               downstream.




               4)    In sediment samples with greater than 50 ppm PCBs


















                                                                             142

                concentrations of congeners 4/10, 5/8, 17, 24/27, 16/32,

                47/48, and 26 were found to be enriched in the sediment

                compared to the original Aroclors. Congeners 5/8 were in the

                greatest abundance (29.2 percent on average).     In six of the

                samples, congeners 5/8 were found to comprise a large

                percentage of the sample (43.7 percent on average).



                5) Congeners with a greater than two percent increase relative

                to a 50/50 mix of Aroclor 1248 and Aroclor 1254 were 5/8, 19,

                17, 24/27, 16/32, 26 and 47/48. Congeners 5/8, 17, 24/27 and

                16/32 were also prominent congeners in the Hudson River where

                anaerobic dechlorination has been reported to occur.



                6) Congeners with a greater than two.percent decrease relative

                to a 50/50 mixture of Aroclor 1248 and 1254 were 18, 28/31,

                52, 44, 70/76, 66/95, 56/60, 101, 77/110, 132/153/105, and

                138/163.  In the Hudson River, congeners 44, 70/76, 66/95,

                132/153/105, and 138 were also found in diminished quantities.



                7) Harbor sediment samples had congener patterns similar to

                the original Aroclors and to the river sediments containing

                less than 50 ppm.




                8.2. Water

















                                                                           143

                    The total PCB concentrations in water were highest

               several miles downstream from the source.    The water samples

               were less contaminated directly at the source and in the

               harbor. The variation of total PCBs in water is most likely

               due to the transport of PCBs downstream. The fact that the

               highest concentrations were found several miles downstream

               from the source is not surprising. Water concentrations at

               the source would be lower because of constant inflow of "clean

               water" at Rochester Park and subsequent dilution of the

               contaminated river water.

                   Other findings are summarized below:



               1) With distance   from the source, the water developed a

               congener profile that was similar to the sediments.

               Presumably, this   is due to the entrainment of sediment

               particles in the   water column as a result of turbulent

               scouring.



               2) Higher concentrations of the lighter chlorinated congeners

               were found in the dissolved phase compared to the particulate

               phase. These results would be expected based on the partition

               coefficients and solubility constants for these congeners.



               3) Congeners 4/10, 5/8, 17, 16/32, 41/64/71 were found to be
















                                                                           144

               in high concentrations in the water'relative to the original

               Aroclors used at the source. These congeners were found to

               be prominent congeners in the sediment.



               8.3. Fish

                  The congener distribution in fish bore little resemblance

               to the sediment or water distribution. From a modelling and

               regulatory perspective, this finding is significant because

               some researchers have advocated using s ediment as a predictor

               of congener distributions in fish. Clearly, for rivers such

               as the Sheboygan River or Hudson River, use of these models

               is inappropriate since the fish are enriched in higher

               chlorinated congeners while the sediment is not.

                  Specific findings are listed below:

               1) In contrast to the water distribution, the f ish do not

               contain the lighter chlorinated congeners. This is consistent

               with previous studies of fish that show the lighter

               chlorinated congeners are metabolized and the heavier

               chlorinated congeners are bioaccumulated.



               2) There was a positive correlation between the weight percent

               fat in fish and PCB content.



               3) Fish samples were analyzed for total PCBs using both


















                                                                           145

               capillary column and packed column techniques.      Total PCBs

               measured by both techniques were similar.



               4) Congeners 28/31, 47/48, 49, 41/64/71, 66/95, 101, 118,

               77/110, 132/153/105 and 138/163 were prominent in fish.

               Congeners 28/31, 70/76, 66, 153/132/105, 110, 95 are the top

               three congeners in Aroclor 1248 and Aroclor 1254 respectively.

               Congeners 47/48, 49, 41/64/71 were found to be in abundance

               in the sediment.     Congeners 77, 105, and 118, which are

               considered to be among the most toxic PCBs, coelute so their

               relative importance (in terms of concentration in fish) is not

               clear.




               8.4. Concluding Remarks

               Further studies of anaerobic dechlorination of PCBs is

               warranted to determine if environmental conditions can be

               enhanced to encourage reductive dechlorination of PCBs.

               Specifically, investigations are needed to identify the

               microorganisms responsible for dechlorination, to determine

               the necessity of additional substrates (cometabolism), and to

               determine the type of anaerobic environment necessary

               (nitrogen atmosphere versus carbon dioxide atmosphere).

                   In addition, further research is warranted to determine

               the feasibility of using anaerobic microbial degradation as
















                                                                           146

               a large-scale treatment option for PCBs.        Currently, the

               Environmental Protection Agency (EPA) is conducting a pilot

               project on anaerobic microbial degradation of PCBs using

               highly contaminated Sheboygan River sediments.      At present

               however, much remains to be learned before microbial anaerobic

               degradation can be thought of as a efficient and cost-

               effective, large-scale treatment option.










          Bibliography
          Ahmed, M., and D.D. Focht. 1973. Degradation of Polychlorinated
               Biphenyls by two species of Achromobacter. Can. J.
               Microbiol., Vol. 19.

          Alexander, M. 1981. Biodegradation of Chemicals of
               Environmental Concern. Science, Vol. 211. pp. 132-138.

          Alexander, M. 1985. Biodegradation of Organic Chemicals.
               Environ. Sci. Technol., Vol. 18. No. 2. pp. 106-111.

          American Society of Testing and Materials (ASTM). 1987. Vol. 4.08
               Soil and Rock; Building Stones ASTM, Philadelphia, PA.

          Baker, J.E., Capel, P.D. and S.J. Eisenreich. 1986. Influence of
               Colloids on Sediment-Water Partition Coefficients of
               Polychlorobiphenyls Congeners in Natural Waters. Environ.
               Sci. Technol. Vol. 20. No. 11. pp.1136-1143.

          Bedard, D.L., Haberl, M.L., May, R.J., and M.J. Brennan. 1987a.
               Evidence for Novel Mechanism of PCB Metabolism in
               Alcaligenes eutrophus H850. Appl. Environ. Microbiol.,
               Vol.53. No. 5. pp. 1103-1112.

          Bedard, D.L., Wagner, R.E., Brennan, M.J., Haberl, M.L. and J.F.
               Brown. 1987b. Extensive Degradation of Aroclors and
               Environmentally transformed Polychlorinated Biphenyls by
               Alcaligenes eutrophus H850. Appl. Environ. Microbiol. Vol.
               53., No. 5. pp. 1094-1102.

          Blasland and Bouck Engineers. 1988. Preliminary Report on Results
               of Sheboygan River and Harbor Remedial Investigations.
               Syracuse. New York.

          Bopp, R.F., Simpson, H.J., Olsen, C.R. and N. Kostyk. 1981.
               Polychlorinated Biphenyls in Sediments of the Tidal Hudson
               River, New York. Environ. Sci. Technol., Vol. 15. No. 2.
               pp. 210-216.

          Boyer, S.K., McKenna, J., Karliner, J. and M. Nirsberger. 1985. A
               Mild and Efficient Process for Detoxifying Polychlorinated
               Biphenyls. Tetrahedron Letters. Vol. 26. No. 31. pp. 3677-
               3680.


          Brown, J.F., Wagner, R.E., Bedard, D.L., Brennan, M.J., Carnahan,
               J.C., May, R.J. and T.J. Tofflemire. 1984. PCB
               Transformations in Upper Hudson Sediments. Northeastern
               Environmental Science, Vol. 3. No. 3/4. pp. 166-178.

          Brown, J.F., Bedard, D.L., Brennan, M.J., Carnahan, J.C., Feng,
               H. and R.E. Wagner. 1987a. Polychlorinated Biphenyl
               Dechlorination in Aquatic Sediments. Science, Vol. 236.
               pp. 709-712.










          Brown, J.F., Wagner R.E., Feng, H., Bedard, D.L., Brennan, M.J.,
               Carnahan, J.C. and R.J. May. 1987b. Environmental
                Dechlorination of Polychlorinated Biphenyls. Environmental
               Toxicology and Chemistry, Vol. 6. pp. 579-593.

          Bruggeman, W.A., Marton, L.B.J.M., Kooiman, D. and 0. Hutzinger.
               1981. Accumulation and Elimination Kinetics of Di, Tri, and
               Tetra chlorobiphenyls by Goldfish after Dietary and Aqueous
               Exposure. Chemosphere. Vol. 10. No. 8. pp. 911-832.

          Bruggeman, W.A., Oppenhuizen, A., Wijbenga, A. and 0. Hutzinger.
               1984. Bioaccumulation of Super Lipophilic Chemicals in
               Fish. Toxicology and Environmental Chemistry. Vol. 7. pp.
               173-189.


          Brunner, W., Sutherland, F.H., and D.D. Focht. 1985. Enhanced
               Biodegradation of Polychlorinated Biphenyls in Soil by
               Analog Enrichment and Bacterial Inoculation. J. Environ.
               Qual. Vol. 14. No. 3. pp. 324-328.

          Burkhard, L.P. 1984. Physical-Chemical Properties of
               Polychlorinated Biphenyls: Measurement, Estimation, and
               Application to Environmental Systems. PhD Thesis. Water
               Chemistry Program, University of Wisconsin, Madison,
               Wisconsin.

          Burkhard, L.P., Armstrong, D.E., and A.W. Andren. 1985a.
               Partitioning Behavior of Polychlorinated Biphenyls.
               Chemosphere. Vol. 14. No. 11/12. pp.1703-1716.

          Burkhard, L.P., Armstrong, D.E. and A.W. Andren. 1985b. Henry's
               Law Constants for Polychlorinated Biphenyls. Environ. Sci.
               Technol. Vol. 19. No. 7. pp.590-596.

          Califano, R.J., O'Connor, J.M. and J.A. Hernadez. 1982.
               Polychlorinated Biphenyl Dynamics in Hudson River. Striped
               Bass. I Accumulation in Early Life History Stages. Aquatic
               Toxicology. Vol. 2. pp.187-204.

          Chantry, W. 1989. Biodegradation of PCBs Sorbed to Sewage Sludge
               Lagoon Sediments in An Aerobic Digester. PhD Thesis.
               Department of Civil and Environmental Engineering,
               University of Wisconsin, Madison, Wisconsin.

          Chen, M., Hong, C.S., Bush, B. and G.Y. Rhee. 1988. Anaerobic
               Biodegradation of PCBs by Bacteria from Hudson River
               Sediments. Ecotoxicology and Environmental Safety, Vol. 16.
               pp. 95-105.

          Clark, R.R., Chian, E.S.K. and R.A. Griffin. 1979. Degradation
               of Polychlorinated Biphenyl by Mixed Microbial Cultures.
               Appl. Environ. Microbiol. Vol. 37. No. 6. pp. 680-685.

          Connor, M.S. 1984. Fish/Sediment Concentration Ratios for Organic










               Compounds. Environ. Sci. Technol. Vol. 18. No. 1. pp. 31-
               35.

          Crane, J. 1986. Preliminary Report for PhD examination.
               University of Wisconsin, Madison, Wisconsin.

