Document gL5ggkwnJ9eZ16OzDw8Yn2pV

MEMORANDUM DEPARTMENT OK HEALTH, EDUCATION', AND U'CLF.W: > PUBLIC HEALTH SERVICE FOOD AND DAUC ADMINISTRATION to ; William Papageorge Manager, Product Acceptability BATE: July 21, 1976 FROM : Chief, Epidemiology Unit SUBJECT; Final Report of the Subcommittee on the Health Effects of Polychlorinated Biphenyls and Polybrominated Biphenyls. A copy of the above report is enclosed for your information. Cv+JJkj Frank Cordle, Ph.D., M.P.H. Chief, Epidemiology Unit, HFF-102 Bureau of Foods Food and Drug Administration Washington, DC 20204 erratum Table numbers proceed from 9 to Table 10 There is no HONS 211836 FINAL REPORT OF THE SUBCOMMITTEE ON THE HEALTH EFFECTS OF POLYCHLORINATED BIPHENYLS AND POLYBROM!NATED BIPHENYLS JULY 1976 DEPARTMENT OF HEALTH, EDUCATION AND WELFARE WASHINGTON, D.C. MOMS 211637 COSTENTS Members and Consultants, Subconmittee on the Health Effects of PCBs and PBBs .,, , ............................................. .... ,., v Subgroups and Their Members........... Vn Purpose and Specific Objectives.................. ..................................... ... INTRODUCTION ................................................................................................... 1 GENERAL SUMMARY AND CONCLUSIONS ........................................................ 4 CHEMISTRY..................................................................................................... 4 METABOLISM AND BIOCHEMICAL TOXICITY............................................ ANIMAL TOXICOLOGY............................................... 12 HUMAN EXPOSURE........................................................................................ 17 GENERAL RECOMMENDATIONS ........................................................................ 25 DETAILS) RECOMMENDATIONS........................................................................ 27 CHEMISTRY................................................................................................... 27 METABOLISM AND BIOCHEMICAL TOXICITY......................................... 28 ANIMAL TOXICOLOGY..................................................................................30 HUMAN EXPOSURE...................................................................................... 31 CHEMISTRY OP PCBs AND PBBe.................................................................. 34 Introduction............................................................................ 34 Chemistry of Chlorinated Biphenyls............................................35 A. Synthesis of Chlorinated Biphenyls........................... 35 B. Non-metabollc Alteration of Chlorlneted Blnhenyls............................................................................... .. 37 Oxidation ...................................................................... .. ^ Hydrolysis and Alcoholysis................................................ 38 Photochemistry.......................................................... 38 HONS 2118^8 C. Chemical Structure Related to Occurrence, Fate and Effects of Chlorinated Biphenyls.................... uz 0. Determination of PCS Residues......................................48 E. Chlorinated Dibeniofurans............................................ 61 Chemistry of Broolnated Biphenyls....................................................68 A. Comparison of Broainated Biphenyl with Chlorinated Biphenyls...................................................... 68 B. Non-metabolic Alteration of Bromiaated Biphenyls................................................................................ 70 Oxidation and Hydrolysis............................................... 70 Photochemistry...................................................................... 70 C. Broainated Biphenyls and Determination of Residues............................................................................. 72 D. Possibility of Bromiaated Dibeniofurans In Commercial PBB Mixtures........................................... 74 METABOLISM OP BIOCHEMICAL TOXICITY OF PCBs AMD PBBs......... 75 Effects of PCBe on Biochemical Function......................... 75 The Comparative Biochemical Toxicity of Aroclor 1254 and Firemaster BP-6 .......................................................................... 83 Metabolism of PCB and PBB Mixture .......................................... 87 The Metabollam of Individual PCSs......................................... 96 ANIMAL TOXICOLOGY ................................................................................. 100 Introduction...................................................................... 190 Acute Toxicity................................................................................... 100 Polychlorinated Biphenyls .................................................. 100 Polybrominated Biphenyls ...................................................... 102 Sub-Acute and Reproductive Effects of PCBs, PBBs and Chlorinated Dibeniofurans ......................................... 107 HONS 211839 A. Mammalian Sub-Acute Effects of ?CBs.......................lq7 B. Avian Sub-Acuce Effects of PCBs................................in C. Mammalian Reproductive Effects of PCBs................114 D. Mammalian Sub-Acute and Reproductive Effects of PBBs...................................... .......................... LIS E. Chlorinated Dibenzofurans - Mammalian Toxicity................................................................................... 122 F. Chlorinated Dibenzofurans - Avian Toxicity....................................................................................123 G. Brominated Dibenzofurans - Matsnallan Toxicity................................................................................... 124 H. Longterm Toxicity Including Tuaorogenssis .... 124 Chlorinated Biphenyl ...................................................... 124 Decachloroblphenyl ........................................................... 129 Chlorinated Dibenzofurans and Chlorinated Naphthalenes ........................................................................ 129 Immunosuppressive Effects of PCBs and PBBs ... 129 Mutagenicity and Teratology ...................................... 130 Polychlorinated Biphenyls .................................... 130 Teratology ...................................................................... 131 Chlorinated Dibenzofurans .................................... 132 Toxicity of Chlorinated Naphthalenes .................. 133 HUMAN EXPOSURE TO POLYCHLORINATED BIPHENYLS......................... 140 Introduction.........................................................................................1^0 Dietary Exposure to PCBs............................................................. 1^0 PCB Residue in Fish........................................................................ 1^4 PCB Exposure in Rice Oil............................................................. 160 Occupational Exposure to PCBs .................................................. 168 HONS 211840 Air and Water Exposure to PCSs 182 Residues of PCS in Human Tissue and Hilk.........................184 HUMAN EXPOSURE TO POLYBROMINATED BIPHENYLS...........................187 Introduction.......................................................................*............... 137 Human Exposure................................................................................... 138 HONS 211841 SUBCOMMITTEE ON THE HEALTH EFFECTS OF POLYCHLORINATED BIPHENYLS AND POLYBROMINATED BIPHENYLS Chairperson Albert C. Kolbye, Jr., M.D. Food and Drug Administration Members (M) and Consultants (C) Dr. John Buckley (C) Environmental Protection Agency Mr. Jerry Burke (M) Food and Drug Administration Dr. Frank Cordle (M) Food and Drug Administration Mr. Paul Corneliussen (M) Food and Drug Administration Dr. David Firestone (M) Food and Drug Administration Dr. Lawrence Fishbeln (M) Food and Drug Administration Dr. Gary FIsm (M) National Cancer Institute Dr. Georgs Fries (C) U.S. Department of Agriculture Mr, Albert Gardner (M) Food and Drug Adnlnlatratlon Dr. Larry Garthoff (M) Food and Dr-e Administration Ms. Helga Gerstner (C) Oak Ridge National Laboratory Dr. Joyce A. Goldstein (C) Environmental Protection Agency HONS 211842 v Ml. Bitty Hackliy (C) National Marina Fisheries Sarvlca Or. Charlas F. Jelinek (M) Food and Drug Administration Or. Louis Kaaza (M) Food and Drug Administration Or. Ranata Kimbrough (M) Cantar for Olsaasa Control Mr. Thomas E. Kopp (C> Envlronmsntal Protaction Agancy Or. Yuoh Ku (M) Food and Drug Administration Or. Richard L. Lahman (C) Oapartmant of Coaaarca Or. William L. Marcus (C) Envlronmsntal Protaction Agancy Or. H. B. Matthews (M) National Inatituta for Envlronmsntal Haaltb Sciancas Or. J. 0. McKinney (M) National Inatituta for Envlronmsntal Hsalth Sciancas Or. Josaph McLaughlin (C) Consumer Product Safety Collision Or. John A. Moora (M) National Inatituta for Envlronmsntal Health Sciancas Or. Irwin Pomsrants (M) Food and Drug Administration Dr. Richard A. Shodan (M) National Inatituta for Occupational Safety and Health Mr. John A. Roach (Hi Food sad Druf Administration Dr. Samual I. Shibko (M> ' Food and Drug Administration Dr. Raymond E. Shapiro (M) Food and Drug Administration HONS 211843 Dr. Richard H. Tasks (M) Food and Drug Administration Dr. William Trottar (M) Food and Drug Administration HONS 211844 vii SUBCOMMITTEE ON THE HEALTH EFFECTS OF POLYCHLORINATED BIPHENYLS AND POLYBBOMINATED BIPHENYLS Chairperson: A. C. Kolbye, Jr., M.D. Subgroups sad Thair Members 1. Chemistry J. Burka D. Firastons T. Kopp J. McKinney I. Poaarantz, Chairperson J. Roach W. Trotter 2. Mecabolisn and Biochemical Toxicity G. Frlaa A. Gardner L. Carthoff J. Goldstein Y. Ku H. Matthews, Chairperson J. Moore 3. Animal Tonicity and Careloosenasis J. Buckley L. Flshbela G. Fleas L. Lease E. Llabrougb, Chairperson W. Marcus S. Shibko R. Teaks 4. Kin Exposure F. Cordle, Chairperson P. Corneliuseen vi HONS 211845 C. Jellnek B. Hackley R. Lehman J. McLaughlin R. Rhoden R. Shapiro Staff Support H. Geretner J. Moore R. Shapiro HONS 211846 Subcommittee on the Health Effects of PCBs and PBBs Purposed 1. "Assemble, review and interpret data that assess the health significance of polychlorinated biphenyls." 2. "Formulate recosaendatlons as to future research needs." Specific Objectives; 1. Chemistry: Assess the Impurities thee may be found in PCBs and PBBs, the "hardness" of the data giving their potential for accumulating in the food chain. 2. Metabolism and Biochemical Toxicity: Assess the Metabolism of PCBs and PBBs and the possible contri butlon of specific Isomeric homologs to the observed toxicity; evaluate poeslble effects at the cellular level; and define existing dose-response relationships for the phenomena. 3. Animal Toxicity and Carcinogenicity Assess the short and long term effects. Including carcinogenis of exposurm to PCBs and PBBs In various animal models, their reletionship to known human effects, and the establishment of dose-response curves. HONS 211847 x 4. Human Exposure Quantity known human exposures, accidental pulsed, and Long range, their relationship to possible overt symptomatology, and Che establishment of possible dose-response curves. HONS 211848 xi INTSODL'CTIOM PCBs were reportedly first synthesized in 1801 by Schmid: and Schultz, but were not available commercially until 19301 (Hubbard, 1964). From 1930 until 1970, when their distribution in the U.S. was voluntarily restricted by Monsanto to closed systems, the use of PCBs in a wide variety of industrial applica tions had steadily increased. 