          Crane, J. 1990. The Influence of Water/Particle Partitioning on
               the Transport of Water Column PCB congeners in a
               Contaminanted Lake System. PhD Thesis, Water Chemistry
               Program, University of Wisconsin, Madison.

          Das, B.M. 1986. Soil Mechanics Laboratory Manual. Engineering
               Press, San Jose, CA.

          Dekock, A.C. and D.A. Lord. 1988. Kinetics of Uptake and
               Elimination of Polychlorinated Biphenyls by an Estuarine
               Fish Species (Rhabdosargus Holubi) after Aqueous Exposure.
               Chemosphere. Vol. 17. No. 12. pp.2381-2390.

          DiToro, D.M., Jeris, J.M. and D. Clarcia. 1985. Diffusion and
               Partitioning of Hexachlorobiphenyl in Sediments.  Environ.
               Sci. Technol. Vol. 19. No. 12. pp.1169-1172.

          Elzerman, A.W. and J.T. Coates. 1987. Hydrophobic Organic
               Compounds on Sediments: Equilibria and Kinetics of Sorption
               in Sources and Fates of Aauatic Pollutants R. Hites and S.J.
               Eisenreich (Eds.) American Chemical Society, Washington
               D.C.


          Erickson, M. 1986. Analytical Chemistry of PCEs. Butterworth
               Publishers. Boston.

          Fessenden, R.J. and J.S. Fessenden. 1982. Organic Chemistry,
               Willard Grant Press. Boston, MA.

          Fathepure, B.Z., Tiedje, J.M. and S.A. Boyd. 1987. Reductive
               Dechlorination of 4-Chlororesorcinol by Anaerobic
               Microorganisms. Environ. Tox. and Chem. Vol. 16. pp. 929-
               934.

          Fisher, J.B., Petty, R.L. and W. Lick. 1983. Release of
               Polychlorinated Biphenyls from Contaminated Lake Sediments:
               Flux and Apparent Diffusivities of Four Individual PCBs.
               Environ. Pollut. Series. B. Vol. 5. pp 12.1-132.

          Furukawa, K., Tonomura, K. and A. Kamibayashi. 1978. Effect of
               Chlorine Substitution on the Biodegradability of PCBs.
               Appl. and Environ. Microbiol. Vol. 35. No. 2. pp 223-227.

          Furukawa, K. 1982. Microbial Degradation of PCBs in
               Biodegradation and Detoxification of Environmental
               Pollutants A.M. Chakrabarty (Ed.). CRC Press, Boca Raton,
               FL.










          Furukawa, K., Tomizuka, N. and A. Kamibayashi. 1983. Metabolic
               Breakdown of Kaneclors (PCBs) and their Products by
               Acinetobacter sp. Appl.   Environ. Microbiol. Vol. 46.

          General Electric Research and  Development Center. 1987.
               Research and Development  Program for Destruction of PCBs.
               Seventh Progress Report. Schenectady, New York.

          General Electric Research and  Development Center. 1988.
               Research and Development  Program for Destruction of PCBs.
               Seventh Progress Report. Schenectady, New York.

          Gooch, J.A. and M.K. Hamdy. 1983. Uptake and Concentration Factor
               of Aroclor 1254 in Aquatic Organisms. Bull. Environ.
               Contam. Toxicol. Vol. 31. pp. 445-452.

          Hansen, L.G. 1987. Environmental Toxicology of PCBs in
               PCBs: Mammalian and Environmental Toxicolog S. Safe (Ed.).
               Springer Verlag, Berlin.

          Hawker, D.W. and D.W. Connell. 1988. Octanol-Water Partition
               Coefficients of Polychlorinated Biphenyl Congeners.
               Environ. Sci. Technol. Vol. 22. pp. 382-387.

          Head, K.H. 1980. Manual of Soil Laboratory Testing. Pentech
               Press, London.

          Hillel, D. 1982. Introduction to Soil Physics. Academic Press,
               New York.

          Horowitz, A., Suflita, J.M. and J.M. Tiedje. 1983. Reductive
               Dehalogenations of Halobenzoates by Anaerobic Lake Sediment
               Microorganisms. Appl. Environ. Microbiol. Vol. 45. No. 5.
               pp. 1459-1465.

          Horvath, R. 1972. Microbial Co-Metabolism and the Degradation
               of Organic Compounds in Nature. Bacteriological Reviews,
               Vol. 36. No. 2.

          Horzempa, L.M. and D.M. DiToro. 1983. The Extent of
               Reversibility of Polychlorinated Biphenyl Adsorption. Water
               Res. Vol. 17. No. S. pp.851-859.

          Huckins, J.N., Schwartz, T.R., Petty, J. and C.M. Smith. 1988.
               Determination, Fate and Potential Significance of PCBs in
               Fish and Sediment Samples with Emphasis on Selected AHH-
               inducing Congeners. Chemosphere. Vol. 17. No. 10. pp.1995-
               2016.

          Hutzinger, 0., Safe, S. and V. Zitko. 1974. The-Chemistry of
               PCBS. CRC Press, Cleveland, OH.

          Kaneko,.M., Morimoto, K. and S. Nambu. 1976. The Response of
               Activated Sludge to a Polychlorinated Biphenyl. Water Res.










               Vol. 10. pp. 157-163.

          Karickhoff, S.W. 1979. Sorption of Hydrophobic Pollutants on
               Natural Sediments. Water Res. Vol. 13. pp. 241-248.

          Karickhoff, S.W. 1981. Semi-empirical Estimation of Sorption of
               Hydrophobic Pollutants on Natural Sediments and Soils.
               Chemosphere. Vol. 10. No. 8. pp. 833-846.

          Keeton, W.T. 1980. Biological science. W.W. Norton and Co. New
               York.


          Klecka, G.M. and S.J. Gonsoir. 1984. Reductive Dechlorination of
               Chlorinated Methanes and Ethanes by Reduced Iron (II)
               Porphyrins. Chemosphere. Vol. 13. No. 3. pp. 391-402.

          Kohler, H.P.E., Kohler-Staub,   and D.D. Focht. 1988.
               Cometabolism of PCB: Enhanced Transformation of Aroclor 1254
               by Growing Bacterial Cells. Appl. Environ. Microbiol. Vol.
               54. No. 8. pp. 1940-1945.

          Krumme, M.L. and S.A. Boyd. 1988. Reductive Dechlorination of
               Chlorinated Phenols in Anaerobic Upflow Bioreactors. Water
               Res. Vol. 22. No. 2. pp. 171-177.

          Lech, J. and R. Peterson. 1983. Biotransformation and Persistence
               of Polychlorinated Biphenyls in Fish in PCBs: Human and
               Environmental Hazards, D11tri, F.M. and M. Karin (Eds.).
               Butterworth Publishers.. Boston.

          Lui, D. 1980. Enhancement of PCB Biodegradation by Sodium
               Ligninsulfonate. Water Res. Vol. 14. pp. 1467-1475.

          Maack, L. and W.C. Sonzogni. 1988. Analysis of Polychlorobiphenyl
               Congeners in Wisconsin Fish. Arch. Environ. Contam.
               Toxicol. Vol. 17. pp. 711-719.

          Mackay, D. 1982. Correlation of Bioconcentration Factors.
               Environ. Sci. Technol. Vol. 16. pp. 274-278.

          Mackay, D. and A.I. Hughes. 1984. Three Parameter Equation
               describing the Uptake of Organic Compounds by Fish.
               Environ. Sci. Technol. Vol. 18. No. 6. pp. 439-444

          Manchester, J. 1988. Seasonal Variation in the Amount and
               Distribution of Atmospheric PCB Congeners. Masters Thesis.
               Water Chemistry Program, Dept. of Civil and Environmental
               Engineering. University of Wisconsin, Madison, Wisconsin.

          Marti, E.A. 1984. PCBs in Sixteen Lake Michigan Tributaries.
               Master's Thesis, Water Chemistry, University of Wisconsin-
               Madison.

          McCarthy, J.F. and J.M. Zachara. 1989. Subsurface Transport of










               Contaminants. Environ. Sci. Technol. Vol. 23. No. 5. pp.


          Mikesell, M.D. and S.A. Boyd. 1988. Enhancement of
               Pentachlorophenol Degradation in Soil through Induced
               Anaerobiosis and Bioaugmentation with Anaerobic Sewage
               Sludge. Environ. Sci. Technol. Vol. 22. No. 12. pp. 1411-
               1414.


          Mudroch, A., Onuska, F.I., and L, Kalas. 1989. Distribution of
               Polychlorinated Biphenyls in Water, Sediment and Biota of
               Two Harbours. Chemosphere. Vol. 18. No. 11/12. pp. 2141-
               2154.

          National Academy of Science. 1979. Polychlorinated Biphenyls.
               National Academy of Science. Washington, D.C.

          Niimi, A.J. and B.G. Oliver. 1988. Influence of Molecular Weight
               Molecular Volume on Dietary Absorption. Can. J. Fish.
               Aquatic Sci. Vol. 145. pp. 222-227

          Niimi, A.J. and B.G. Oliver. 1989. Distribution of
               Polychlorinated Biphenyl Congeners and other Halocarbons in
               Whole Fish and Muscle among Lake Ontario Salmonids.
               Environ. Sci. Technol. Vol. 23. No. 1. pp.83-88.

          Oliver, B.G., Charlton, M.N. and R.W. Durham. 1989.
               Distribution, Redistribution and Geochronology of PCB
               congeners and other Chlorinated Hydrocarbons in Lake Ontario
               Sediments. Environ. Sci. Technol. Vol. 23. pp. 200-208.

          Oliver, B.G. and A.J. Niimi   1988. Trophodynamic Analysis-of
               Polychlorinated Biphenyl Congeners and other Chlorinated
               Hydrocarbons in Lake Ontario Ecosystem. Environ. Sci.
               Technol. Vol. 22. pp. 388-397.

          Opperhuizen, A., Velde, E.W., Cobas, F.A.P.C., Liem, D.A.K. and
               J.M.D. Steen. 1985. Relationship between Bioconcentration
               in Fish and Steric Factors of Hydrophobic Chemicals.
               Chemosphere. Vol. 14. No. 11/12. pp.1871-1896.

          Opperhuizen, A. and S.M. Schrap. 1988. Uptake Efficiencies of
               Two Polychloro Biphenyls in Fish after Dietary Exposure to
               Five Different Concentrations. Chemosphere. Vol. 17. No. 2.

          Palmer, D.T., Linkfield, T.G., Robinson, J.B., Genthner, B.R.S.
               and G.E. Pierce. 1989. Determination and Enhancement of
               Anaerobic Dehalogenation: Degradation of Chlorinated
               organics in Aqueous Systems. U.S. Environmental Protection
               Agency Research and Development, Risk Reduction Engineering
               Lab, Cincinnati, OH. EPA/600/S2-88/054.

          Parkinson, A. and S. Safe. 1987. Mammalian Biologic and Toxic
               Effects of PCBs in PolVchlorinated-Biiphenyls: Mammalian and










               Environmental Toxicology. Springer-Verlag. New York.