1, Commercial PCBs are complex mixtures. Arochlor 1254, for example, contains 69 different molecules (Zitko, 1971), which differ in the number and position of chlorine atoms. This is a chemical representation of a PCB molecule. C carbon atom, X * Cl (chlorine atom) or H (hydrogen atom). The basic parent molecule contains two benzene rings connected by a carbon-carbon bond. When all the X* and X are hydrogen, the molecule is called biphenyl. When several of the hydrogen atoms ars replaced by chlorine atoms, the molecule is called a PCB. X and X* are used to indicate that the chlorine atoms are on different benzene rings. For example, the chemical structure for 2,5,2*,S'-tetrachlorobiphenyl is: e ci Positional Isomers have tho same number of chlorine arums, but they ai located at different positions on the ring. 2, 5,2',5'-tetrachlorobiphenyl and 3,4,5',4'-cetrachlorobiphenvL (TCB) are isomers. They are different molecules with differ ent chemical, physical, and biological properties. The chemical structure for 3,4,3',4'-TCB is: HONS 211849 The problem of polychlorinated biphenyls (PCBs) became of national concern in 1971, when several accidental contaminations of foods were reported. In addition, the extent of the environ mental contamination, and its persistence, were not precisely known. Subsequently, various regulatory actions were taken by the concerned agencies involved and. with the cooperation of the only C.S. producer, the situation was felt to be under control. Then, In 1975, the reported high levels of contamination, with PCBs, of Hudson River fish, refocused national attention on this environmental contaminant. It was soon apparent that the actions and control measures of the early 1970*s had not succeeded in totally reducing, or even substantially alleviating the problems associated with this environmental pollutant. On November 17-19, 1975, a National Conference on Polychlori nated Biphenyls, sponsored by the Environmental Protection Agency in cooperation with the Department of Agriculture, Council on Environmental Quality. Department of Health, Education and Welfare and Department of the Interior, was held in Chicago, Illinois. This conference brought together the latest data, and scientific expertise, to consider all aspects of the various problems asso ciated with PCBs, and the means for their possible rssolutlon. In his Introductory remarks to the Session on Health Effects a-d Human Exposure, Dr. David P. Rail, Session Chairman, announced the formation of a Subcommittee on PCBs of the DHEW Committee to HONS 211850 Coordinate Toxicology and Related Programs. The Subcommittee was given the charge to: 1. Assemble, review and interpret data chat assesses the health significance of polychlorinated biphenyls. 2. Formulate recomnendatlons as to future needs. The accidental contamination of animal feeds with polybrominated biphenyls (PBBs), and consequent human exposures, that occurrec in the State of Michigan, has been approached not only on its own basis, but in comparative relationship to the PCBs, as well. Therefore, the scope of this Subcommittee was expanded to the Subcommittee on the Health Effects of PCBs and PBBs. Members of this Subcommittee came from within the Oenartment of Health, Education and Welfare, as well as including consultants from other Federal Agencies concerned with this difficult problem. The report which follows represents the culmination of this Subcommittee's efforts. HONS 211851 GENERAL SUMMARY AND CONCLUSIONS CHEMISTRY The great complexity of PCB commercial mixtures has provi^r-d the difficult task of separating the components of these mixtures and determining the complete Identity of the compounds. Efforts to define these mixtures have been quite successful over the past several years (Hutzlnger et al., 1974). Identification of the component chemicals in PCB residues presents another difficult problem. Little progress has been reported in this area. Since the fate of components of comnerclal PCB mixtures entering the environment cannot be followed directly, inferences must be drawn about chemical alteration of chlorinated biphenyl compounds in the environment mainly based on laboratory studies. The known resistance of aryl chlorides to chemical oxidation and hydrolysis presumably was a major factor in the promotion of PCBs for industrial uses. Exposure to non-metabollc environmental agents Is unlikely to result In significant oxidation or hydrolysis of chlorinated biphenyls. However, since appreciable photoaltera tion of chlorlanted biphenyls has been obeerved using either sun light or sunlight-simulating lamps In the laboratory (Hutzlnger et al., 1974) as energy sources, it is highly probable that photo chemical changes occur In the environment. These photochemical studies have indicated that reductive dechlorination to biphenyls HONS 211852 of lover chlorine content Is a predominant alteration route although formation of more polar compounds, including a chlori nated dlbenzofuran, has also been reported (Hutzinger et al., 1974; Crosby and Mollanen, 1973). The diverse conditions of the environment make it difficult to predict accurately the end products and rates of photoalteration of chlorinated biphenyls. From a comparison of carbon-halogen bond-energies, bromlnated biphenyls might be expected to be more readily altered by light than chlorobiphenyls. But because of the lower volatility and different end uses of bromoblphenyls, they may not be as readily exposed to light as the chlorinated analogs. A typical PCB residue from fish resembles the Aroclor 1254 mixture more closely chan It does other Arodors (Zlcko et al., 1972; Velth, 1975). Considering the major components of Aroclor 1254 (Sissons and Welti, 1971) It appears chat penta-, hexa-, and heptacMorlnated biphenyls tend to concentrate at this trophic level In the biosphere. It is of considerable interest, therefore to carry out comparative accumulation, metabolism and toxicity studies e^haslzlng these higher chlorinated biphenyls using compounds of known chlorine substitution pattern. Individual chlorinated biphenyls are now quite accessible through synthesis and racent studies have begun to obtain these types of biological data. Studies using a group of five symmetrical hexachlorobiphenyl Isomers In chicks (McKinney et al., In press) and in mice (Biocca HONS 211853 1975) Indicated that distinct difference in coxicity are possible and these differences could be related to structure. The isomers with chlorine substituents in the 4,4' positions appeared to accumulate more rapidly in adipose tissue, showed increased acti vity in the liver and greater overall toxicity (McKinney, 1975; McKinney et al., in press). Comparison of two pentachlorbiphenyls in laying chicks again indicated the one isomer with 4,4'-substitution to have greater toxic effects (Ax and Hansen, 1975; Ax et al., 1976). Structure-activity relationships, particularly for biphenyls known to be major components of commercial FOB mixtures, could be useful in assessing the potential hazard of these compounds. A major pert of determining human exposure to FOB residues in foods is evaluating the adequacy of the analytical methodology used and the significance of the date obtained. The methods applicable to FCB determination are complex and to be judged adequate should give acceptably reproducible results in the hands of experienced analysts. The degree of success of inter-laboratory collaborative studies helps measure the adequacy of a method. PCB residues are multicomponent mixtures. Many consnon chlori nated pesticides ere extracted from samples along with the FCBs. Procedures for separation of FCB residues from interfering pesti cides therfore becOne important as a prerequisite to quantitation. Several different quantitation techniques have been used (Hutzinger et al., 1974). Comparison of FCB residue date is difficult where standardization of the quantitation procedure is lacking. HONS 211854 It his been shown that recovery of PC8 mixtures purposely added to food simples, using commonly applied analytical methodo logy, varies with the average level of chlorination. Recoveries tend to decrease with increasing chlorine level (Stalling et al., 1972). Analytical procedures for determination of PCB and PBS (the latter is predominantly a hexabromobiphenyl) residues are essen tially similar. They differ somewhat In cleanup of the extract and In the use of a higher column temperature for gas chromato graphic determination of the PBB residue. The limit of quantitation for PCBs Is generally 0.2 ppm for Individual foods and about 0.0S ppm for Total Diet compositee using Food and Drug Administration methodology. For PBB residues, the limit of quantitation Is approximately 0.03 ppm In fats and 0.01 ppm in non-fatty foods. Interlaboratory studies of analytical methods for PBB determination have not been reported. The chlorinated dibenzofurana (Cl-DBFs) have become a focus of concern as the class of contaminant compounds In comerclal PCBs most likely to contribute significantly to the toxicity of the PCB mixture. This concern presumably stems mainly from the demonstrated toxicity of 2,3,7,8-tetrachlorodlbenzo-p-dioxln and the simile .ty In structure between dibenzo-p-dloxln and dibenzofuran (NICHS Conference, 1973; also aee Appendix of Chemistry Report) and from the little toxicity data chat has been published for Cl-DBFs (Bauer et al., 1961). Recent daca indicated HONS 211855 Che Coxicicy of 2,3,7,8-Cetrachlorodibenzofuran approaches ehac of che analogous dibenzo-p-dioxin ac lease in chicks and in guinea pigs (Moore ec al.. 1926). If che chlorinaced dibenzo-p-dioxins can serve as an example, chlorinaced dibenzofuran coxicicy can be expected co vary wich number and posicion of chlorine aeoms on Che parent ring syscert. Relationships between structure and Coxicicy will emerge as more Cl-DBFs are synchesized and tested for toxicity. A range of Cl-DBFs (from dichloro chrough hexachloro) have been reported in various Aroclors (Roach and Pomerantz, 1974b; Bowes at al., 1975a) and two specific compounds, the 2,3,7,3tetrachloro- and 2,3,4,7,8-pentachloro-dlbenzofurans, have been identified (Bowee et al., 1975b). Quantitation of Cl-DBF contami nants in PCB mixtures, a difficult procedure, suggests total Cl-DBF levels in the low parts per million range (Nagayaoa et al., 1975; Bowes et al., 1975a). The presence of Cl-DBF in a synthesized, individual, symmetrical chlorinated biphenyl has been reported (Moron et al., 1973). Chemical analysis of these individual chloroblphenyls may be neceseary prior to their use in toxicologies and other biological studiee. There have been no published positive findings of Cl-DBFs in environmental st plea or in foode other than rice oil. The finding of Cl-DBFe in such samples would not necessarily implicate conmerclal PCBe as the source of these contaminants since Cl-DBFs (generally the higher chlorine laVfls) have been reported in other HONS 211856 industrial ehemicale (Firestone et al., 1972; Schwetz et al., 1974; Villanueva at al., 1974). It is alao possible that Cl-DBFs nay be formed or altered in the environment. METABOLISM AMD BIOCHEMICAL TOXICITY The effect of PCBe and PBB on the hepatic mixed-function oxidase (MFO) enzymes has been the moet thoroughly studied of any biochemical parameter chat they are known to alter. On a molar basis, the PBBe are approximately five-fold more potent than the PCBs in inducing increased levele of the MFO enzymes (Farber and Baker, 1974). Among the PCBs it appears as if their potency increases with increasing chlorination and chloriae sub stitution in the pare>ortho> meta positions respectively (Echobichon, 1975). However, this induction of the MFO enzymes is not unique to these compounds and the induction jossrved is well within the range observed with many other xenobiotics. The PCBa are somewhat unique as MFO inducers la that they Induce the formation of both Type I and Type II P-450 (Alvares et al., 1973), but this induction may have been due to the fact that the comerclal formulation used in the study was a mixture of twenty or more PCBs which were metabolized to an even greater number of metabolites. In addition, many, if not most comercial PCB formulation contain trace amounte of chlorinated dlbenzofurans (Araki, 1974). These compounds may be up to 170 times more potent as MFO inducers than the PCBs. The chlorinated dlbenzofuran concentration of Amarclcan 9 HONS 211857 PCB formulation* i* usually quite low when they are produced, how ever, the effect of long-term is unknown. In any case, since induction of the MFO enzymes may result in increased hormone metabolism or carcinogea activation, exposure to the PCBs and PB3s should be limited on that basis alona. PCBs and PBBe administered at relatively high concentrations, usually 50 ppm or higher in the diet, to laboratory animals have bean shown to cause prophyrla (Goldstein at al., 1975), disfunction of the thyroid (Bastomalcy, 1974), inhibition of various enzymes (Pardlni, 1971), changes in the liver to body weight ratios (Garthoff at al., 1975) various disorders of the liver (Kimbrough at al., 1975), and to altar the level or utilization of corticosteroids (Vasserman et al., 1973) and vitamins A, D and E (Cecil at al., 1973; Vong at al., 1974; Combs et al., 1975). Certain of tha less chlorinated PCBs have also been shown to hava a mild astroganlc effect when tested in the immature rat and mouse (Kihlstrom, 1973; Nelson, 1974). However, moat of these parameters have not been studied in sensitive species or demonstrated at low exposura levels in laboratory animals. The available data imply that the PCBs and PBBs containing aim or fewer halogen atone are readily absorbed from the gut of higher animals. Ths available data also imply that the PCBs are not excreted to an appraciable axcent prior to metabolism to mors polar compounds, and that long tarm PCB storaga is In the skia and a'dipose tissue (Matthews and Anderson, 1975a). Studies of PBB 10 HONS 211858 metabolism ars not yet available. Since tissue samples from birds and navels with known exposures to PCBs usually contain only those PCBs with five or more chlorine atoms, it is assumed that the less chlorinated PCBs have bean metabolized and excreted. On the other hand there is little evidence that fish can metabolize any PCB and an analysis of fish tissue usually shows a PCB pattern vary similar to that to which the fish were exposed (Stalling and Mayer, 1972). Laboratory studies have demonstrated that the rats of PCB metabolism and thus excretion is approximately inversely propor tional to the degree of PCB chlorination so long as there are two adjacent unsubstituted carbon atone on the biphenyl molecule. When two adjacent unsubstituted carbon atoms are not present the biological half-life of the given PCB may be a matter of years and accumulation of high tissue concentrations with continued exposure is inevitable (Matthews and Anderson, 1975a). It should be noted that the msjor constituent of Firemaster BP-6 does not have two edjacent unsubstltutad carbon atone and that the corre sponding PCB has been shown to have an extremely long half-life in the laboratory rat and probably the human population as well. It should also be noted that metabolism can also result in further coaplieations of the PCB problem because those PCBs which are most readily metabolized and excreted ars those which are most likely-to be metabolized via arena oxide intermediates. And where as the reactivity of arena oxides varies greatly and often HONS 211859 ultimately determines the toxicity, mutagenicity or carcinogenicity of the parent compound (Jerlna and Daly, 1974), the PCBe offer such a range of degree and position of substitution that it would not be unlikely to find that one or more of these PCB metabolites would have the proper stability to be a mutagen or a carcinogen. Thus a move from the more highly chlorinated PCB formulations to the leas chlorinated ones may help solve the long-term residue problem only to intensify other problems. ANIMAL TOXICOLOGY Gleaning the information that is now available on animal toxicology makes it obvious that different coMerclal mixtures of PCBe elecit different toxic responses in animals, and that different animal species vary in their susceptibility to the toxic effects of PCBs. Reproduction is severely affected in mink at a dietary level of 5 ppm Aroclor 1254, and a slight effect is still noted at a dietary level of 1 ppm (Ringer at al., 1972). In rhesus monkeys reproduction was reduced at a dietary level of 2.5 ppm of Aroclor 124B (Allen 1975). In rats a dietary level of 20 ppm Aroclor 1254 depressed reproduction while in the same rat strain (Sherman), in a study conducted simultaneously in the same laboratory, a dietary level of 500 ppm Aroclor 1260 was necessary to reduce reproduction (Linder at al., 1974). In comparative studies done with European products, Phenoclor DB6 and Clophen A60, and the American product, Aroclor 1260, the HONS 211860 European products were more toxic to chickens than the American product (Voa & Koeman, 1970). The difference in toxicity in this study was attributed to contamination of the European products with chlorinated dibanzofurans and perhapa chlorinated naphthalenes. However, if the differences in the effect on reproduction by the different Aroclors are coopered, the contamination with chlorinated dlbenzofurans nay not be the decisive factor. Hepatic porphyria has been produced in a numbar of species, namely the chicken, rabbit, Japanese quail and rat, by a number of commercial mixtures, such as Clophen A60, Phenoclor 0P6, Aroclor 1016, 1242, 1254, and 1260 (Vos and Koeaan, 1970, Vos and Sseas, 1971, Iverson at el., 1975, Goldstein at el., 1974, 1975). It (hspetic porphyria) has not been reported in the monkey, mink or human (see Yusho) mammalian species quite susceptible to the toxic offsets of PCSa in other ways. Aroclor 1254 end Aroclor 1242 produced hepatic porphyria in the female ret at doses lover then Aroclor 1016. Again, different coserciel mixtures produce this toxic effect at different doaage levels. The hepatic porphyria occurs concomitantly with an incraeee in ALA synthetase in the liver. Mixed function oxidases era also Induced in the liver and comparative studies with PCB isomers have suggestsd that if the 4,4' positions on the biphenyl ring ere occupied by chlorine atoms the effect la most pronounced (Ecoblchon and Comeau, 1975). The liver is the primary target organ for PCBa in the rat. Early chengea Include hepatomegaly with a concomitant increase 13 HONS 211861 In smooth endoplasmic retiulum, lipid accumulation at higher dietary levels (20 ppm for Aroclor 1254 and 1260) and ultrastructural changes such as atypical mitochondria and the formation of "fingerprints" in the hepatic cytoplasm ("Fingerprints" consist of concentrically arranged membranes surrounding lipid vacuoles). An increase in mitotic figures and cell breakdown are increasingly noted with either higher dosas or longer exposure (Kimbrough, 1975). These changes are not ae pronounced with Aroclor 1242 or 1016. Similar changes have been reported in the primate (Allen, 1975). In the mouse, liver tumors have been produced with Aroclor 1254 (Kimbrough and Linder, 1974) and Kanechlor 500 (Ito at al., 1973) and in the rat with Kanechlor 400 (Klmura and Baba, 1973) and Aroclor 1260. Some tumors have also been produced with Aroclor 1242, 1254 and 1260 (Calandra 1976) in rata in a separate study. In one study with Aroclor 1260, some of the tumors were classified as hepatocellular carcinomas (Kimbrough at al., 1975). The dietary levels of the PCBe in all of these studies were 100 ppm or more, a high level when compared to the daily average human intake of PCBs but not very high for subgroups with a high Intake of fish from polluted waterways or in soma occupational situations. This is emphasised by the fact that the dietary levels which produce tumors and the relatively low dietary levels which affect reproduction (2.5 ppm in the monkey and 1 ppm in the mink) of some coMerclal PCB mixtures do not represent a no effect level. It is presently not known what the no effect levels are. Other uncertainties 14 MONS 211862 contribute to these problems. The Yusho oil which caused s poison ing outbreak in Japan wss not only contaminated with PCSs but also with high levels of chlorinated dibenzofurans. In the primate, in addition to the effect on the liver, the gastric mucosa (Allen and Borback, 1973), the skin, and the Meibo mian glands are also affected at comparatively low dietary levels and the bone marrow is depressed (Allen 1975) while the gastric mucosa of the rat is only affected at exceedingly high doses (Kimbrough, 1974). Whether the dog also shows an sffect on the gastric mucosa needs clarification. In the rabbit, atrophy of the thymus is a toxic manifestation in addition to liver pathoLogy (Vos and Beema, 1971). Fluid accumulation occurs in primates, chickens, and finches. The lymphatic system is also affected in minks. The few studies conducted with some hexachloroblphenyl isomers demonstrated that the 3,4,5,3',4*,5* hexachloroblphenyl was the most toxic while 2,3,6,2',3',6' hexachloroblphenyl was the least toxic isomer (Blocca at al., 1975), Penta-, hexa- and heptachloroblphenyls are preferentially retained in mamellan adipose tissue for extremely long periods of time (Kimbrough, 1975) (at fairly high concentrations in the ret for a recovery period of 16 months). Whether this has an influence on the toxicity of PCB mixtures has not been determined. The toxic effects of the contaminants of PCBs have not been extensively studied. While it is assumed that chlorinated 15 HONS 211863 naphthalene* are toxic within the same dosage range as the chcr:nated biphenyls, the chlorinated dlbenzofurans probably have a greater toxicity. It is assumed that 2,3,7,8 tetrachlorodibenzoCuran Is the most toxic of this group of compounds. This compound also shows marked species variation. While the single oral LD50 in guinea pigs is between 5 and 10 ug/kg bodywaight, 1000 ug/kg TCDF given orally had no affect on rats. Mice are equally insen sitive to the toxic effects of TCDF (Moore et al., 1976). For the bromlnated biphenyls limited toxicity data are only available on mixtures containing predominately hexa- and oetabromoblphenyl. Both mixtures differ sufficiently in isomeric composition so that their toxic effects may quantitatively be quite different and also different from the PCBs. Again tha problem of toxic contaminants has not been resolved. If toxic effects are similar to PCBs then, at least in some species such as mink and monkey, longterm low level exposure should result in meaaureable toxicity. Additional animal studies are needed to resolve some of these problame. Poor metabolism and excretion of the bromlnated biphenyls may lead to long retention of these compounds predominantly in adipose tissue with accumulation to very high levels on continued exposure. Whether this would lead to sufficient recirculation of the chemicals to cause toxic effects on target organs in presently not known. HONS 211864 16 HUMAN EXPOSURE Several reports (Jellnek & Corneliussen, 1975; Humphrey, 1976; Kutz and Yang, 1975; Dennis, 1975; Kleinhert, 1975; Hesse, 1975; Yobs, 1972; Kutz and Strassmman, 1975) provide evidence chat would indicate that a substantial proportion of the population of the United States has been exposed to PCB's, Minimal human exposure of the population Co PCB's has occurred from food, air and water, while significant human exposure appears to be limited to sports fishermen consuming fresh water fish from contaminated streams and lakes, and to occupational exposure in Industrial workers. Jellnek & Corneliussen (1975), in reviewing data from the FDA Total Diet Study (1971-1975) report that all food classes of the total diet have declined to no PCB occurrences except In these meac-flsh-poultry composites. About 40 percent of these composites continue to contain detectable residues of PCB's, although only traces have been detected In the latter years. The fact that levels in these composites have declined to only traces further support the Inference that the meat, poultry and eggs no longer contain detectable PCB's and that the low level findings are probably due to the fish In these co^oaltes. This would Imply that the PCB levels In the diet may have "bottomed out" ' and may remain static until such time as there Is a change in the PCB residues in fish, HONS 211865 Data compiled from studies sponsored by the National Marine Fisheries Service (NMFS) also support the continuing presence of PCB residues In fish. A compilation of PCS data, representing the results of all the measurements known to NMFS on PCB'a in fish used in the U.S. diet, indicates several important points: (1) while at one time or another, some PCB measurements have been made on many fish, there is an inadequacy of information on PCB residues in the most liqiortant fish items in the fish diet; (2) sampling and analysis have been sporadic and not designed to measure trends in human exposure; and (3) systematic surveys of neither the l^tortant items nor the species moat liksly to be contaminated have been undertaken. However, these survey data do show that in general U.S. fish eaters include a wide variety of fish items in their diet and that some 93 percent of the U.S. population (197 million) consume fish, with the average annual consumption of fish being 15 pounds/year/fieh eater. At present, it is difficult to estimate all human exposure to PCB from eating fish, either from the population as a whole or subgroups at higher risk of consuming large quantities of fish with higher PCB residues. Fragmentary evidence from NMFS data suggests that the exposure of the population as a whole from PCB residues in ingested fish is probably well below 19 meg/day/ consumer, based on PCB levels which are estimated to be below 1 ppm and 19 g of fish consumsd/day. This can b coaparsd to HONS 211866 the reeults of the FDA Total Diet Study where it has been estimated thet the overall PCS daily intake is on the order of 5-10 meg/day for the general population. The lower FDA estimate Is based on the methodology of the Market Basket Survey where fish are purchaeed at the consumer level and would not be applicable to diets which include a high consumption of fish from certain areas with high PCB residues. A recently completed study (Humphrey et el., 1976) has attested to assess some of the consequences of human exposure to PCBs from the high consumption of fish from contaminated areas. The results of this study show thet a group of sports fishermen consumed an average of 24-25 pounds of fish/person/yeer, with the highest individual exposure for a two year period reported as 190 pounds per year. PCB residues in cooked fish ranged from 0.36 to 5.39 ppm. Although there was a wide range of blood PCB levels for each quantity of fish consiaed, there was a highly significant correlation between the reported quantity of Lake Michigan fish consumed and the concentration of PCB in the blood of study perticipents, with the higher reported fish consumption being associated with higher PCB blood levels. The blood values ranged from a low of 0.007 ppm in the control group (fish consump tion less than 6 pounds/year) to a high of 0.366 ppm in the exposed group (fish consumption 24-25 pounds/year). These investigators calculated that the amount of PCB ingested by the exposed group could average 46.5 mg/year and ranged from HONS 211867 14.17 to 114.31 mg/year. While no systematic adverse health effects could be demonstrated in the exposed group when compared to controls, the investigators caution that any long-term chronic effects are unknown at the present time. Additionally, it can be concluded that exposure similar to those reported in this special group will continue and there is the likelihood that as sportsflshing becomes more popular, larger numbers of people may be exposed la a similar way. Although human exposure to PCBa from air and water is probably minimal, there seems little question thee such exposure does occur. Samples of ambient air collected in Florida, Missi ssippi and Colorado, show that PCBa were present at all locations. The average concentration at each of the three locations was approximately 100 nanograme per cubic meter of air. Studies of surface water from the major drainage basins of the United States report the widespread occurrence of PCBs in both surface water and bottom sediments. Mean residue levels of PCBs in the surface water ranged from 0.01 to 0.05 mcg/llter, with a maximum residue level of 20.0 meg/liter. In Wisconsin, effluents from cooling water in aluminum foundries contained PCBa ranging from 11.5 to 335 ppb. Effluents from paper mills ranged from 0.01 to 25 ppb, Analysis of snow malt water from Wisconsin cities showed PCB residue levels of 0,17 to 0.24 ppb, suggesting that fallout of PCBa from the air may be an important source of PCBa entering the waters of the state. 