          Parsons, J., Veerkamp, W. and 0. Hutzinger. 1983. Microbial
               Metabolism of Chlorobiphenyls. Toxicological and
               Environmental Chemistry. Vol. 6. pp. 327-350.

          Parsons, J.R. and D.T.H.M. Sijm. 1988. Biodegradation Kinetics of
               Polychlorinated Biphenyls in Continuous Cultures of a
               Pseudomonas Strain. Chemosphere. Vol.17. No.9. pp.1755-
               1766.


          Paull, R.K. and R.K. Paull. 1976. Geology of Wisconsin and
               Upper Michigan. Kendall/Hunt Publishing. Dubuque, Iowa.

          Pavlou, S.P. and R.N. Dexter. 1979. Distribution of
               Polychlorinated Biphenyls (PCBs) in Estuarine Ecosystems.
               Testing the Concept of Equilibrium Partitioning in the
               Marine Environment. Environ. Sci. Technol. Vol. 13. No. 1.

          Quensen, J.F., Tiedje, J.M. and S.A. Boyd. 1988.      Reductive
               Dechlorination of Polychlorinated Biphenyls by Anaerobic
               Microorganisms from Sediments. Science. Vol. 242. pp. 752-
               754.


          Rhee, G.Y., Bush, B., Brown, M.P., Kane, M. and L. Shane. 1989.
               Anaerobic biodegradation of Polychlorinated Biphenyls in
               Hudson River Sediments and Dredged Sediments in Clay
               Encapsulation. Water Res. Vol. 23. No. 8. pp. 957-964.

          Rubenstein, N.I. Lores, E. and N.R. Gregory. 1983. Accumulation
               of PCBs, Mercury and Cadmium by Nereis Virens, Mercenaria
               Mercenaria and Palaemonetes Pugio from Contaminated Harbor
               Sediments. Aquatic Toxicology. Vol. 3. pp.249-260.

          Rubenstein, N.I., Gilliam, W.T. and N.R. Gregory. 1984. Dietary
               Accumulation of PCBs from Contaminated Sediment Source by a
               Demersal Fish (Leiostomus Xanthurus). Aquatic Toxicology.
               Vol. 5.. pp. 331-342.

          Rusling, J.F. and C.L. Miaw. 1989. Kinetic Estimation of
               Standard Reduction Potentials of Polyhalogenated Biphenyls.
               Environ. Sci. Technol. Vol. 23. No. 4. pp. 476-479.

          Safe, S. 1984. Microbial Degradation of PCBs in Microbial
               Decfradation of-Organic Compounds D.T. Gibson (Ed.). Marcel
               Dekker. New York.

          Safe, S., Bandiera, S., Sawyer, T., Zmudzka, B, Mason, G.,
               Romkes, M., Denomme, M.A., Sparling, S., Okey, A.B., and T.
               Fujita. 1985. Binding to the 2,3,7,8 -TCDD Receptor Protein
               and AHH Induction--Halogenated Biphenyls. Environ. Health.
               Persp. Vol. 61. pp.21-33.










          Safe, S. 1987. Determination of 2,3,7,8-TCdd Toxic Equivalent
               Factors (TEFs): Support for the Use of the in Vitro AHH
               induction Assay. Chemosphere. Vol. 16. No. 4. pp. 791-802.

          Shaw, G.R. and D.W. Connell. 1984. Physicochemical Properties
               Controlling Polychlorinated Biphenyl Concentrations in
               Aquatic organisms. Environ. Sci. Technol. Vol. 18. pp. 18-
               23.

          Shiu, W.Y. and D. Mackay. 1986. A Critical Review of Aqueous
               Solubilities, Vapor Pressures, Henry's Law Constants and
               Octanol-Water Partition Coefficients of the Polychlorinated
               Biphenyls. J. Phys. Chem. Ref. Data. Vol.15. No. 2.
               pp.911-929.

          Simmons, M.S., Bialosky, D.I. and R. Rossman. 1980. PCB
               Contamination in Surficial sediments of NE Lake Michigan.
               J. Great Lakes Res. Vol. 6. No. 2. pp. 167-171.

          State Laboratory of Hygiene. 1989.    Methods and Quality Control
               Manual. organic Chemistry Unit. Wisconsin State Laboratory
               of Hygiene. Madison, Wisconsin.

          Steen, W.C., Paris, D.F., and G.L. Baughman. 1978. Partitioning
               of Selected Polychlorinated Biphenyls to Natural Sediments.
               Wat. Res. Vol. 12. pp. 655-657.

          Stryer, L. 1981. Biochemistry. W.H. Freeman and Co. San
               Francisco.

          Suflita, J.M., Horowitz, A., Shelton, D.R. and J.M. Tiedje.
               1982. Dehalogenation: A Novel Pathway for the Anaerobic
               Biodegradation of Haloarozatic Compounds. Science. Vol.
               218. pp. 1115-1117.

          Swackhammer, D.L. 1985. The Role of Water-Particle Partitioning
               and Sedimentation in Controlling the Fate and Transport of
               PCBs in Lakes. PhD Thesis. Oceanography and Limnology.
               University of Wisconsin. Madison, Wisconsin.

          Swain, W. 1983. An Overview of the Scientific Basis for Concern
               with Polychlorinated Biphenyls in the Great Lakes in PCBs:
               Human and Environmental Hazards. F. DILtri and M.A. Xamrin
               (Ed.). Butterworth Publishers, Boston, MA.

          Tucker, E.S., Saeger, V.W. and 0. Hicks. 1975. Activated Sludge
               Primary Biodegradation of Polychlorinated Biphenyls. Bull.
               Environ. Contam. Toxicol. Vol. 14. No. 6.

          United States Environmental Protection Agency. 1989. Fact sheet
               Sheboygan River and Harbor Update. September 1985. Region 5.
               Office of Public Affairs, Chicago, IL.

          van der Oost, R. Heida, H. and A. Opperhuizen. 1988.










                Polychlorinated Biphenyl Congeners in Sediment, Plankton,
                Molluscs, Crustaceans and Eel in a Freshwater Lake.
                Implications of Using Reference Chemicals and indicator
                organisms in Bioaccumulation Studies. Arch. Environ.
                Contam. Toxicol. Vol. 17. pp. 721-729.

          Wade, R.S., Harlin, R. and C.E. Castro. 1969. Note: Organic
                Matter and Reduced Iron Porphyrins. Journal of the American
                Chemical Society. Vol.91. No. 26. pp. 7530.

          Wisconsin Department of Natural Resources. 1989. Sheboygan River
                Remedial Action Plan. Wisconsin Department of Natural
                Resources, Madison, Wisconsin.

          Wong, P.T.S. and K.L.E. Kaiser. 1975. Bacterial Degradation of
                PCBs 11 Rate Studies. Bull. Environ. Contam. Toxicol. Vol.
                13. No. 2.

          Woods, S.L., Ferguson, J.F. and M.M. Benjamin. 1989.
                Characterization of Chlorophenol and Chloromethoxybenzene
                Biodegradation during Anaerobic Treatment. Environ. Sci.
                Technol. Vol. 23. pp. 62-68.

          Wu, S. and D.M. Gshwend. 1986. Sorption Kinetics of Hydrophobic
                Organic Compounds to Natural Sediments and Soils.      Environ.
                Sci. Technol. Vol. 20. No. 7. pp.717-725.

          Zeikus, J.G. and M.R. Winfrey. 1976. Temperature Limitations of
                Methanogenesis in Aquatic Sediments. Appl. Environ.
                Microbiol. Vol. 31. pp99-107.





























            Concentrations of Toxic Polychlorinated Biphenyl Congeners

                        in Sheboygan River (USA) Sediments











                                       by   W. Sonzogni

                                            L. Maack


                                            T. Gibson


                                            J. Lawrence




                                            Laboratory of Hygiene

                                            and Water Chemistry Program

                                            UnIversity of Wisconsin

                                            Madison, WI 53706























               In the U. S. polychlorinated biphenyls (PCBs) were
               produced in mixtures (Aroclors) of the 209 different PCB
               congeners. The Aroclors dif f er not only in the congeners
               contained.in the mixture, but also in the weight percent
               of   the   congeners.      Until   recently   almost    all
               environmental and biological samples were analyzed for
               PCBs by matching the chromatographic pattern to the
               pattern of pure Aroclors (Webb and McCall 1973; Erickson
               1986), and results were reported as an Aroclor or mixture
               of Aroclors.      How'ever, with the refinement. of. high
              .resolution capillary gas chromatographic techniques, it
               is now possible to identify and quantify individual PCB
               congeners in environmental samples (Mullin et al. 1984;
               Maack and Sonzogni 1988).      Note that in this paper
               congeners will be referred to by their International
               Union of Pure Applied Chemistry (IUPAC) numbers and by an
               abbreviation of their structure. For example, 3, 3', 4,
               4' tetrachlorinated biphenyl is referred to as congener
               77 (34-34).

               Interest in individual congener concentrations has also
               increased because recent toxicological data indicate the
               potency of congeners varies widely (Safe 1987; Tanabe et
               al. 1987). Those congeners presently thought to be most
               toxic are the non-ortho chlorinated PCBs, since these co-
               planer compounds structurally resemble the highly potent
               2,3,7,8-tetrachlorodibenzo-dioxin (TCDD)     Congeners 77
               (34-34), 126(345-34) and 169 (345-345) are non-ortho
               chlorinated and most resemble dioxin. They are believed
               to be the most toxic congeners, at least in terms of
               dioxin-like properties (Safe et al. 1985; Tanabe et al.
               1987). Congener 81 (345-4) also has no ortho chlorines;
               however, it is structurally different enough from TCDD
               that it is not considered to be as toxic as the compounds
               above. The mono-ortho congeners, such as 105

               Send reprint requests to W. Sonzogni at the above
               address.









                 (234-34) , 118 (245-34) , 123 (345-24) , 114 (2345-4) and
                 167 (245-345) are also thought to be toxic (have toxic
                 properties related to dioxin) , although they are less
                 potent.(Safe et al. 1985; Safe 1987).

                 Unfortunately, most of the congeners mentioned above are
                 difficult to analyze, even by high resolution gas
                 chromatography.    These compounds tend to co-elute with
                 other congeners.      Special separation techniques are
                 required to identify them. To date, the only separation
                 techniques reported in the literature are (1) carbon
                 column   absorption     (Stalling    et    al.   1983)    and
                 multidimensional gas chromatography (Duinker et al.
                 1988).    The purpose here is to present results of
                 analyses of river sediments for these toxic PCB congeners
                 using a multidimensional chromatography technique.

                 MATERIALS AND METHODS

                 Sediment samples were col  lected from the Sheboygan River,
                 a Wisconsin tributary to Lake Michigan.        The river is
                 polluted with PCBs from the mouth to about 22.6 km (14
                 miles) upstream.      Waste hydraulic fluids containing
                 Aroclor 1248 and Aroclor 1254, were the source of the
                 contamination (David 1990). The polluted se'?--tion of    the
                 river is a U.S. federal "Superfund" site as well as       one
                 of the Great Lakes "Areas of Concern" As defined by       the
                 U.S./Canadian International Joint Commission.