20 HONS 211868 In Michigan, testing of 900 samples of industrial effluents shoved 22 percent had PC8 residues greater than 0.5 mcg/liter, 8 percent greater than 1.0 mcg/liter, 6 percent greater than 10 meg/ liter and 2 percent greater than 100 mcg/liter. With sludge dis posal taking place by incineration, landfill and crop or pasture application, the continuation of PCBs in the environment seems obvious. Adverse human health effects resulting from PCS exposure have coma primarily from studies of occupational exposure and from human exposure through the ingestion of contaminated rice oil in Japan. Schwartz (1936) provided some of the earliest reports of adverse health effects due to occupational exposure in the U.S., in which he described skin lesions and symptoms of systematic poisoning among workers who were reported to have inhaled ehlorodiphenyls. There have been numerous reports over the ensuing years describing cutaneous eruptions and of systematic manifestations as well, among marine electricians, machinists, capacitor and transformer manufacturing workers, and others occupationally exposed to PCBe. The skin lesions described by Schwartz in 1936 have come to be designated as ''chloracne." Part of the chloracne lesion resembles adolescent acne, but is generally more severe and the lesion distribution is inconsistent with adolescent acne. Hara (1969) and Kasegawa at al., (1973) have reported dermato logic ailments which Include "brown chromodermatosls'' of the 21 HONS 211869 dorsal joints of the hands and purple-like eruptions of the face and neck. However, Hasagawa et al., (1973) performed a health survey of workers in carbonless copy paper factories, two years after the use of PCS in such processes had ceased, and reported no dermal effects nor liver function, urine or blood test abnor malities. PCB blood levels ware reported as 0.01 - 0.02 ppm. Typical clinical findings in the human exposure to PCB which occurad in Japan in 1966, and which resulted from the ingestion of rice oil contaminated with Kaaachlor 400 included chloracne and increased pigmentation of the skin, increased eye discharge, transient visual disturbances, feeling of weakness, numbneaa in limba and soma disturbance in liver function. Adult Yuaho patienta had protracted clinical diseaae with a slow regression of symptoms and signs. In the dose-reaponae epidemiologic study, the average cumulative intake of PCB'a leading to overt symptoms was 2,000 mg, with the lowest dose leading to overt symptoms being 300 mg. However, Kurataune et al., (1975) have introduced a new factor into the Yuaho incidence with the finding that the rice oil contained chlorinated dlbanzofurans (Cl-DBF) at 3 ppm. Nagayama et al., (1975) reports that the toxicity of (Cl-DBF) is said to be from 200 eo 500 times that of PCB. Whether or not the (Cl-DBF) con taminant la the crucial toxic substance producing the symptoms observed in the Yuaho incident, or whether the exposure to the high levels of Kanechlor 400 produced the observed effects, or 22 HONS 211870 whether there wee en inter-active process taking place is unknown. The date necessary for determining reasonably accurate time and dose exposure to PBS in individuals In Michigan is either unavailable or non-existent. Attempts to secure accurate dietary intake with the PBS levels in food and the duration of consump tion have been unsuccessful. It Is hoped that data to be received from the Michigan Department of Public Health may provide some basis for crude estimates. While there appears to be no evidence at the moment to indi cate acuta health effects from exposure to PBB, any chronic effects remain largely unknown. A large scale epidemiological study is expected to get underway shortly to Identify all the farm family members from quarantined farms; a large group of study subjects secondarily exposed to PBB through the purchase of farm products on a regular basis from quarantined farms and a control group of individuals not exposed to PBB contamination. This study will continua efforts to Identify any acute or chronic effects of PBB exposure through physical examination, biochemistry tests, and dietary histories. Efforts will continue to assess the original Fireaaster BP-6 for the presence, If any, of chemical contaminants which might present human health hasards. In the meantime, a ten fold safety factor for PBBs when cohered to PCBa appears both reasonable and acceptable based on all currently available scientific data. 23 HONS 211871 Much work remains to be done concerning the toxicity of PCBe and PBBe and che association of these compounds with any demon strable adverse human health effects. To accomplish some of these needs a list of recommendations follows. HONS 211872 GENERAL RECOMMENDATIONS Mora collets and additional metabolism studies with various individual PCBs and PBBs should be undertaken. Attention to the formation of possible toxic metabolites of these compounds should be considered, as well aa detecting and quantifying any specific biochemical parameters which may be affected as a result of such exposures. Based on the above, lifetime feeding studies should be conducted with selected commercial PCBs and PBBs mixtures, as well as selected individual PCBa and PBBs. Why some species seem to be more susceptible to the toxic effects of PCBs, and which are most closely related to man in their response should be determined. In addition, more experi mental data ia needed for the occupational PCBa settlnga; i.e., via the respiratory or dermal route. Additional toxicological evaluatlona should be made of fish and other foods which contain high levels of environmentally accumulated PCBa and other con taminants. Appropriate toxicological studies should be undertaken with PBBs in order to be able to make some predictions on possible human effects. Tha quantitative evaluation of halogenated biphenyl exposure to men with respect to blood and body fat lavela, and any possible health-effects, needs further study, With respect to PCBa, this may be accomplished by identifying a population of individuals consuming large amounts of fish contaminated with high levels of HONS 211673 ?CBs. In addition, those exposed Industrially should be Included In these efforts. With respect to PBBs, appropriate studies should be Initiated with Michigan farm families exposed through consump tion of contaminated meat and dairy products. Individual chemical components of PCB residues should be qualitatively identified and procedures for improving quantitation of these residues should be investigated. Commercial ?CB (and PB8) mixtures need to be analysed further to determine which chlorinated dlbenzofurans (or, in the case of PCBs, whether any brominated dibenzofurans) are present as contaminants. Specific chlorinated dibeuzofursn compounds must be synthesized for use in development of analytical procedures and to aid in identification of contami nants. Analytical procedures are needed to permit examination of foods for presence of chlorinated dlbenzofurans. Toxicological studies should be carried out on chlorinated dlbenzofurans and brominated dlbenzofurans. HONS 211874 DETAILED RECOMMENDATIONS CHEMISTRY Investigate procedures (such as Webb and McCall's, J. Chromatog. Sci. 11, 366--373 (1973)) for possible improvement of the qusntltstlon step In the analysis of PCB residues. Present procedures for analysis of PCBe yield a "PCB residue" that is cononly examined using electron capture GC. Individual components of this residue should be identified and it should be determined whether any of the electron capture GC peaks can be attributed to Cl-DBPs or Cl-naphthalenes. Analyze commercial PCB mixture (e.g. Aroclors) to determine which Cl-DBPs are present as contaminants. Qualitative and where possible, quantitative studies should be carried out. Synthesized, individual chloroblphenyls should be examined for presence of chlorinated dibenzofurans before metsbolisn or toxicity studies sre csrried out with the biphenyl. (The importance of this is shown by the reported formation of 2,3,7, 8-CI4 DBF as a side-product in the Ullnann coupling reaction used to synthesize the sywetricsl 2,2'(4,4',5t5'-hexachlorobiphenyl.) Synthesize, purify and characterize by physical and chemical means a range of Cl-DBPs to be used for analytical procedure development se reference standards for confirmation purposes and for suitable toxicological studies. HONS 211875 Develop a suitable procedure for analysis of edible fish for Cl-DBFs, emphasising recovery and quantitation of 2,3,7,8-tetrachlorodibenzo furan. Analyze a commercial mixture of brominsted biphenyls (Flrenaster BP-6) for bromlnated dibenzofuran content. Analyze long-used heat exchange, transformer and capicltor fluids for chlorinated dibenzofurans. Synthesize specific 1*C or tritium radiolabeled chlorinated biphenyls that may be needed for toxicity and metabolism studies. Study the reported conversion of certain chlorlaatad biphenyls to Cl-DBFs under sialight or sialight-elaulating conditions. If feasible, extend the work to additional Cl-bipheayls. Simulating conditions applicable to heat exchange units and/or transformers, determine whether Cl-DBF content of FCB mixtures increases when the PCBs are heated and/or axpoeed to air. Carry out laboratory studies, similatlng environmental conditions, to determine alteration products from Cl-DBFs when irradiated or heated. MTOBOLISH AW) BIOCHEMICAL TOXICITY Long-term feeding studies spproschlng life-time should be conducted with the less chlorinated FCB formulations. These 28 HONS 211876 studies should include at Isaac two of cha laaa chlorinated PCB formulations and a series of carefully selected individual PCBa. Further and sore complate metabolism studios should be done with the more highly chlorinated PCBs, for comparison with those done with PCBs having five or leas chlorine atoms. The effect of chlorine position on PCB metabolism and arena oxide formation should be further elucidated. The formation of specific chlorinated dlbenzofurans as possible PCB metabolites should be studled, their biochemical toxicity in mammalian species determined and the magnitude of their toxicity ascertained. These studies will also include those specific chlorinated dlbentofurane identified in new and used PCBa. If possible, arena oxide metabolites of several PCBs should be synthesized and their toxicology investigated. Wichln an isomeric series of PCBa, (e.g., tetrs or hexa), metabolism of the more toxic members should be compared to those which are lasa toxic. of individual PBBa, comparable to those for PCBs, ffriPil't be studied. These data should be related to known toxicltles. HONS 211677 The toxicology of known FCB metabolites should be sCudlad. Porphyrin axcration by tha paopla and animals axpoaad in tha Michigan PBB incldant should ba chackad. Paopla who ara suapactad to hava racalvad high exposures of PCBa or PBBs should be chackad for any changes in enzyme levels-- e.g., protein-bound iodine, antipyrene metabolism, serum cholesterol, etc. Tha adsorption, distribution and excretion studies of halogensted dibenzofurans and PBBa should bn studied. Further studies of possible PCB metabolism by fish. Study the degree of chlorination and length of exposure on the estrogenic effects and reproductive organs with particular emphasis on the lower chlorinated PCBa (e.g., 1016). ANIMAL TOXICOLOGY Inhalation studies with "heated" PCB mixtures simulating occupational exposure and dermal toxicity studlaa should be conducted. Studies as to why some species seem to be more susceptible to the toxic effects of PCBs than others and of these species, which are most closely related in their response to humans should be undertaken. 