                 Sediment cores were collected in December of 1988         and
                 April of 1989 using a metal corer 90 cm (3 ft) long       and
                 7.6 cm (3 in.) in diameter. All samples were collected
                 ,near the original source of PCBs at Rochester Park (about
                 22.6 km upstream from the mouth). A hydraulic extruder
                 was used to remove the sediment from the corer.           The
                 cores were segmented into 15 cm sections and kept cool
                 (40C) until analysis.

                 Sediment was air dried and sieved to form a homogeneous
                 sample. PCBs were extracted from 50 g sediment samples
                 by soxhleting for eight hours with acetone and hexane
                 (1: 1) . Granular copper was: added to the soxhlet f lask to
                 insure   uniform     boiling    and    to    remove     sulfur
                 interferences. After concentrating the sample, anhydrous
                 sodium sulfate was added to remove water from the
                 extract. The hexane-acetone solution was then exchanged
                 for 2,2,4-trimethyl pentane (iso-octane) .         PCBs were
                 separated from other contaminants using Florisil and
                 silica gel. Hexane and 6% ethyl ether were the eluting
                 solvents used for the Florisil fractionation and hexane
                 for the silica gel fractionation. The first silica gel
                 fraction was subsequently used for PCB quantitation.
                 Hexane was exchanged with isooctane prior to gas
                 chromatographic analysis. Samples were concentrated to









                10 or 50 mL prior to injection of an aliquot into the gas
                chromatograph.

                PCB congeners were quantitated using a Siemens Sichromat
                2-8 multidimensional gas chromatograph. The instrument
                was equipped with two 30 m capillary columns (DB-5 and
                DB-1; J and W Scientific). Each column was connected to
                an individual "Ni electron capture detector. The columns
                were also contained in separate ovens. Eluate from the
                DB-5 column passed through the column's detector
                producing a chromatogram similar to that produced in
                conventional     high   resolution     gas    chromatography.
                However, when a 'IT-piece" connecting the DB-5 column to
                the DB-1 column was activated, eluate was diverted to the
                DB-1 column. Thus, a selected part of the eluate (e.g.,
                the eluate eluting between a specific time interval) was
                cut to the DB-1 column.       The cut is accomplished on a
                real time basis according to pneumatic (gas flow)
                differences between the two columns, the timing,of which
                is controlled by the analyst.

                The temperature of the oven containing the DB-5 column
                was ramped from 900C to 1600C at a rate of 200C per
                minute, then from 1600C to 2600C at a rate of 40C per
                minute. The final temperature was held for one minute.
                The temperature of the column containing the DB-1 oven
                was held at 1600C for 20 minutes and then increased at a
                rate of 40C per minute to 2400C. The final temperature
                was held for eight minutes. The injector temperature was
                2500C. the detectors were heated to 3000C. The carrier
                gas and make-up gas were hydrogen and nitrogen,
                respectively.

                By cutting a portion of the eluate after it passed
                through the DB-5 column to and through the less polar DB-
                1 column, increased separation of the compounds in the
                cut was obtained.     The chromatogram produced from the
                second (DB-1) column and its associated detector
                represented the response of the cut components, while the
                chromatogram of the first column (DB-5) represented the
                rest of the compounds in the injected mixture              (the
                chromatogram for the first column is interrupted by the
                cut).   For a further discussion of the instrument, see
                Duinker et al. (1988).

                For the eight congeners studied, pure standards were used
                for identification (based on retention times) and
                quantification. A detector limit of 1 ng/g, based on a
                signal to noise ratio of approximately 3, was made.
                Although replicate analyses were made to check results,
                no alternate method was available to confirm the
                identification of the congeners in the unknown.












               RESULTS AND DISCUSSION

               Results of the analyses of Sheboygan River sediment
               samples f or total PCBs and eight "toxic" congeners are
               summarized in Table 1.      Each of these congeners was
               .identified and quantified in at least one of the samples.
               Note that all sediment core samples analyzed had total
               PCB concentrations greater than 50 Ag1q.

               Congeners 118 (245-34), 105 (234-34), and 77 (34-34) were
               detected in all of the samples reported here.
               Concentrations ranged from about 5 to 1500 ng/g.        The
               remaining toxic congeners, 81 (245-4), 114 (2345-4), 167
               (245-345), 126 (345-34), and 169 (345-345), were detected
               less frequently in the analyzed samples. Concentrations
               of these congeners ranged from non-detectable to slightly
               over 100 ng/g.

               Very few measurements of toxic congeners in environmental
               samples have as yet been reported in the literature.
               Tanabe et al. (1987) reported various levels of congeners
               77 (34-34), 126(345-34), and 169 (345-345) in fish,
               marine mammals, and terrestrial animals in Japan.
               Duinker et al. (1988) reported seal blubber from the
               Dutch Wadden Sea contained 100, 800 and 5200 ng/g of
               congeners 77 (34-34), 118 (245-34) , and 105 (234-34) ,
               respectively.   Congeners 126 (345-3A), 81 (345-4), 167
               (245-345), and 114     (23 45-4) were all found in seal
              ,blubber at concentrations less than 10 ng/g. Note that
               for the Sheboygan sediments reported here, congeners 77
               (34-34), 118 (245-34), and 105 (234-34) were also found
               at concentrations higher than the rest of the congeners.
               To the best of the authors' knowledge, no other data on
               concentrations of these congeners in aquatic sediment
               have been published.

               Table 2 presents the weight percents of the congeners in
               the samples. Also included are the weight percents of
               congeners in several Aroclors as determined by Duinker et
               al. (1988). The weight percents of the toxic congeners
               in the sediments were generally lower than those found in
               Aroclors 1248 and 1254 (the primary PCB mixtures
               discharged to the river) and in the other Aroclor
               mixtures listed in Table 2. The weight percents of the
              ,most prominent toxic congeners (77,,118, and 105) were
               about one order of magnitude lower than the weight
               percents of these congeners in Aroclor 1248. Some weight
               percents for congener 81 (345-4) were higher than
               reported for the Aroclors, although the congener was
               detected only in four of the eight samples reported.

               Anaerobic reductive dechlorination of PCBs, similar to
               that reported by Brown et al. (1987), Quensen et al.
               (1988), and Rhee et al. (1989), has been reported to









               occur in Sheboygan River sediments           (David 1990).
               Dechlorination of non-ortho chlorinated congeners has
               been found to occur more readily than dechlorination of
               ortho   chlorinated PCBS      (Brown and Wagner        1990).
               Congeners 77 (34-34), 118 (245-34), and 105 (234-23) are
               all   non-ortho    chlorinated    PCBs,   suggesting     that
               dechlorination might play a role in the depletion.

               of the toxic congeners, congener 118 (245-34) was found
               in Sheboygan River sediments in the highest weight
               percent. Congener 118 is also found in all the Aroclors
               in Table 2 in the highest weight percent of the toxic
               congeners. It also appears to be the most common toxic
               congener in environmental samples.        It was f ound in
               highest    concentrations    (relative    to   other    toxic
               congeners) in seal blubber (Duinker et al. 1988) , as, well
               as in the sediments collected for this study. Congener
               118, unlike the other toxic       congeners, can of ten be
               resolved and quantified without multidimensional gas
               chromatography or other special techniques. It has thus
               been reported in a variety of matrices, such as fish and
               the blood of sport fish eaters (Maack and Sonzogni 1988;
               Fiore et al. 1989 and Sonzogni et al. 1991).

               The eight toxic PCBs studied here appear to be present in
               the sediment, but in relatively low concentrations
               compared to total PCBs or other more abundant
               congeners. The concentrations of the eight congeners are
               low relative to the congeners with which they co-elute as
               well. For example, toxic congener 77 (34-34) co-elutes
               with congener 110 (236-34), but 77, in the sediments
               studied here, made up less than 4 percent of the total of
               the co-eluting pair. It should again be emphasized that
               the toxic congeners have been so labeled because of their
               dioxin-like structure and the potential health effects
               of dioxin. More research needs to be conducted on the
               toxicological effects of the PCB congeners. The effects
               of exposure to environmental levels needs to be
               considered as well as toxicological endpoints other than
               that for dioxin.     For example, the neurotoxicological
               impacts of specific congeners or combinations of
               congeners needs to be studied. Perhaps new analytical
               toolst such as multidimensional gas chromatography, will
               help make such research more feasible.

               Acknowledgements: The authors gratefully acknowledge the
               sampling assistance of William Wawrzyn and Thomas Aartila
               of the Wisconsin Department of Natural Resources. This
               research was supported under a grant from the Wisconsin
               Coastal Management Program.










             Table 1. Concentration of various toxic PCB congeners in Sheboygan River
             sediment samples of different total PCB concentrations.


               Total PCB
               in sample                         Congener (IUPAC Number)
                (Ag/g)              77    118         105     167      114        126       169     81
                                                            (ng/g)

                          50.3      31    140         80      44       NDa        ND        ND      ND

                       1050        360    1480        490     80       110        10        19      90

                          76.9      24    129         27      ND       ND           9         9     ND

                          97.8      21    318         41      ND       ND         ND        ND      ND

                         256        46    395         15      ND       ND         ND        ND      75

                          73.4        5   103         11      ND       ND         ND        ND      ND

                          97.5      11    162           6     ND       ND         ND        11      ND

                         124        16       86       11      ND       ND         ND        ND      50



             "ND   not detected (concentration less      than 1 ng/g)








             Table 2. Weight Percent of Various Toxic PCB Congeners in
             .Sheboygan River Sediment Samples and Weight Percent of
              Congeners in Several Aroclors'.



                Total PCB
                in Sample                                Congener (IUPAC Number)
                 GLg/g)                77       118              167         114       126      169      81
                                                               (weight
                             50.3     0.06    0.27       0.15    0.09        - -b
                           1050       0.03    0.14       0.05    0.01        0.01     <0.01    <0.01    0.01
                             76.9     0.03    0.16       0.04                          0.01     0.01

                             97.8     0.02    0.33       0.04    -  -        - -
                            256       0.02    0.15       0.01    -  -        - -                         0.03

                             73.4     0.01    0.14       0.01    -  -        - -
                             97.5     0.01    0.17       0.01    -  -        - -                0.01
                            124       0.01    0.07       0.01    -  -        - -                         0.04
                     Aroclor 1242     0.50    1.80       0.33    <0.01      <0.01      <0.01    <0.01  <0.01
                     Aroclor 1248     0.30    3.35       0.55    <0.01      <0.01      <0.01    <0.01  <0.01
                     Aroclor 1254 <0-01       8.45       2.03      0.05     <0.01      0.08     0.08   <0.01
                     Aroclor 1260 <0-01       1.15       0.08      0.15     <0.01      0.05     0.05   <0.01

              'Aroclor weight percents f rom. Duinker et al. 1988
              bCongeners not detected in samples (weight percents less than 0.002%).










                References


                Brown JF, Bedard DL, Brennan MJ, Carnahan JC, Feng H,
                      Wagner     RE    (1987)     Polychlorinated       biphenyl
                      dechlorination in aquatic sediments. Science
                      236:709-712
                Brown JF, Wagner RE    (1990) PCB movement, dechlorination,
                      and detoxication in the Acushnet Estuary Environ
                      Toxicol Chem 9:1215-1233
                David M (1990) PCB congener distribution in Sheboygan
                      River sediment, fish and water.        MS Thesis, Water
                      Chemistry Program, Univ of WI, Madison 159p
                Duinker JC, Schultz DE, Petrick G (1988) Multidimensional
                      gas chromatography with electron capture detection
                      for    the    determination      of   toxic      congeners
                      polychlorinated biphenyl mixtures.             Anal Chem
                      60:478-482
                Erickson MD      (1986)   Analytical Chemistry of' PCBs.
                      Butterworth Publishers, Stoneham, MA
                F iore BJ, Anderson HA, Hanrahan LP, Olson LJ, Sonzogni
                      WC (1989) Sport fish consumption and body burden
                      levels of chlorinated hydrocarbons; A study of
                      Wisbonsin anglers. Archives of Environ Health
                      44:82-88
                Maack     L,     Sonzogni     WC     (1988)     Analysis       of
                      polychlorobiphenyl congeners in Wisconsin fish.
                      Arch Environ Contam Toxicol 17:711-719
                Mullin MD, Pochini CM, McCrindle S, Romkes M, Safe SH,
                      Safe LM (1984) High resolution PCB analysis:
                      Synthesis and chromatographic properties of all
                      209 PCB congeners. Environ Sci Technol 18:468-476
                Safe S, Safe L, Mullin M (1985) Polychlorinated
                      biphenyls: Congener-specificanalysis of commercial
                      mixture and a human milk extract. J Agric Food
                      Chem 33:24-29
                Safe S (1987) Determination of 2,3,7,8-TCDD toxic
                      equivalent factors (TEFs) ; Support for the use of
                      the in vitro AHH induction assay.             Chemosphere
                      16:791-802
                Sonzogni W, Maack L, Gibson T, Degenhardt D, Anderson H,
                      Fiore B (1991) Polychlorinated biphenyls congeners
                      in blood of Wisconsin sport fish consumers. Arch
                      Environ Contam Toxicol, in press
                Stalling DL, Smith LM, Petty JD, Hogan JW, Johnson JL,
                      Rappe      C,   Buser    HR     (1983)    Residues       of
                      polychlorinated           debenzo-p-dioxins            and
                      debenzofurans in Laurentean Great Lakes fish.           In:
                      Tucker RE, Young AL, Gray AP (ed) Human                 and
                      environmental risks of chlorinated dioxins              and
                      related compounds, Plenum
                Tanabe S, Kannan N, Subramanian A, Watanabe S, Tatsukawa
                      R (1987) Highly toxic coplanar PCBs: Occurrence,
                      source, persistency, and toxic implications to
                      wildlife and humans. Environ Pollut 47:147-163
                Webb  RG, McCall AC (1972) Identities of polychlorinated
                      biphenyl isomers in aroclors. J Assoc Offic Anal
                      Chem 55: 746-752









                PCB Dechlorination in the Sheboygan River, Wisconsin

          Extended Abstract to be Published in Proceedings of a Workshop on
                  Biological Remediation of Contaminated Sediments


                    by William C. Sonzogni and Margaret M. David
                 Laboratory of Hygiene and Water Chemistry Program
                               University of Wisconsin
                                 Madison, WI 53706


              The Sheboygan River in Wisconsin flows into Lake Michigan at

          the city of Sheboygan, located 90 km north of Milwaukee. Due to

          the high concentrations of PCBs in the river sediment, the

          Sheboygan River and Harbor area has received national attention.

          The main source of contamination was from a die casting plant

          located in the Village of Sheboygan Falls. The contamination

          source area is about 22 km upstream from the mouth of the river.

               Hydraulic fluids containing PCBs were used by the die

          casting plant from 195.9 to 1971 (11). Apparently, a large fire

          occurred at the plant prior to 1959 that was caused by combustion

          of the hydraulic fluids then in use. Fluids containing PCBs were

          subsequently put in use because of their fire resistance. Based

          on interviews and available records, a product called Pydrol F9

          was used between 1959 and 1969 and a product called Chemtrend

          HP30 was used between 1970 and 1971. Pydrol F9 contains Aroclor

          1248, while Chemtrend HF30 contains, mostly Aroclor 1254 with a

          small percentage of Aroclor 1248. In 1971 the use of hydraulic

          fluids containing PCBs ceased.

              Material from the plant (oil soaked rags, hoses and other

          refuse) and soil from around the plant was used to construct a

          low dike at the edge of the Sheboygan River. The dike sloped at






 4o



          a 45 degree angle to the river, so erosion of the diked material
          into the river occurred relatively easily (11). Concentrations

          of PCBs in the soil samples were as high as 120,000 gg/g. The

          die casting plant is the only known major source of PCBs to the
          river, therefore, the congeners deposited in the sediments were

          most likely the components of Aroclor 1248 and 1254.

               In an article in science it was reported that biological

          reductive dechlorination of PCBs was occurring in Hudson River

          sediment. There was evidence that anaerobic dechlorination was

          also occurring in other aquatic sediments, including Sheboygan

          River sediments (4). The Sheboygan evidence was based on

          observations of chromatograms obtained from the U.S. Army Corps

          of Engineers.

               As result of the published reports that dechlorination could

          occur and because of new analytical capabilities to do congener

          specific PCB analysis, research was begun to examine the

          distribution of PCB congeners in the Sheboygan River sediment and

          to determine whether anaerobic dechlorination may be occurring.

          The intent was to determine the congener distribution in

          Sheboygan River sediment and assess whether some transformations

          had occurred. Results of congener distributions in Sheboygan

          River sediment relative to distributions in Aroclors will be

          summarized below as well as information on the occurrence of

          "toxic" congeners. Finally, a summary of evidence so far to

          degrade PCBs in the laboratory using bacteria from Sheboygan

          River sediments will be made.

               Total PCB concentrations ranged from 1586 Ag/g found









          downstream from the source to 0.04 gg/g above the source
          (considered to represent background levels). Although there is

          considerable variation in the sediment PCB concentrations, in

          general, the values were found in areas of sediment deposition in

          the river. In the individual cores, the top segment of core (0-

          15 cm) and the bottom segment of core (45-60) cm had relatively

          low concentrations of PCBs. The highest concentrations were

          found in the 15-45 cm segments.

               Sediment samples were also analyzed for PCB congeners using

          high resolution gas chromatography. Samples containing total PCB

          concentrations greater than 50 Ag/g appeared to be enriched with

          the lower chlorinated congeners whereas those with less than 50

          Mg/g PCBs were not. Samples containing 50 gg/g or more PCBs and

          significantly higher concentrations of mono- and di- chlorinated

          congeners when compared to aroclors 1248 and 1254 which were

          originally introduced into the river. Using a mulltivariate ANOVA

          statistical test, samples containing PCB concentrations less than

          50 Ag/g, were found to be statistically different from Aroclor

          1248, Aroclor 1254 and an equal parts mixture of Aroclors 1248

          and 1254.(p values were <0.05). In sediment samples containing

          PCB concentrations less than 50 Ag/g, the homolog patterns were

          more similar to the patterns of the Aroclor'1248 and 1254 than

          the more contaminated sediments.

              The most prominent congeners in the sediments with total PCB

          concentrations greater than 50 Ag/g were (IUPAC #) 5/8, 17,

          16/32, 47/48, and 28/31. Relative to the, original Aroclors,

          particularly high concentrations of congeners 5 and 8 were seen







          (congeners 5 and 8 co'elute). To confirm the presence of
          congeners 5/8 six of the samples containing high concentrations
          of PCBs (342.8 Ag1g on average) and high concentrations of
          congener 5/8 (43.7 percent on average) were analyzed using an

          electron impact gas chromatography mass spectrometer. All

          samples contained high concentrations of dechlorinated congeners.

          In samples containing less than 50 gg/g of PCBs, the most

          prominent congeners were similar, but their weight percents were

          generally reduced.

              The result from the sediment analyses indicate that the PCB

          congeners and their respective weight percentages in sediments

          with high PCB concentrations are significantly different from the

          Aroclors originally introduced into the river. Although

          physical-chemical processes such a sediment-water partitioning

          are important in determining the distribution of congeners in

          sediments, it is unlikely that it is the dominant process

          influencing the distribution of congeners. Sediment-water

          partition coefficients generally increase with molecular weight

          and thus an enrichment of the higher chlorinated congeners in the

          sediments not lower chlorinated congeners as observed would be

          predicted.

               Diffusion of congeners out of the sediment and into the

          water is slow relative to sedimentation rates and is inversely

          related to partition coefficients (7,8). Therefore, a

          distribution enriched in the higher chlorinated PCBs would be

          predicted (opposite of what was observed in this study).

              Another possibility to account for the change in congener








          patterns in abiotic chemical reactions. PCBs have been shown to
          undergo abiotic reductive dechlorination in the labor atory;

          however, the conditions in the laboratory (high temperatures,

          excess base, and the presence of a catalyst) are considerably

          different from those in the environment (3). In general, it is

          thought that there are very few abiotic pathways which completely

          mineralize organic contaminants (1).

               It is possible, however, that the Aroclors undergo

          biological dechlorination. Recent work by Quensen et al. (9),

          Chen et al. (6), Rhee et al. (10), and Brown et al. (4,5) suggest

          that PCBs can undergo.anaerobic microbial degradation. Several

          results in this study suggest such a process.

               First, there is a shift in the congener pattern from the

          higher chlorinated congeners to the lower chlorinated congeners

          as observed by Quensen et al. (9) in a laboratory experiment and

          as noted by Brown et al. (4,5) in a field study of hudson River

          sediments. This enrichment in lower chlorinated congeners cannot

          be accounted for by physical-chemical partitioning relationships

          or diffusion processes.

               Second, there appears to be a structural selectivity as to

          which congeners are depleted in the sediment. Congeners

          containing chlorines in the ortho position are enriched, whereas

          congeners containing chlorines in the meta or para position are

          depleted. This is consistent with the results obtained by

          Quensen et al. (9) and Brown et al. (4) in their anaerobic

          microbial dechlorination work.

               Third, several congeners are found in abundance that would







          not be expected based on physical-chemical partitioning
          relationships or on the original weight percentages present in

          the Aroclor mixtures. In sediment samples with concentrations of

          PCBs above 50 Ag/g, congeners that were significantly enriched

          are 5/8, 19, 17, 24/27, 16/32, 26, and 47/48. Congeners that

          were significantly depleted are 18, 28/31, 52, 44, 70/76, 66/95,

          56/60, 101, 77/110, 132/153/105, and 138/163. These changes are

          comparable to Brown et al.'s (5) findings.

               Fourth, the concentration of PCBs in the sediment appears to

          be important. Microbial degradation is often restricted to areas

          of high substrate concentrations. For example, toluenel xylene,

          and naphthalene are metabolized by bacteria at high

          concentrations but not at low concentrations (2). Threshold

          concentrati ons exist for many contaminants and are the minimum

          concentration of a chemical which is needed to support growth of

          a microbial population (2). Below the threshold concentration,

          additional energy sources must be found to support growth of the

          population, since the organism is no longer able to completely

          mineralize the substrate. Based on the different chromatographic

          patterns seen for high and low concentrations of PCBs in the

          Sheboygan River, it may be that a threshold concentration exists

          for the anaerobic dechlorination of PCBs.