30 HONS 211876 Subacute and chronic toxicity studies should be conducted with hexabromoblphenyl (Firemaster FF-l) In order to be able to make son* predictions on human PBB toxicity. The toxicity of fish containing high levels of PCBs which have been accumulated by environmental exposure should be evaluated. It should be established whether and which toxic impurities are present in ueed PCBe, such as transformer oil and capacitor fluid. Long-term feeding studies, approaching life-time, should be conducted with laboratory animals and selscted commercial mixtures of PCBs and PBBa as well as selected individual PCBs and PBBs. Further and more complete metabolism studies should be done with individual PCBs and PBBa. The formation of toxic metabolites of these compounds should be considered end carefully Investigated. Efforts to detect and quantitate specific biochemical mechanisms which are affected or altered by exposure to PCBa or PBBs should be continued. HUMAM EXPOSPIB Polychlorinated biphenyls Recommend that a population of fish eatars be identified who are consuming substantial amounts of frssh watar fish with high levels of PCBs. This study population should bs followsd prospectivsly with adequate diatary historiss, blood and body fat levels HONS 211879 >, of FCBs, a health questionnaire, liver function tests and other biochemical studies. Recommend a similar study be carried out in an industrial population exposed to FCBs. Systematic studies to identify sub-groups of the total U.S. population who consume high levels of fish with more accurate measures of PCB exposure. Polybrominated biphenyls Recommend that a large scale epidemiological study be carried out to (1) identify all farm family members from quarantined farms in Michigan, (2) identify those individuals with secondary exposure to PBB contamination through the purchase of dairy products from quarantined farms on a regular basis, and (3) a suitable control group of non-exposed farm famillea. These groups should be followed to assess any short and long term adverse health effects related to PBB exposure. Subgroups of these farm families from quarantined farms should be studied by: 1. Repeat FBB blood levels. 2. Matched blood-body fat PSB levels. 3. Pregnant female-infant studies. . - 4. A battery of biocnemical studies. HONS 211680 Ancillary Studies Toxicology studies In non-human primates and rodents with hexabroainated biphenyl and any brominated contaminants of hexabrominated biphenyls. Toxicology studies In several animal species (mice, rats, dogs, non-human primates using hexabroainated biphenyl and several PCBs to determine species differences in response to the effects of these compounds. Toxicology studies feeding hexabroainated biphenyls and selected PCBs singly and in combination to determine If there are different or additive effects when fed in combination. HONS 211881 CHEMISTRY OF PCBs AND PBBs I. Introduction. A recent book by Huttinger et al. (1974) reviews the chemis try of chlorinated biphenyls through the end of 1973. This wide-ranging review also includes discussions of metabolism and determination of chlorinated biphenyls. The present report reviews areas of chloroblphenyl and bromobiphenyl chemistry held to be pertinent to the Subcommittee' concern for possible health effects of these types of compounds. No attempt at complete coverage of these subject areas is claimed. Findings reported since 1973 are particularly stressed. Non-metabolic alteration of the halogenated biphenyls is considered in terms of agents most available to Induce chemical change in the environment. Expected differences between chlorinated biphenyls and brominated biphenyls in their environ mental chemistry are commented on. It is possible that certain of the chlorinated biphenyl compounds will be shown to have relatively greater potential hazard for health. Therefore, occurrence and fate in the environment and me^sbolism and toxic effects in animals are related to chemical structure of the biphenyls. For the general public, food Is presumed to be the major HONS 211882 source of chlorinated biphenyl compound*. The important concepts related to analytical methodology for PCB (and P3B) residues, particularly in foods, are reviewed. Alternative means of quantitation of these residues and problems in the quantitation techniques are discussed. The pertinent aspects of chlorinated dlben2ofurans, as known contaminant* in commercial PCBa and as potentially significant environmental contaminants, are reviewed. In reviewing the chemistry related to health effects of the PCBa and PBBs, it must be remembered that environmental contami nation by these two coMarcisl chemical mixtures la vastly different in magnitude. The former have been steadily released into the environment, in many countries, presumably ovar decades, and are now found to be a pervasive, world-wide contaminant. The number of chlorinated biphenyls reaching the environment probably number nearly 100 different compounds. The PBBs, encompassing a small number of chemical structures to begin with, are of concern due to a single, fairly recent contamination incident, apparently limited to the State of Michigan. tl. Chemistry of Chlorinated Blohenvla A. Synthesis and analysis of Commercial Mixtures . Commercial preparation of mixtures of chlorinated biphenyls by.reaction of biphenyl with chlorine has been described by Hubbard (1964). A more recent discussion of this subject, HONS 211883 including the newer preparation designated as Aroclor 1016, is now available (Mleure et a!., 1976). Analysis of the chlorinated biphenyl content of various American commercial PCB mixtures (Aroclor) has been reviewed by Hutzinger et al. (1974). An estimated 40 to 60 different chlorinated biphenyl compounds are present in each of the higher chlorinated commercial mixture. (There are 209 possible compounds obtainable by substituting chlorine for hydrogen on from one to tan different positions on the biphenyl ring system; see Appendix.) PCB commercial mixtures produced In the U.S. and elsewhere have been shown to contain classes of compounds other than the chlorinated biphenyls: chlorinated napthalenes and chlorinated dlbenzofurans (Cl-DBFs), for example (Vos et al., 1970; Roach and Pomerantz, 1974 a,b; Bowes et al., 1975a). The possibility that naphthalene and dlbenzofuran contaminate the technical biphenyl feedstock used in preparation of the commercial PCB mixtures has not been excluded. Available toxicity information Indicates that, of the identified types of contaminants In commercial PCB mixtures, Cl-DBPs pose the greatest potential hazard (See Section II.E on Chlorinated Dlbenzofurans). Aa a result, the Cl-DBFs have became the focus of studies on contaminants In these mixturesMo report has appeared of an attempted complete content analysis of the trace l^iuritles in a commercial PCB mixture. HONS 211884 B. Hon-metabolic Alteration of Chlorinated Biphenyls Chlorinated biphenyls, as is typical of aryl chlorides generally, are quite stable to chemical alteration. Consideration of possible non-matabollc alteration routes for these compounds in the environment suggests air oxidation, aqueous hydrolysis, and photoalteration in sunlight as potential reactions to be investigated, (Thermal decomposition would be of concern in connection with incineration conditions meant to destroy waste chlorinated biphenyls). Study of these reactions under the complex and variable sets of conditions existing in the environ ment is difficult. Chemical reactivity of chlorinated biphenyls has therefore been studied almost entirely under the more con trolled conditions of the laboratory. Oxidation PCBe are fairly stable to oxidation under moderate conditions. PCBa are stable under conditions which essily oxidize DDE, and can be asperated from DDE by oxidizing DDE to the more poler dlchlorobenzophenone prior to colmn chromatographic sspsrstlon (Trotter, 1973). To effect the oxidation of DDE, the PCB-DDE solution Is refluxed is 2.5X chronic acid-acetic acid on a steam bath. Some cf the loa ,r chlorinated biphenyls, however, are not recovered after the oxidation. It Is reported (Uelngarten, 1961) that mono-, di-, and trichloroblpheayls are oxidized in 7.5X HONS 211885 37 chromic acid-acetic acid to chair respective (chloro-) benzoic acids. With vigorous oxidizing conditions in che environment, some PCB oxidation, especially of the lower chlorinated biphenyls may occur. It is difficult, however, to assess the total extent of possible environmental oxidation of PCBs. Hydrolysis and Alcoholysis PCBs are fairly stable to hydrolysis under moderate conditions. When refluxed with 21 KOH in ethanol, PCBs are stable (Trotter, 1973; Young and Burke, 1972), A halogen on a chloroblphanyl molecule is not easily displaced in a nucleophilic substitution reaction. Under vigorous conditions the 4 and the 4,4' positions of decachloroblphenyl are found to be the most susceptible to chlorine displacement by hydroxide and mathoxide ions. Decachlorobiphenyl can be hydrolysed to octachloro-4,4'-blphenylol when treated with aqueous alkali at high temperatures in an autoclave (Smith, 1943; Societe d'Electro Chiaie, 1962). Heating 2,3-dichlorobiphenyl with sodium mathoxide produces 2-chloro-5-bipbenylol (deCrauw, 1931). Decachloroblphenyl when treated with sodium mathoxide in pyridine yields the 4-methoxyand 4,4'-dimethoxy-chlorobiphenyl (Binns and Suschitzky, 1971). It la probable that the non-metabolic hydrolysis or alcoholysis of PCBs in the environman is limited. Photochemistry Exact environmental conditions for the photochemistry of 38 HONS 211886 environmental contaminants, including PCBa, are sometimes difficult to simulate in the laboratory. Limited understanding exists concerning the parallel of Laboratory photoreactions and reaction rates under non-envlronmental conditions and possible environmental photolyses. The exact environmental conditions under which residues may photoreact can be diverse and difficult to evaluate- Unknown or discounted factors In the environment, such as possible sensiters or quenchers, may be ioportant. The frequency (energy) and Intensity of a laboratory photolysis light may not be coi^srable with environmental conditions. 3000 A probably represents the practical lower limit of the ultra violet (UV) portion of sunlight (Crosby, 1969). The residue must be exposed to sufficient light to react appreciably. It Is necessary to consider the physical state of the residue exposed to light In the environment. The residue may exist as a solid or liquid (s.g., a film), a solution or a vspor. In photolyzlag solutions the solvent can have an li^ortant effect. Photoreactiona which proceed In one solvent may not proceed or may proceed at a different rate In another solvent. A muddy river containing a residue may yield different rssults cohered to e laboratory photolysis In hexane, Laboratory photolyses under a variety of conditions which simulate most. If not all, of the environmental conditions of photolysis could be extremely sign ificant and relevant. In the absence of these, we must rely on the questionable extrapolation of laboratory photolyses under non-envlronmental conditions. 39 HONS 211887 The photochemistry of PCBs has been described by Hutzinger, et al.,(1972; 1974). In contrast to oxidation and hydrolysis, PCBs are fairly easily photoreacted under certain laboratory conditions. PCB photochemistry has been studied in various solutions, as a thin film, and as a vapor, In hydrocarbon solvents progressive reductive dechlorination of the PCB is the predominant photochemi cal reaction (Hutzinger et el., 1974). Irradiation of solutions of chlorobiphenyls (Hutzinger et el., 1972) and trapped gas liquid chromatographic (GLC) effluents (Hannan, et al.,197]) have shorn that PCBs with higher chlorine content dechlorlnate more readily. 