               To confirm that microbial processes are actually responsible

          for degrading PCBs, laboratory experiments have been conducted

          similar to those reported by Quensen et al. (9) and Rhee et al.

          (10). Using bacteria extracted from Sheboygan sediments,

          degradation was attempted using growth medium and anaerobic








          conditions suitable for microbial dechlorination of PCBs.

          However, to date no dechlorination has been observed in the
          experiments. The reason for the lack of dechlorination activity
          is not clear, but it is suspected that the conditions that favor
          degradation are very complicated (e.g., may involve very precise
          Eh conditions and may involve several different species or stains

          or organisms) and may be difficult to consistently reproduce in

          the laboratory.

              Finally, Sheboygan sediments have been analyzed for the

          presence of non-ortho or coplanar PCBs. These congeners are

          believed to be the most toxic (at least in terms of dioxin like

          toxic properties), but generally coelute with other congeners

          using capillary column gas chromatography. A multidimensional

          ("heart cutting") gas chromatograph that uses two high resolution

          columns in series was used to separate coeluting congeners.

          Results to date indicate that several congeners of toxicological

          interest are found in sediment samples, albeit at low

          concentrations. Congeners 118, 105 and 77 were detected in 83

          percent of the samples analyzed, at average concentrations of

          about 0.25, 0.06, and 0.04 Mg/g respectively. The average

          composition of these congeners was 0.13, 0.03, and 0.02 percent,

          respectively. congeners 81, 114, 167, 126, and 169 were also

          detected in some of the samples all at concentrations less than

          0.03 Ag1g. While the concentrations of these congeners are low

          relative to total concentrations of PCBs or to the congener they

          coelute with, the fact that they are present may be important

          toxicologically. Research is  ongoing in this area.








               This work was supported by grants from the Wisconsin Coastal

         Management Program and the Wisconsin Sea Grant Program.



         References




           1. Alexander, M. (1981). Biodegradation of chemicals of

               environmental concern. Science, 211: 132-138.



           2.  Alexander, M (1985) Biodegradation of organic chemicals.

               Environ. Sci. Technol., 18:106-111.



           3.  Boyer, S.K., J McKenna, J. Karliner, and M. Nitsberger

               (1985). A mind and efficient process for detoxifying

               polychlorinated biphenyls. Tetrahedron Letters, 26; 3677-

               3680.




           4.  Brown, J.F., D.L. Bedard, M.J. Brennan, J.C. Carnahan, H.

               Feng, and R.E. Wagner (1987a). Polychlorinated biphenyl

               dechlorination in aquatic sediments. Science, 236: 709-712.



           5.  Brown, J.F., R.E. Wagner, H. Fend, D.L. Bedard, M.J.

               Brennan, J.C. Carnahan, and R.J. May (1987b). Environmental

               dechlorination of polychlorinated biphenyls. Environ.

               Toxicology and Chemistry, 6: 579-593.



           6. Chen, M., C.S. Hong, B Bush, and G.Y. Rhee (1988).

               Anaerobic biodegradation of PCBs by bacteria from Hudson










                River sediments. Ecotoxicology and Environ. Safety, 16: 95-

                105.



            7.  DiToro, D.M., J.M. Jeris, and D. Clarcia (1985)'. Diffusion

                and partitioning of hexachlorobiphenyl in sediments.

                Environ. Sci. Technol., 19: 1169-1172.



            8.  Fisher, J.B., R.L. Petty, and W. Lick (1983) Release of

                polychlorinated biphenyls from contaminated lake sediments:

                flux and apparent diffusivities of four individual PCBs.

                Environ. Pollut. Series B., 5: 121-132.



            9i  Quensen, J.F., J.M.'Tiedje, and S.A. Boyd (1988). Reductive

                dechlorination of polychlorinated biphenyls by anaerobic

                microorganisms form sediments. Science, 24211 752-754.



           10.  Rhee, C.Y.j B. Bush, M.P. Brown, M. Kane, and L. Shane

                (1989). anaerobic biodegradation of polychlorinated

                biphenyls in Hudson River sediments and dredged sediments in

                encapsulation. Water Research, 23: 957-964.



          11. Wisconsin Department of Natural Resources (1989). Sheboygan

                River remedial action plan. Environmental Quality Division,

                Madison, Wisconsin.








































                        SUMMARY OF LABORATORY EXPERIMENTS TO



                     DECHLORINATE PCBs USING BACTERIA EXTRACTED



                           FROM SHEBOYGAN,RIVER SEDIMENTS









              In May of 1989 the first experiments were run to induce

         dechlorination of PCBs in the laboratory. Anaerobic bacteria

         were extracted from three sediment cores using the procedure that

         will be described subsequently with the exception that a chopped

         mean anaerobe media was used. PCBs were added to nine vials to

         produce concentrations of 60 ppm, and to nine vials to produce

         concentrations of 600 ppms (to give 18 incubating vials,

         excluding controls).

              After initial PCB analysis, this run was abandoned for the

         following reasons. First, the sediment from which the anaerobes

         were extracted had low PCB concentrations (18 ppm) so that

         dechlorinating anaerobes may not have been established in the

         sample. Second, the media used (a standard chopped meat

         carbohydrate based anaerobic media) was different than used by

         Quenson et al. (1988) in their original work. This media

         presented sampling and extraction problems for PCB analysis.

         Thus, a new experiment was started.

              In the second experiment medium was prepared after Quensen

         (1988 and personal communication, Michigan State University,

         1989).









            The composition of the medium (in mg/L) is listed below:

                                       KH2PO4                 270

                                       K2HP04                 350

                                       NH4Cl                  530

                                       CaC12                    15

                                       MgC12                  100

                                       MnC12                     0.5

                                       ZnCl2                     0.05

                                       CUC 12 - 2 H20            0.038

                                       COC12 - 6H20              0.05

                                       FeC12 - 4H20              20

                                       H3BO3                     0.05

                                       NaMn04 - 2H,20            0.01

                                       N'C12 - 6H20              0.05

                                       Na2SeO3 - 5H20            0.05

            The medium was not buf f ered with NaHC03 nor              reduced with sulfide,

            as these additions were noted in Quensen's                procedures as

            optional. After autoclaving for about thirty minutes, the pH of

            the mineral medium was adjusted to 7.0 with dilute sodium

            hydroxide.

                   A sediment sample was obtained from an area of known high

            PCB contamination with the expectation that it would contain

            bacteria capable of dechlorinating PCBs. An additional sample

            was obtained from a site, upriver from the contamination source,

            to serve as an uncontaminated sediment source. The samples with

            high PCB contamination, which tested anaerobic to *methylene blue,

            were placed in anaerobe jars with 10 mL of sterile water.









               Twenty grams of uncontaminated sediment and 35 mL mineral

          medium were added to each of eight serum bottles in a CO     /H2 glove

          box. The bottles were left open for 24 hours in the glove box,

          injected with 50 gL ethanol, sealed, incubated at 370C until

          methane production was observed, and, after two weeks, were

          autoclaved at 1210C for 1 hr.

               Anaerobic microbes were obtained from the PCB-contaminated

          sediment by first transferring the sediment from the anaerobe jar

          to a glove box. In the glove box (C02/H2 atmosphere) a sample of

          several hundred grams of PCB-contaminated sediment was shaken

          vigorously with 500 mL of the mineral medium. After 15 min

          settling, the liquid was decanted and saved. Aliquots (35 mL) of

          the decanted liquid were added to each of the eight autoclaved

          serum bottles of uncontaminated sediment.

               Aroclors 1248 and 1254 (1:1 mixture dissolved in

          dimethylsulfoxide) were added to serum bottles (containing

          sediment and inoculated medium) in two concentrations. The

          resulting concentrations of PCBs in the vials were 55 ppm and 436

          ppm. Duplicate vials were prepared at each concentration. The

          remaining four bottles, used as controls, were inoculated with

          anaerobic medium, but were then twice autoclaved for 15 minutes

          before addition of the Aroclor standards. All eight sealed serum

          bottles were stored in the dark at room temperature (23-260C).

               Part way through the experiment it was feared that the

          sediment sample from which the bacteria were extracted may not

          have had enough dechlorinating bacteria. Therefore, an

          additional sediment sample was obtained to maximize the









         probability of the existence of dechlorinating anaerobes. The

         sample obtained was subsequently analyzed for PCBs and it

         contained more than 100 ppm of PCBs. Therefore, this sample

         should have contained dechlorinating bacteria. This sediment was

         extracted and the extract added to the media as described above.

         However, PCBs were spiked to give a resulting concentration of

         436 ppm. Two vials were set up this way (no additional controls

         were prepared).

              Samples of slurry, 2 mL each, were withdrawn from the vials

         for PCB analysis on an approximately monthly schedule. The

         .samples were withdrawn into centrifuge tubes with 2 mL acetone

         and 10 mL hexane, agitated on a mechanical shaker for 10 min,

         decanted, agitated with an additional 10 mL hexane for 10 min,

         decanted and the liquid added to the previous extracts. The

         liquid was then blown down to near dryness and taken into about 3

         mL iso-octane. The iso-octane extracts were then each eluted

         through a column of 8g f lorisil and 2g Na2SO4 with 200 mL 6%

         ether/hexane. An aliquot of the sample was diluted in iso-octane

         until packed column GC screening indicated a suitable

         concentration for congener analysis with capillary GC. The

         samples were then analyzed for PCB congeners using a Hewlett

         Packard 5880 gas chromatograph equipped with a

         60 m DB-5 coated capillary column.

              The PCB concentrations measured in the monthly analyses were

         only about 30 percent of the expected concentration. Apparently,

         the procedure used to extract PCBs from the media was incomplete.

         However, the distribution of PCB congeners that were extracted









         were nearly identical to the PCB congener distribution of the

         original Aroclors added. Cross streak plate tests of the samples

         after three months did reveal anaerobic activity (Clostridium and

         Bacillus organisms were tentatively identified). Consequently,

         despite'anaerobic conditions, anaerobic dechlorination apparently

         did not occur (at least not measurably) in this experiment.

              Part way through the experiment, the two vials that were set

         up using extracts from the sediments with PCBs greater than 100

         ppm were opened in the glove box and cysteine sulfide was added

         along with 40 mL of ethanol. The cysteine sulfide was added in

         an amount equivalent to the amount of sodium sulfide specified by

         Quenson (Michigan State University, East Lansing, MI, 1989;

         personal communication) as an optional addition. These chemicals

         ensure that fully reducing conditions are occurring in the

         sample. It was thought that this addition would remove any

         uncertainty posed by not originally adding the sulfide.

              After seven months the incubation was ended. All analyses

         up to this point indicated that no dechlorination was occurring,

         although the amount of PCBs extracted from subsamples was always

         less than expected. Either the vials were subsampled in such a

         way that the subsample obtained was unrepresentative (unlikely),

         or the extraction procedure was not fully removing the PCBs from

         the sediment/media mixture.