0.1Z hexane solutions of 2,2',5,5'-tetrachlorobiphenyl and 2,2',4,4*,5,5'-hexachlorobiphenyl were each irradiated with 3100 A light in a Rayonet reactor (Hutzinger et al., 1972). After 24 hours of Irradiation, 33Z of the 2,2',5,5*-tetrachloroblphenyl was unreactad while < 1Z of the 2,2*4,4',5,5'-hexachloroblphenyl remained. In dechlorlnating PCBs by photoexcitation, ortho chlorines preferentially cleave (Ruzo et al.t 1974). With PCBs containing only aeta and para chlorines, aeta chlorines are lost preferentially. The rate of dechlorination is faster in alcohol solvents, such as aethanol (Huatert and Korte, 1972) than in hydrocarbon solvents. Irradiation of 2,2*,4,4',5,5'-hexachloroblpheayl In aethanol yields, in addition to reductive dechlorination, ring aethoxylatlou (Ruzo et al., 1974). Photochemical studies qf PCBs In aqueous solutions may offer a significant relevance to environmental PCB photochemistry. A 0.4Z Aroclor 1254 suspension in water-dioxane (7+3) in the presence of sodium bicarbonate with 10 HONS 211888 air bubbling through the mixture wae irradiated with 3100 A light. Two thin layer chromatographic fractions of the Aroclor 1254 after irradiation corresponding to the "addition of water to chlorobiphenyIs" and a "carboxy fraction" were found (Hutzlnger et al,, 1972). Presumably PCBs formed by reductive dechlorination were also produced. Black light irradiation of Aroclor 1254 as a thin layer film in the presence of water yields a "carboxy fraction" and a fraction whose mass spectrum Indicates hydroxychloroblphenyls (Hutzlnger et al., 1972). Sunlight irradiation of certain chlorobiphenyls as a thin film without the presence of water gives reductive dechlorlaated products (Hutzlnger et al., 1974). Chlorinated terphenyls and quaterphenyls are also produced by the black light and sunlight irradiation of certain chloroblphenyls as a thin film (Hutzlnger et al., 1974). Vapor phase photolysis of PCBs may be extremely relevant especially for the more volatile components of Aroclors with low chlorine content. Sunlamp irrad iation of the vapor phase of a refluxing suspension of 2 2',5,5'tetrachloroblphenyl and water yields "carboxy" products (Hutzlnger et al., 1972). These laboratory photoreactions represent chemical conversions which may proceed in the environment. The photoreactloo rates under diverse environmental conditions and the extent of PCB degradation or reaction in the environment are extremely difficult to assess. u HONS 2U89 The reported low-yield conversion of certain chlorinated biphenyls to Cl-DBF is discussed under Chlorinated Dlbentofuran, Section II.E. C. Chemical Structure Related to Occurrence, Fate end Effects of Chlorinated Biphenyls The physical and chemical properties of chlorinated biphenyl compounds and casmercial mixtures of PCBs vary greatly depending on the degree and position of chlorine substitution on the biphenyl ring system. Of particular loportance to their environmental occurrence and fate are the properties of volatility, watar solubility, bioaccumulation, biodegradability and photostablllty. Volatility, water solubility and bioaccumulation are of more importance as mechanisms of Introduction Into and transport within the environment. Biodegradation and phoCodegradation are more important as mechanisms of removal. There is data (Mleure at al., 1975) which Indicates that the higher chlorinated biphenyls are lass volatlla and less water soluble than the lower chlorinated biphenyls. These two factors would tend to enhance the ratio of lower to higher chlorinated biphenyls in the environment. There is no clear data to assess the involvement of chlorine position in volatility and water solubility, but is llkaly to be of less importance. Therefore, the lower chlorinatsd biphenyls are more llkaly to volatilize into and be selectively transported In the aqueous environment. HONS 211890 Tha relationships between structure and bloacctimulatlon factors for PCB isomers are only beginning to be assessed (McKinney, 1975; Sugiura et al, 1975) with the availability of purified specific lsomars for study. However, it appears (Zltko et al., 1972; Veith, 1975) that tha peak patterns from gas chromatography found in environmental samples representing tha biosphere moat cloaaly resemble tha higher chlorinated com mercial mixture, Aroclor 1254. This suggests a selective accumulation of the morea highly chlorinated components of the commercial mixtures in biological material. This also Indicates that the higher chlorinated biphenyls are able to find their way into the environment in spite of their poorer volatility and water solubility. Pharmacokinetic studies (Matthews, 1975) with selected radiolabeled chlorinated biphenyl compounds have confirmed the trend toward increasing biological half-life with increasing chlorine number. However, these findings may be the result of both higher accumulation rates and lower elimi nation rates for the higher chlorinated biphenyls. It may be of Interest to note that fish from lower and intermediate levels of the food web have been found (Zitko et al., 1972) to contain lower amounts of hexa and higher amounts of tetra and penta than higher trophic level white and silky sharks and aquatic birds This suggests that selectivity in bloaccumulatlon is also a function of bioepeclee. Again the effects of varying chlorine position are leae HONS 211891 clearly understood, end the best assessment of this comes from study of two or more members of an Isomeric series of the chlorinated biphenyLs. Recent work with five symmetrical hexachloroblphenyl isomers in chicks (McKinney er al., in press; Goldstein et al., in press) and subsequently in mice (Biocca. 1975) has demonstrated that separate and distinct differences in Isomer toxicity are possible end that the differences are related to chemical structure via effects of varying chlorine substitution on compound lipophillcity and metabolism. The hexachloro isomers studied were 3,4,5,3',4*,5'-; 2,4,6,2',4',6'-; 2,3,4,2* ,3' ,4' 2,4,5,2',4',5'-; 2,3,6 2\3\6*- and 2.3,5 2*, 3',5'-. Those hexa-isomera with 4,4'-substitution appear to be more rapidly accumulated and may be more slowly metabolized, l.e., these isomers showed greater accumulation in adipose tissue and increased activity in the liver and overall greater toxicity. These differences are believed to be associated with differences in molecular polarizability and to be measureable by spectro scopic and chromatographic techniques. Other workers (Ax et al., 1976; Ax and Hansen, 1975) have comparad the toxicity of two pentachlorobiphenyls In laying chicks. The isomer with 6,4*-subbticutloo (2,4,5,3',4') showed higher average embryonic mortality and teratogenicity In unhatched eggs but .lower decreased fertility in '-he chick than 2,3,6,2',3* the Isomer not chlorinated in the 4-position, Although it is difficult to assess the role of metabolism here, the data would suggest a HONS 211892 nor* rapid accumulation of the 2,4,5,3'4'-isomer In the egg. Other workers (Bush et al., 1974) have already observed a correlation between PCB content of eggs and embryo mortality and teratogenicity. Various other workers (Ecoblchon and Comaau, 1975; Johnstone at al., 1974; Hill et al., 1974) studying a range of chlorinated biphenyls have generally observed the importance of degree of chlorination as well as position of chlorlns sub stitution (especially 4,4'-subetltutlon) In overall biological effectiveness. Although some of the co^iouads studied are unrealistic as components of coerclal PCB mixtures, they have served as models to demonstrate the ii^ortance of chlorine nuo&er and position. Nevertheless, of 50 possible components of Arodor 1254 Identlfed by Sissons and Welti (1971) as many as 19 (38t) could have 4,4*-substltution. It is of Interest to note here that one of the possible persistent Isomers found In Yusho patient tissue has been tentatively identified (Kuratsune, 1975) as the 2,3,4,5,3',4*-heptachlorobiphenyl. This Isomer has been prepared (McKinney, personal cosmunication) and its toxlcty will be determined especially In relation to the highly toxic 3,4,5,3',4',S*-hexa Isomer previously tested. It hae been only recently that hydroxylated metabolites of PCBa have been Isolated and Identified (Jansson et al., 1975) in-environmental samples. The study of the metabolism of the commercial mixtures (deFreltaa and Norstrom, 1974) themselves has HONS 211893 received little attention for obvious reasons. Recently, the metabolism of purified chlorinated btphneyls, some radiolabeled for quantitation purposes, has been studied in a number of biological systems. Although there are a large number of publications in the literature on this subject, this report will concern Itself with those studies which seem to support general ities in terms of the effects of varying chlorine number and position on metabolism with particular reference to mammalian systems and to realistic components of Aroclors. The distribution and excretion of a series of four radio labeled (^C) chloroblphenyls which have degrees of chlorination similar to and are themselves constituents of Aroclors 1221, 1242, 1254, and 1260 have been studied (Matthews, 1975) in the male rat. These studies have clearly shown an increasing biological half-life with increasing chlorine number for the series (4-; 4,4'-; 2,4,5,2',5'- and 2,4,5,2',4',5'-) which is believed to be related to the rate of metabollam and the ease of formation of the arene oxide Intermediate. These compounds were also dosed at realistic exposure levels (0.06 to 6.0 mg/kg) which showed that little if any of tha chlorobiphanyl is excreted in unchanged form, Moet other metabolic studies (Hutzinger et al., 1974) with selected chj.oroblphenyls have dealt with much higher dose levels (generally resulting in excretion of much unchanged biphenyl) and unlabeled compounds which are of little quantitative va-lua In assessing the effects of chlorine degree and position. HONS 211894 There has been no similar study with selected chlorinated biphenyls within an isomeric series to assess the effects of chlorine position on metabolism with the possible exception of an incomplete study (Hass et al., unpublished) on the identifi cation of metabolites from the excreta of chicks (and mice) fed symmetrical hexachlorobiphenyl Isomers. It is essential that this be done with symmetrical isomers (in order to simplify the probelm of interpretation) and preferably in two or more isomeric series (tetrss and hexes). The various metabolism studies have generally shown the occurrence of polar hydroxylated compounds as major components of the metabolite mixture. As the degree of chlorination Increases, hydroxylatlon can be concurrent or concomitant with dechlorination (Hutzlnger et al., 1974). There is also evidence for the formation of methyl ethers (Safe, 1975) and methylchlorobiphenyls (Hass et al., unpublished) as metabolites. There Is an Increasing body of evidence (Safe et al., 1975) to support the arena oxide intermediate in hydroxylatlon, especially (Chen et al., Division of Nutrition and Disposition, in press) in cases where vicinal (1-2,-adjacent) unsubstituted carbons are found In the molecule. The corresponding dlhydrodlol and/or catechol generally occur along with the phenolic metabolites. Although! it has not been studied in msMSlian systems, there is at least one report (Baxter et al., 1975) that studies of MONS 211895 Individual chlorinated biphenyls do not accurately predict the rates of metabolism of the same compound In simple mixtures. Therefore, the overall problem of metabolic degradation of the commercial mixtures may be complicated by the possibility that certain FCBs are potent enzyme Inducers but poor substrates and vice verse with various degrees In between. This Is further compllcsted by the fact that certain chlorinated biphenyls occur in optically active forma and only one enantiomer may be biologically active and undergo enzyme Interactions. The existence of nine of the major, and ten of the minor, constituents of Aroclors 1242, 1254, and 1260 In optically active forma has been predicted (Kaiser, 1974). D. Determination of PCB Residues The determination of PCB residues has been reviewed In detail by the WHO Task Group report on "Environmental Health Criteria for Polychlorinated Biphenyls and Terphenyls" (WHO Task Group, 1975) and In "Chemistry of PCBs" (Hutzlnger et el., 1974). PCBs are lipophilic, quite similar In this respect to DDE, the metabolite of DDT. An analyses for PCB residues follow procedures the same as or similar to those used for multiple raalduee of orgt ichlorine pesticides (WHO Task Group, 1975; Hutzlnger et al., 1974; Food and Drug Administration, 1968-1975, Horvitz, 1975; Hearing Clerk, Dept. HEW, 1973), The Individual steps that follow sampling are extraction of raalduee from the sample. Isolation of residues from coextractives (cleanup). 48 HONS 211896 separation of PCBs from interfering chlorinated hydrocarbon pesticides, quantitation, and confirmation of identity. In general analytical methods capable of completely extracting residues of organochlorins pesticides from sample substrates and of quantitatively recovering the pesticides through subsequent cleanup procedures are also capable of achieving quantitative analysis for PCBs. The llpophlxiclty of chloroblphenyls which increases with increasing chlorine content may Influence their recovery through some analytical methods (Stalling at al., 1972). For example, recoveries through the frequently used cleanup step, partitioning of PCB from a petroleum ether or hexane solution of fat or oil to acetonitrile, exesed 95 percent for Aroclor 1242 but drop to 75-80 percent for Aroclor 1260 (Hearing Clerk, Dept. HEM, 1973). Overall ability of typical analytical methodology to recover PCBs added in vivo to food samples is shown by two interlaboratory studies con ducted within the Food and Drug Administration (FDA) and one study conducted for the Association of Official Analytical Chemists (AOAC) (Food and Drug Administration, 1971; Burks, 1972; Food and Drug Administration, 1973; Sawyer, 1973). Using method ology described in the FDA Pesticide Analytical Manual and in the Book of Official Methods of the AOAC, average recoveries and co efficients of variation for determination of Aroclors added as unknowns to different foods were: Aroclor 1254-fish, 74+9 per cent; Aroclor 1254-infant chicken, 89 + 22 percent; Aroclor 1242chicksn fat, 101 + 13 percent; Aroclor 1248-chicken fat, 96 + 0 U MOHS 211897 percent;Aroclor 1254-flsh, 75 + 14 percent; Aroclor 1260-fish, 75 + 15 percent. The three studies involving fish required seperecion of the PCBs from the DDT group before quantitation. Gas chromatography with electron capture detection is the moat widely used procedure for determination of PCB residues. Gas chromatographic columns with methyl silicone liquid phases are widely used under conditions that typically separate the Arodors and PCB residues into about 15 peaks (Fishbeln. 1972). The 10 percent DC-200 (or OV-101) column described In the FOA Pesticide Analytical Manual separates Aroclor 1254 into 14 peaks (Armour, 1972). These analytical column do not give a highly defined representation of the Aroclor or PCB residue; several of the peaks In Arodors result from mixtures of more than one chloroblphenyl (Sissons and Welti, 1971; Stalling and Huckens, 1971; Webb and McCall, 1972; Webb and McCall, 1973). A column prepared fro* purified Apleson L has been shown to separate a 54 percent chlorine PCB mixture into over 40 peaks (Jensen and SundatrSh, 1974), Separation of the PCB residue Into three fractions by chromatography on charcoal prior to examination on the Apleson L colissn has made it possible to characterise end quantitate nearly 60 technical PCB components. This method provides a way to get much needed, more detailed information on the compos1ton of PCB residues end might be used in special studies. The increased time and complexity of this approach would probably limit its application in regular monitoring analysis. 50 MONS 211898 The need to confirm residue Identity end the procedures available are similar for PCBs and pesticides. The multipeak gas chromatographic pattern of a PCB residue may be very similar to that of a conmercial Aroclor or, as in most biological samples, the residue peak pattern may be changed to varying degrees from that of a given Aroclor. The use of column chromatographic or chemical reaction procedures to separate PCBs from orgenochlorine pesticides Increases the certainty of the gas chromatographic identification. Procedures readily available to the residue lab oratory for confirming the Identity of PCBs include: halogen specific gas chromatographic detectors (Hearing Clerk, Dept. HEW, 1973), stability of the residue peak pattern after refluxing the extract with alcoholic alkaline solution (Young and Burke, 1972), perchlorlnatlon of the residue to the decachloroblphenyl derivative (Berg et al., 1971; Armour, 1973), and thin layer chromatography (Fehrlngar and Westfall, 1971; deVos and Pest, 1971). Hass spectrometry Is not readily available to many laboratories conducting analyses for PCBs nor would the expense justify regular use in monitoring programs. However, the use of mass spectrometry is encouraged for reeldues and samples which are unusual or signi ficant and for the occasional examination of a so-called routine sample. Two especially critical considerations are associated with the determlnetlon of PCB residues; (1) their separation from potentially Interfering orgenochlorine pesticides, particularly 51 MQNS 211899 DDT, TDE, and DDE or the multicomponent chlordane or toxaphene, and (2) quantitative measurement of che multicomponent PCB residue which. In biological organisms, is usually changed in relative amounts of chloroblphenyl components from commercial Aroclors or which may result from mixture of PCB from different sources. To achieve reliable residue results It Is essential that the analyst correctly make critical judgements and Interpretations In dealing with mixed residues of PCBs and organochlorlne pesticides and In quantitation of the multicomponent PCB residue. There Is no substitute for analyst experience in this analysis. PCBs are separated from certain pesticides In the usual cleanup procedures, a.g., adsorption chromatography on Florlsll or alumina or by gal permeation chromatography (Food and Drug Administration, 1968-1975; Stalling at al., 1972; Holden and Marsden, 1969). When DDT, TDK, DDE, chlordane, or toxaphene are present, ancillary procedures designed to separate these chemicals from PCBs must be used. The DDT analogs, particularly DDE, are the pesticide residues most frequently encountered in samples and DDE is the most difficult to separate from the PCBs. The effective electron capture gas chromatographic response for DDE is 20-30 times greater than for an equal weight of Aroclor 1254 and the response for DDT is only slightly less than that for DDE (Food and Drug Administration, 1968-1975). Without proper treatment these pesticides can readily interfere In the deter mination of PCBs. Some procedures utilising colun chromatography HONS 211900 <1? on alumina or Florist 1 separate DDT sod TDE and intentionally collect DDE and PCBs In the teas fraction (Holden and Marsden, 1969; Reynolds, 1969), which Is analyzed. In these cases the analyst oust exclude the DDE region of the gas chromatogram from the quantitative measurement of PCBs. Column chromatography on charcoal also has been proposed for separation of PCBs from DDE, DDT, and other organochlorlne pesticides (Berg et al., 1971). Column chromatography on silicic add Is probably the most widely used procedure for separating PCBs from DDT and analogs (Food and Drug Administration, 1968-1975; Armour and Burke, 1970). The technique Is fsirly lengthy and difficult to reproduce, requiring empirical standardisation In each laboratory (Sawyer, 1973; Masunoto, 1972; Edwards, 1974). The separation of DDE Is probably not 100 percent effective but under Ideal conditions practically all DDE can be separated; the uaual Inconsistency Is for a portion of the DDE to be eluted from the colum with the PCBs, requiring allowances to be made In PCB quantitation. The lower chlorinated PCBs present the greateet difficulty In separation from the DDT group by coliasn chromatographic procedures. DDE can also be separated from PCBs by oxidation to the dlchlorobenzophenooa, followed by a chromatographic separation of this more polar derivative from PCBs. DDT and TDE nay be dehydrochlorlnated to their respective olefins and similarly separated along with DDE (Mulhern et al., 1971; Collins et al., 1972; Trotter, 1974). .This technique has not received as much application as the chroma tographic procedures, probably because pesticides and lower HONS 211901 53 chlorinated biphenyls are destroyed or changed, preventing their determination. A variation on this approach used sodium dlchromate plus a minute amount of sulfuric acid, rather than dehydrochlorlnatlon followed by oxidation with chromium trloxlde in acetic acid. The dlchromate reagent Is reported to convert DDE quantitatively to the dichlorobenaophenone without affecting DDT, TDE, or any of the chloroblphenyls (Jensen and Sundatrom, 1974). Quantitation of PCS reaiduea Is done In most laboratories by comparison of measurements made on the multicomponent electron capture gas chromatograms of the residue and a known quantity of reference material. The PCB residue In biological aamplea, as mentioned earlier, is very likely to be changed In the relative amounta of chloroblphenyl componenta from any one Aroclor or the realdue may be a mixture of PCBe from different sources (Food and Drug Admlniatratlon, 1968-1975; Jensen and Sundstrm, 1974; Cook, 1972). The reaponse of the electron cap ture detector variea with the number and location of chlorine atoms in the biphenyl molecule (Gregory, 1968; Zitko at el., 1971). With the electron capture system described in the FDA Pesticide Analytical Manuel, the response/tailt/weight increases about 6 fold from Aroclor 1242 to Aroclor 1260 (Hearing Clerk, Dept. HZW, 1973). The halogen specific microcouloaatrlc and electrolytic conductivity detectors respond proportionally to the weight of chlorine present and in theory could provide a more accurate measurement of a PCB residue that is not exactly the MONS 211902 sum composition as the reference (Hearing Clerk, Dept. HEW, 1973). However, lower sensitivity, difficulty in mainraining optimum performance, and limited availability in residue labor atories has limited the use of these detectors. Unless the PCB residue and reference Aroclors are exactly the same in chlorobiphenyl composition, the gas chromatographic determination cannot be considered accurate; the greater the difference between residua and reference composition, the greater the deviation between determined and the true residue level (Hearing Clerk, Dept. HEW, 1973; Seezhoold and Stout, 1973). The most frequently used approaches to quantitation have been selected for the reference the Aroclor with the most similar gas chromatographic pattern to the residue and; (1) compared the response of a single peak in the residue with the response of the counterpart peak in the Aroclor reference; or (2) cohered the total response for several peaks from the residue to the total response for the counterpart peeks in the reference; or (3) compared the total response for all peaks in the residue with the total response for all peaks in the reference; or (4) in a greater effort to duplicate the residue peak pattern, prepared a reference made up of one or more Aroclors to simulate the gas chromatographic pattern of the residue, and compared the total response for all peaks of the residue to the total response for all peaks in the reference (Sawyer, 1973; Beeshold and Stout, 1973). Response has been measured in terms of both peak height and area (Sawyer. 1973). The latter approach which utilizes an Aroclor or s mixture 5 HQNS 211903 of Aroclors co duplicate the residue peak pattern is recommended by the A0AC for quantitation of PCB residues in certain foods (Horwitz, 1975). This is a currently accepted and practical way to quantitate PCB residues. Perchlorlnatlon of PCBs to decachlorobiphenyl has been suggested ss a means to improve precision sad consistency la PCB determination (Berg at al., 1971; Armour, 1973). This approach will not Improve the accuracy of quantitation, however, because proportions of Individual chlorobiphenyls in the PCB residue remain unknown; equal weights of individual chlorobiphenyls of different chlorine content result in different weights of decachlorobiphenyl. Contaminations found in antimony pentschloride, the perchlorlnatlon reagent, also detract from this procedure for quantitation purposes (Trotter and Toiag, 1975). An approach to quantitation which appears to have practical merit as well as offering improved accuracy in quantitation has been advanced by Webb and McCall (1973). The PCB reaidue is quantitated peak by peak In comparison to reference Arodors which heve been characterized as to the number of chlorines sod the fraction of total Aroclor weight represented by each electron capture peak after separation on a widely used (methyl silicone liquid phase) gas chromatographic column. The availability of carefully characterized Aroclors and evaluation in practice are required to fully evaluate the merits of this procedure. HONS 211904 Precision In the interlaboratory determination of PCBs is slightly less then with the coneon orgsnochlorlne pesticides. In several lnterlaboratory studies involving biological sample* end paperboard containing either added Aroclors or actual residues of PCB the coefficients of varistion are about + 20 percent. The results of studies conducted by the AOAC and the FDA involving samples containing added Aroclors have been mentioned above (Food and Drug Administration, 1971; Burke. 