              For a final analysis for PCBs after the seven month

         incubation, special procedures were taken to ensure better

         extraction of the sediment/media mixture. The vials (2)

         containing bacteria extracted from sediment with high PCBs and to









         which the sulfide was added were analyzed by this special

         procedure. These vials were most likely to show dechlorination.,

              PCBs were first extracted from the decant of the vial with

         50 mL of hexane. The extraction was repeated four times and the

         extract was saved. The residue left in the vial was then

         vigorously shaken with 35 mL of acetone in order to suspend all

         the material. The vial was then sonicated for three minutes to

         break up the particles as finely as possible to ensure good

         contact with the solvent. After the contents settled, the

         acetone was poured off and saved with the hexane extracts. This

         acetone extraction procedure was repeated three more times, so

         that the sonicated sediment/media mixture was extracted a total

         of four times with acetone.

              The sediment/media mixture was further extracted with 35 mL

         of hexane. The mixture was sonicated for three minutes during

         each of the four successive extractions. After settling, the

         decant from each extraction was saved.

              For PCB analysis, all the decants following extraction were

         mixed together. The resulting mixture was concentrated to 10 mL

         by rotoevaporation. The concentrated solution was cleaned up on

         a co lumn containing 22.0 g of florisil and two I cm layers of

         Na2SO4 (to remove water and other interferences). The.column was

         eluted with 220 mL of 6% either/hexane. The solvent leaving the

         column was evaporated by blow-down. Iso-octane was added to

         adjust the volume to 250 mL. Finally, a 1.0 mL aliquot was

         diluted to 50 mL with iso-octane.

              Following screening by packed column chromatography, the









         iso-octane extracts were analyzed by capillary column gas

         chromatography. For the two vials analyzed by this procedure,

         88% and 89% of the PCBs were recovered. Thus, repeated

         extractions of sonicated samples was effective at extracting PCBs

         from the sediment/media mixture.

              Despite the good recoveries, no evidence was found of

         dechlorination. Congener patterns closely resembled the

         distributions of congeners originally spiked into the vials.

         Consequently, none of the laboratory experiments produced

         evidence that bacteria extracted from Sheboygan River sediments

         produce dechlorination.

              Despite the negative results of the experiments conducted,

         it is believed that more work would produce laboratory evidence

         of dechlorination. Several other laboratories around the country

         have now reported successful laboratory PCB dechlorination

         experiments. Dr. G-Yull Rhee of the New York Department of

         Health has now duplicated Quenson et al.'s experiments with

         bacteria extracted from the Hudson River (Rhee et al. 1989).

         However, Dr. Rhee (New York State Department of Health and School

         of Public Health, SUNY at Albany, Albany, NY, 1990; personal

         communication) has indicated that getting the bacteria to grow

         properly is very tricky. He indicated that in his work it would

         not be unusual to have only one of four replicates show

         dechlorination. Both Dr. Rhee and Dr. Quenson agree that the

         factors that may effect dechlorination in the laboratory are not

         clear. For example, it is not clear whether the addition of

         sulfide and ethanol are necessary or what electrode potential









         should be obtained for the organisms to grow. Both believe that

         a consortium or organisms, rather that an individual species, is

         responsible for the dechlorination activity. Thus, while these

         studies laboratory experiments were not positive, it is believed

         that additional work, including a larger number of replicate

         samples, would produce evidence of dechlorination. The

         laboratory experiments of this study do show, however, that the

         conditions under which dechlorination occurs are not clear and

         that more needs to be learned if anaerobic dechlorination is to

         be used as a bioremediation tool.




         References




         1.   Quensen, J.Fi, J.M. Tiedje, and S.A.Boyd (1988). Reductive

              dechlorination of polychlorinated biphenyls by anaerobic

              microorganisms form sediments. Science, 242: 752-754.



              Rhee, C.Y., B. Bush, M.P. Brown, M. Kane, and L. Shane

              (1989). Anaerobic biodegradation of polychlorinated

              biphenyls in Hudson River sediments and dredged sediments in

              encapsulation. Water Research, 23: 957-964.


































             An Evaluation of the Potential for PCB Dechlorination in

                           Confined Disposal Facilities






















                                     Wisconsin State Laboratory of Hygiene
                                                   University of Wisconsin
                                                        Madison, WI 53706











         EXISTING CONFINED DISPOSAL FACILITIES

              Dredging of Great Lakes harbors to maintain navigation

         dates back to the early 1800's. Natural siltation from tributary

         rivers prompted periodic dredging, and dredging has been

         generally accepted as necessary and normal harbor maintenance.

         Typical disposal methods were to side cast the dredgings to

         adjacent areas or, mor e commonly, to deposit the material into

         deeper waters.

              Until the mid 1960's immediate  economic concerns  governed

         the focus of this disposal. However,  a new awareness of lake

         eutrophication problems led to a questioning of open water

         disposal techniques. The U.S. Army Corps of Engineers (1969) in

         a study titled "Dredging and Water Quality Problems in the Great

         Lakes" concluded that "in-lake disposal of heavily polluted

         dredging must be considered presumptively undesirable." In 1970,

         Congress generated PL 91-611 to create the Dike Disposal Program

         for the Great Lakes (Anonymous, 1989)., The intent was to hold,

         in a confined facility, a ten year supply of dredgings, and to

         initiate a Dredged Material Research Program (DMRP). The U. S.

         Army Corps of Engineers (COE) conducted the DMRP at their

         Vicksburg (Mississippi) Waterways Experimental Station (WES).

             The DMRP study was a comprehensive project, involving

         universities, private research laboratories, and other federal

         agencies. The main conclusions of the DMRP study, as indicated

         in U.S. Environmental Protection Agency (1987), were fourfold:









              1.   No single disposal alternative is suitable

                   for a region, nor should any single alternative be

                   dismissed from consideration.

              2.   Environmental considerations.are stronger than other

                   considerations to necessitate long.range regional

                   planning as a lasting, effective solution to disposal

                   problems.

              3.   Fears concerning short term release of contaminants to

                   site waters are unfounded especially if the geochemical

                   environment is not changed.

              4.   To be environmentally effective, a confined site must

                   retain a high percentage of finer particles, since this

                   is where contaminants are carried.

              Since 1978 there has been some additional dredged material

         research. However, the research focus has shifted from

         eutrophication to toxic substances. During the late 1970's and

         1980's many harbor sediments were found to contain high

         concentrations of heavy metals and organic pollutants, such as

         PCBs. The research has not, however, provided a clear

         understanding of the impact that toxic substances in a CDF may be

         having on the Great Lakes or what happens to toxic chemicals over

         time when placed.in a CDF.

              Existing confined disposal facilities (CDFIS) in the Great

         Lakes basin are listed in Table 1. Other than the Superior/

         Duluth facility (constructed in 1978), Wisconsin's CDFIS are all

         sited in or along Lake Michigan. In addition to the six

         Wisconsin facilities listed in Table 1, three additional sites









        have been proposed (Menominee, Sturgeon Bay, and Sheboygan) for

        construction. The U. S. COE is responsible for dredge and fill

        operations, including the operation of CDFIS. Guidelines and

        criteria for dredging and filling are, however, set by the U.S.

        Environmental Protection Agency (EPA).

             The Sheboygan Harbor was dredged by the COE in the mid

        1950 Is and annually up to 1969 when dredging stopped due to
        controversy over the presence of heavy metals. Some contaminated

        harbor sediments were removed in 1981 and 1984. As stricter

        disposal regulations, particularly for PCBs have been put into

        effect, dredging has been halted due to the lack of a means of

        disposal.

        GENERAL DESIGN OF CONFINED DISPOSAL FACILITIES

             Each CDF is unique to the needs of the individual site.

        Upland CDFIS generally have earthen dikes, whereas in-lake CDFIS

        generally are encased by stone dikes to withstand wave action.

        The overall design goals are to hold within the facility as high

        a percent of the sediment particles dredged as possible.

             The design of CDFIS has evolved since the first facility was

        constructed in the early 1960's to a point where there are now

        certain design similarities (U.S. Army Corps of Engineerst

        1986). Designs now allow limited permeability so precipitation

        can drain into the lake. As environmental requirements became

        more stringent, steel sheet pile was incorporated into the

        design. This pile was used in the three Wisconsin CDFIS, thus

        further ensuring the dredged material is retained in the dike.









              Recently constructed dikes have utilized a core of limestone

         which "cements" over time. A layer of locally  available lake

         sand is now typically incorporated by layering the sand. on the

         disposal side of the dike. This sand is uniform, has low

         permeability, and will not change or deteriorate in time. Some

         designs incorporate a weir with a skimmer to retain possible

         sediment overflow as the permeability of the CDF approaches zero.

         PCBIS IN CDFIS

              Very little data are available on PCB concentrations within

         CDFIS. Published data exists for only a few sites. The

         concentrations of PCBIS would be expected to vary from one CDF to

         another depending on the concentration of PCB's in the sediment

         which was dredged. Since PCB's in sediment can vary widely even

         within a small area (sediments are not homogeneous),

         concentrations within CDF's also undoubtedly vary widely.

              The few data that are available on PCB's measured from

         samples collected from CDF's indicate relatively low total PCB

         levels. Clark (1988) reported a value of 1.1 Ag/g. from

         sediments in the Calumet harbor CDF. Rathburn (1988), in a study

         of the Saginaw Bay diked facility, reported concentrations of

         PCB's congeners in water collected from inside the facility

         (indike), immediately outside the facility in Saginaw Bay

         (outdike), and at an adjacent site in the Saginaw River channel.

         An example of these data are shown in Table 2. The data show PCB

         concentrations are higher in water inside the facility than

         outside the facility. However, concentrations in water outside

         the facility are generally higher than PCBs in the reference









         station (Saginaw River channel).

              The specific congeners found in highest concentration in the

         Saginaw Bay study (Table 2) are similar to those found in highest

         concentration in the Sheboygan River. For example, co-eluting

         congeners 28/31 are the most prevalent in the Saginaw Bay study

         samples (Table 2), as they were in most Sheboygan samples.

             Whether or not PCB's leak from a CDF continues to be a major

         uncertainty.   The Saginaw Bay study was initiated to find the

         extent of leakage (as well biological uptake of the PCB's). The

         general conclusion of the study was that no appreciable leakage

         from the CDP was occurring.

              If polychlorinated biphenyls (PCBs) can be dechlorinated in

         river sediments, a logical question to ask is whether PCBs

         contained in confined disposal facilities (CDFs) also undergo

         bacterial dechlorination. If dechlorination does occur in

         confined disposal facilities, these facilities would be an

         attractive way of detoxifying dredged PCB contaminated sediment.

         In the case.of the Sheboygan River, these is interest in building

         a marina in Lake Michigan just north of the harbor. If such a

         facility could be designed in concert with the concept of a

         confined disposal facility, it would be especially attractive if

         it also promoted dechlorination of any PCBs in the sediment.

              In this study the likelihood of dechlorination occurring in

         confined disposal facilities along the Wisconsin shore of Lake

         Michigan was considered. Attempts were made to gather

         information on PCBs in confined disposal facilities to see if any

         evidence for dechlorination was available. Evidence for










         dechlorination in similar facilities, notably sewage sludge

         lagoons, was also sought. Finally, some samples were taken from

         the confined sludge facility off of Milwaukee and analyzed for

         PCBs.


              Two samples were collected from the Milwaukee CDF. The CDF

         might be expected to contain relative high amounts of PCBs

         because of the industry in the area. However, s pecific data on

         the PCB content of the dredged material originally placed in the

         facility was not found. Nor wasany data found from past

         measurements of PCBs made on samples taken from the facility.

              Two "soil" samples were taken from the Milwaukee facility

         with the permission of the Corps of Engineers. Because the soil

         was hard, crusted and contained surface vegetation, sampling was

         difficult. It was also unclear what would be a representative

         sample, or where to sample to get high PCB concentrations.

              The two samples analyzed had low PCB concentrations. No

         evidence of dechlorination was discernable from the PCB pattern.

         It was hoped that samples with higher PCB concentrations could be

         obtained, as such samples might show a greater tendency for

         dechlorination (analogous to Sheboygan River sediments). As it

         was, there was no evidence to suggest that dechlorination may be

         occurring, or at least occurring at a rate that would be

         noticeable.

              A question that comes up is whether sediments below the top

         of the facility are anaerobic. Deeper core samples would be

         desirable in this regard, as dredged material several feet below

         the surface might be anaerobic while near the surface the dredged









         material might be aerobic. Based on what was available in the

         literature, such measurements have not been made.

             one indication that dechlorination might not occur (at least

         not at, a meaningful rate), is that a'sewage lagoon with PCB

         contamination has been extensively studied and there is no

         evidence of any PCB dechlorination occurring there. The Madison

         Metropolitan Sewerage District has several lagoons that have been

         filled for many years and now pose a disposal problem because of

         the high concentration of PCBs (about 50 ppm) in the sludge. The

         lagoons, which do go anaerobic, have been extensively monitored

         over the years. However, monitoring results indicate that the

         PCBs have been very stable over the years (Nemke, J., Director,

         Madison Metropolitan Sewerage District, Madison, WI, 1990;

         personal communication). While there may be special conditions in

         these specific lagoons that prevent dechlorination, one might

         expect the conditions in an anaerobic lagoon to be conducive to

         dechlorination.

             Based on experiments attempting to achieve bacterial

         dechlorination in the laboratory, it is clear that the

         dechlorination process is highly sensitive to a variety of

         factors (some of which are not known or understood). Whether the

         right conditions are in place in CDFs or lagoons is not known.

         It is also clear that for dechlorination to occur in a measurable

         fashion or at a fast enough rate, PCB concentrations must be

         relatively high.  Unless "hot spots" or areas of high

         concentration occur in CDFs, concentrations may be too low to see

         measurable PCB dechlorination.









              Thus, an analysis of the available information suggest that

         dechlorination probably does not occur to any important extent in

         CDFs. More research should probably be conducted to confirm this

         hypothesis, including more rigorous sampling to assay PCBs in

         CDFs. Alternately, research might be conducted on how to promote

         or accelerate PCB dechlorination. Current research conducted on

         sediments from the Sheboygan River in pilot confined treatment

         facilities (in conjunction with Superfund cleanup of the river)

         may uncover some ways of promoting dechlorination that could be

         applied to CDFs.



         b:\cdfl.jan









           Table 1. Great Lakes Confined Disposal Facilities



                                                     size       Capacity
           Location                  Type         (acres)       (cu. yd.)-        Year

           Calumet Harbor,
           Chicago, IL                 L              42        1,300,000        19-84

           Michigan City, IN           U                3.3          25,000      1978

           Bolles Harbor, MI           L              24.6         335,000       1977
           Monroe County, MI           1             685        18,640,000       1978/
                                                                                 1981
           Frankfort, MI               U                           107,000       1982
           Detroit, MI                 1              80        1,900,000        1960
           Dickinson Island,    MI     U             174        2,000,000        1976
           Grand Haven, MI             U              36           310,000       1974
           Holland, MI                 U              27.7         370,000       1977
           Inland Route, MI            U                8.6          19,500      1982
           Monroe, MI                  L              89        4,200,000        1985
           Saginaw Bay, Village of
           Sebewaing, MI               U             180             84,000      1979

           Erie Pier, MN               L              82        1,000,000.       1978

           Buffalo, NY                 L              33        1,500,000        1968
           Buffalo, NY                 L              45        1,500,000        1972
           Buffalo, NY                 L              40        6,900,000        1977

           Cleveland, OH               L              56        2,760,000        1974
           Cleveland,   OH             L              88        6,130,000        1979
           Huron, bH                   1              63        2,150,000        1975
           Lorain, OH                  L              58        1,850,000        1977
           Toledo, OH                  L             150        5,000,000        1977
           Toledo, OH                  L             242        10,000,000       1976

           Erie, PA                    L              23        1,600,000        1979

           Green Bay, WI               1              60        1,200,000        1979
           Green Bay, WI               U             400           @300,000      1965
           Kenosha, WI                 L              25           750,000       1975
           Kewaunee, WI                L              28           500,000       1982
           Manitowoc, WI                              24           800,000       1975
           Milwaukee, WI,              1             454        1,600,000        1975

               L   In-lake adjacent    to land
               I    In-lake CDF island
               U    Upland









            Table 2. PCB Congener Concentrations (ng/L) in Saginaw Bay CDF
                        Water Samples (Prom Rathburn et al. 1988)



                                                                              Sacfina
            Congener Number         Indike            Outdike           River Channel


            004  + 010              0.27              0.04                    0.043
            006                     1.4               0.055                   0.023
            007                     0.061             0                       0.013
            012                     0                 0.032                   0.043
            013                     0.08              0                       -----
            016                     1.2               0.1                     0.04
            017                     3.4               0.19                    0.1
            018                     3.6               0.31                    0.092
            019                     0.3               0.032                   0
            022                     2                 0.13                    0.019
            024                     0.05              0.017                   0
            025                     ----              ----                    0.073
            @027                    0.39              0.036                   0.017
            029                     0.035             0.021                   0
            031  + 028              13                0.96                    0.4
            032                     3.5               0.43                    0.16
            033                     2.1               0.027                   0.052
            037                     ---               -----                   0.064
            040                     1.7               0.1                     0.029
            041  + b7l              3.1               0.23                    0.13
            042                     2.4               0.19                    0.083
            043                     0.5               0                       0
            044                     5.2               0.51                    0.28
            045                     1.2               0.093                   0.03
            046                     0.58              0.051                   0.027
            047  + 048              1.6               0.15                    0.17
            049                                       0.43                    0.23
            051                     0.36              0.013                   ----
            052                                       0.57                    0.17
            053                     1.2               0.1                     0.049
            056  +060               2                 0.25                    0.13
            063                     0.37              0.031                   0.0075
            064                     2.2               0.019                   0.092
            066                     3.4               0.4                     0.23
            070  + 076              3.4               0.41                    0.19
            074                     1.3               0.13                    0.068
            077                     0.65              0.068                   0.034
            081                     0.17              0.024                   0.0083
            082                     0.51              0.057                   0.021
            083                     0.29              0.027                   0 @
            085                     0.78              0.097                   0.052
            087                     0.98              0.13                    0.072
            089                     0.12              0.011                   0
            091                     0.79              0.073                   0.041
            092  + 084              2.6               0.25                    0.14











           100                   0.21            0                     0
           101                   2.1             0.28                  0.092
           105 + 132    153      2 -             0.35                  0.16
           107                   0.18            0.027                 0.013
           110                   3.5             0.4                   0.2
           118                   1.4             0.19                  0.1
           119                   0.2             0.017                 0.015
           128                   0.29            0.044                 0.092
           129                   0               0                     0.0083
           130                   0.14            0.023                 0.028
           134 + 114             0.3             0.04                  0.028
           135 + 144             0.16            0.047                 0.0092
           136                   0.12            0.017                 0
           137 + 176             0               0                     0
           141                   0.2             0.036                 0.028
           146                   0.27            0.044                 0.021
           149                   0.87            0.12                  0.083
           151                   0.23            0.033                 0.021
           156                   0.19            0.025                 0.011
           157 + 200             0               0.027                 0.019
           158                   0.021           0.033                 0.036
           163 + 138             1.4             0.25                  0.15
           167                   ----            ----                  0.0075
           170 + 190             0.48            0.083                 0.051
           172 + 197             0.16            0.021                 0.0092
           173                   0               0
           174                   0.23            0.047                 0.03
           175                   0.23            0.036                 0
           177                   0.16            0.031                 0.02
           178                   0.098           0.039                 0.0083
           180                   0.67            0.13                  0.049
           183                   0.22            0.033                 0.019
           185                   0               0                     0.0045
           187 + 182             0.29            0.059                 0.035
           189                   0.089           0                     0
           191                   0.4             0.016                 ----
           193,                  0.044           0.0093                0.011
           194                   0.14            0.031                 0.016
           196                   0.14            0.032                 0.016
           198                   0.084           0.019                 0.006
           199                   0               0                     0.0047
           201                   0.23            0.052                 0.033
           202 + 171             0.12            0.019                 0.0042
           203                   0.072           0.027                 0.018
           205                   0               0                     0.0033
           206                   0.11            0.037                 0.013
           207                   0.044           0.0093                0.0033
           208 +* 195            0.42            0.072                 0.026
           209                   0.005           0.001                 0.0008


           Total PCBIS          91.4            10.2                   5.2

           Blanks = rejected congeners; zeros = concentrations lower than
           analytical detection limit.












         REFERENCES:



         1.   Anonymous (1989) Diked Disposal Program - Resume (P.L. 91-
              611), Q/A section, U.S. Army Corps of Engineers, Buffalo,
              NY.

         2.   Clark, J.U., McFarland, V.A., and Dorkin, J. (1988)
              Evaluating Bioavailability of Neutral organic Chemicals in
              Sediments: A confined Disposal Facility Case Study. Water
              Quality 88 Seminar Proceedings., Feb. 23-24, Charleston, SC,
              U.S. Army Corps of Engineers, Vicksburg, MS.

         3.   Rathburn, J., Kreis, R., Lancaster, E., Mac, M., and Zabik,
              M. (1988) Pilot Confided Disposal Facility Biomon.itoring
              Study: Channel/Shelter Island Dike Facilityj Saginaw Bay,
              Bay City, Michigan, Report of the Environmental Research
              Laboratory, U.S. Environmental Protection Agency, Duluth, MN

         4.   U.S. Army Corps of Engineers (1969) Dredging and Water
              Quality Problems in the Great Lakes, Buffalo District,
              Buffalo, NY.

         5.   U.S. Army Corps of Engineers (1986) Appendix B, Dredged
              Material Disposal Facilities an the Great Lakes, Draft
              Environmental Impact Statement, Indiana Harbor Confined
              Disposal Facility, North Central District, Chicago, IL.

         6.   U.S. Environmental Protection Agency (1987) Report on Great
              Lakes Confined Disposal Facilities, draft, Great Lakes
              Program Office, Region V, Chicaao. IL.









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