1971; Food and Drug Administration, 1973; Sawyer, 1973); the recoveries of sdded Aroclors ranged from 74 to 101 percent and coefficients of variation from 9 to 15 percent. The levels of Aroclors added in these studies ranged from about 2 to 8 ppm with the exception of 0.2 ppm Aroclor 1254 added to chicken infant food. With samples containing actual residues, 9 laboratories in the AOAC study reported 9.2 ppm + 8 percent for a residue in chicken fst snd 4.5 ppm + 20 percent for a residue in Lake Michigan chubs. The residue in the chubs was determined after separation of DDE, DDT, and IDE by colissn chromatography on silicic acid. In s study by 8 laboratories in cooperation with the International Council for the Coloration of the Sea, PCB residues determined in a fish oil averaged 1.97 pfm + 47 percent. A much better coefficient of variation, about + 11 percent, was obtained when the same fish oil was fortified with en additional 10 ppm PCB (International Council for Exploratoin of the Ses, 1974). In an AOAC study of the method for PCB in paperboard 11 laboratories analysed a paperboard sample manufactured to contain Aroclor 1242 and reported 5.6 ppm + HONS 211905 16 percent (Flnsterwalder, 1974). The inter- or intra-laboratory precision of the determination of PCB in any sample type is improved by use of the saw analytical procedures, especially in quantitation of the residue. ' The lower limit of quantitation for PCB residues will vary among laboratories depending upon the objectives of their analyses and upon the particular details of the analytical methods, especially the sensitivity of gas chromatographic detection and the sample size. The quantitation limit achieved will be higher than for organochlorlne pesticides because of lower effective detector response to the technical PCB mixtures; 20-30 tiass more Arodor 1234 or 30-30 tlws more Aroclor 1242 than DOE is required for the mejor peaks to produce the saw peak height as DOE in electron capture gas chromatogrsphy (Food and Drug Administration, 1968-1975). Other factors which wy restrict the attainment of low limits of quantitation and affect analytical reliability are interfering residues in the sample and laboratory contamination. Fish from some locations contain residues of organochlorlne pesticides and possibly other contaminants at levels that would cause increasing difficulty as the FCB residue decreaws below about one ppm. Contamination of laboratory equipment, reage'"s, and aaeples with PCB from containers, previous samples, control runs, and unknown sources, must be carefully guarded against in analyses where s low level of quanti tation is necessary, for example with htain blood or certain 33 HONS 211906 environmental samples (Trotter. 1975; Jensen et al., 1972; Glam and Wong, 1972). Perchlorlnatlon of the PCB residue to decachloroblphenyl offers the possibility of about 25 fold increase In sensitivity in the gas chromatographic determination (Armour, 1973). However, contaminants present In antimony pentachlorlde severely restrict or prohibit application of this procedure to determination of low lavala of PCBs (Trotter and Young, 1975). Analytical methods used by the FDA for PCB residues also recover chlorinated napthalenes (Armour and Burke, 1971). If present alone, chlorinated naphthalenes would be recognized by the analyst. However, their presence in admixture with PCBs would probably go unrecognized unless amounts were greater than PCBs or the residue was examined by mass spectrometry. Chlorinated naphthalenes are oxidized by procedurea used to differentiate DDE In the preaence of PCBs and could be removed from interfering la the PCB determination (Holmes and Walden, 1972). From examination of procedures used to analyze for chlorinated dlbenzofurana In coMerclal PCB formulations and from analytical studies with chlorinated dlhenxodloxlns It can be Inferred that soma chlorinated dibensofurans may be recovered along with PCBs through analyr' -al aathods using Florlsil and column chromato graphy (Vos, et al., 1970; Roach and Pomarantz, 1974; Bowes, et al., 1975; Porter and Burks, 1971). The chlorinated dlbenzofurans have gas chromatographic properties similar to the PCBs (Bowes, 1975). "OWS 211907 59 Ic Is very doubtful, however, that a relatively snail amount of chlorinated dlbenzofuran In the presence of PCBs would be recognized In the usual analysis for PCBs. Little study has been made of residue analytical methodology for chloroblphenylols (hydroxylated chlorinated biphenyls). Procedures used for their determination differ substantially from those used for PCBs (Bachs and Link, 1973; Zltko et si., 1974). It Is not likely that chloroblphenylols would be recovered through column chromatographic and separation procedures used for PCBs. In the discusslm of PCB residues In foods given In the report of the Himian Exposure Group a large share of the Informa tion cornea from the PDA's surveillance programs. The analytical methodology used In the analyses la described In the PDA Pesticide Analytical Manual, Vol. I, and in Official Methods of Analysis of the AOAC. Quantitation of PCB residues la by gas chromatography with electron capture or halogen specific electrochemical detectors. The total response (eras or peak height) for the PCB residue Is coepared to the total response for the Aroclor reference (or mixture of Aroclors) having the meet similar gas chromatographic pattern. Aroclor reference materials are esch from s single master lot. Results are reported as-ppm of the Aroclor(s) used for the quantitation referenceWith certain exceptions the limit of quantitation for PCBs (baaed on the electron capture detector response to Aroclor 1254) Is 60 HONS 211908 about 0.2 ppm for individual foods and about 0.05 ppm for Total Dlat compos lets. Fish from certain fresh water locations are the food most frequently and consistently found to contain PCS residues. The residues in fish are most often similar In gas chromatographic peak pattern to Aroelor 1254 but usually with higher concentrations of the late eluting (higher chlorination) chloroblphenyl components. E. Chlorinated Dlbenzofurana (See Appendix for structural Information and definitions) In considering the potential hazard to humans of the commercial chemical mixtures of chlorinated biphenyls called PCBe, one must consider which If any Identified trace contaminant In these complex mixtures might contribute significantly to the overall hesard potential of the mixture. Although there is evidence for the presence of chlorinated napthalenes and possibly chlorinated terphonyls in PCBs, the chlorinated dlbenzofuran (Cl* DBF) contaminants are regarded es a greater potential danger for several reasons. In using a chick bloassay to monitor fraction* at Ion of coairclal PCB mixtrues, It was fotaid (Vos et al., 1970) that the fraction most toxic to chicks, and far more toxic chan the other fractions, contained Cl-DBFs and chlorinated naphthalenes. What limited toxicological Information there Is available on these halogenated naphthalenes, terphenyls and dlbenzofurana suggests that only the Cl-DBFs may be more toxic than chlorinated biphenyls by orders of magnitude (Bauer et al., 1961; Koore et aL, 1976) 61 HONS 211909 The extremely high toxicity of some of the chlorinateddlbanzo-p-dloxlns (NIEHS Conference 1973; Schwetz, et al.. 1973) a claas of organic chemicals very similar m structure to the Cl-DBFs, also suggests the latter are likely to be highly toxic. Knowledge of the chemistry and toxicity of various chlorinated dlbenzo-p-dloxln structures and the relationship between toxicity and structure, although recently developed, is more complete than for Cl-DBFs. Therefore, findings In the chlorodioxln field provide Indicators to assess the difficulties lllcsly to be encountered in the study of Cl-DBFs. Chlorinated dibenzofuran contaminants have been reported and confirmed In PCB mixtures manufactured in Germany and France (Tos et al., 1970; Bowes, et al., 1975a) in Japan (Roach and Pomerantz, 1974a; Kagayama, et al. 1975) and In the United States (Arodore ) (Roach and Pomerantz, 1974b; Bowes, et al., 1975e). An additional claim that a d-DBF contaminated an Arodor was not sufficiently supported by the evidence pre sented (Curley et al., 1975). In the PCBs of U.S. manufacture, the range of Cl-DBFs found was from dichloro through hexachloro (see referencee cited above) and sufficient evidence was obtained to show that 2,3.7,8-tetrachloro and 2,3,4,7,8 -pantachlorodlbenzofursn ere presen* in Aroclors (Bowes et al., 1975b). That the Cl-DBFs reported were actual contaminants and not artifacts resulting from the experimental procedures used, was considered in two instances (Roach end Pomerantz, 1974b; Nageyama et al., 1975). 62 HONS 211910 Proper quantitation of the Individual Cl-DBF contaminants is very difficult. It requires full structural identification of the Cl-DBF contaminant, availability of the contaminant as a highly purified reference standard for quantitation purposes and evaluation of the procedure used to concentrate and separate the Cl-DBFs, (e.g. from chloroblphenyls) to determine the capability of the procedure to recover the specific Cl-DBFs In the PCB mixture. Reported attempts to quantitate the Cl-DBFs In PCB suggest contamination levels In the low parts per million range for total chlorinated dibensofurene (Bowes et el., i975a; Nagayarns et al., 1975)- Many of the reported values probably can be taken as minimal in the absence of recovery date. Although the source of the Cl-DBF contaminants In commercial PCB mixtures has not been determined, several possibilities may be considered. The simplest explanation would be the likely presence of dlbenzofuran (parent compound) In the technical grade biphenyl subjected to Che chlorination process. From e consideration of the procedures used in the coMercial synthesis of PCBs (Hubbard, 1964) various chlorinated biphenyls, if substituted in the ortho and orcho-prime positions with hydroxy groups and/or chlorine atoms, might ring dose to form the furanold ring by dehydration or dehydrochlorlnstion. It is important to recall that Cl-DBFs have been reported as contaminants also in other chemicals of commercial importance. Samples of pentechlorophenol (Schwetz et el., 1974) as well as 63 *0N$ 211911 lovar chlorinated phenols (Firestone et el., 1972) and hexachlorobensene, (Villanueva et al., 1974) have been found to contain a range of Cl-DBFs- The ppta level of Cl-DBFs In certain pentachlorophenoIs examined by Schwetz and co-workers (1974) was considerably higher chan the highest level thus far reported In commercial PCB mixtures; the chlorine level in the reported dibenzofuran contaminants of pentachlorophenol ranged from hexa to octa. It la now known that "PCB residues'' obtained from environ mental samples usually do not simply represent an easily Identified original cosercial PCB mixture. For example, there is evidence from the study of electron capture gas chromatograms that certain chlorinated biphenyl compounds are preferentially lost or concentratad in passing through environmental media. This has led to recent studies of the toxicity, metabolism and physiological effects of specific, individual chlorinated biphenyl compounds, synthesized by routes intended to yield a single com pound of known structure. In addition to permitting the development of structure-activity correlations, these studies presumably were intandsd to avoid complications in Interpreting results from the tasting of tha commrcltl chlorinated biphenyl mixtures (Vos and Sotenboom Bam, 1<*`,2), now known to contain toxic Cl-DBFs as contaminants. Intarpretaion of rasults from the testing of individual chlorinated biphenyl compounds, however, may not be entirely straightforward. In the Ullaann coupling reaction of HONS 211912 6 2,4,5-trichlorolodobenzene, lc has baan raportad chat cha highly toxic 2,3,7,8-tetrachlorodlbenzofuran Is formed In 3Z yiald In addition to che expected 2,2'4,4',5.5'-hexechlorobiphenyl product (Moron, et al., 1973), This hexachloroblphenyl Is a significant constituent of PCB mixtures (Sissons and Elci, 1971; Tas and Kleipool, 1972; Jansen and Sundstrom, 1974) and has been singled out for biological studies by several workers (Voe and Notenboom-Ran, 1972; Johnstone ec al., 1974; Hansell and Ecoblchon, 1974). Careful purification and, if necessary, chemical analysis of symmetrical chlorobiphenyls prepared by the Ullmann coupling reaction (for dibensofuran content) Is sug gested as a prerequisite co biological testing of the biphenyl. There are two ocher aspects of the cheelcal relationship betwen chlorinated biphenyls and Cl-DBFs Chat need to be nantloned. Both deal with possible environmental alteration. Evidence has been obtained for photochemical conversion of cercsln chlorinated biphenyls co Cl-DBFs in very low yields (Crosby and Mollanen, 1973). Although the scope of this study was United, formation of Cl-DBF was shown co occur under conditions of sunlight irradiation and In che laboratory using sunllghc-sinulating conditions. Two different ortho-chlorinated biphenyls (the 2,5-dlchloro and 2,2*,5,5'-tetrachloro) were claimed co produce approximately 0.2Z steady-state yields of a nonochloro DBF. The limited reports regarding che photochemical decomposition of Cl-DBFs (Crosby and Mollanen, 1973; Hutzlnger et al., 1973) indicate relatively 65 HONS 211913 rapid daatruetion of tha compound! may taka place in the environment. Since reductive dechlorination la a major route of photochemical alteration of Cl-DBFa, the poaalbllity of dechlorination of highly chlorinated DBFs to more toxic DBFs of lower chlorine content muat be conaldered. The extent to which apeclfic chlorinated biphenyle, found aa major constituents of commercial FCB mixtures, can be photochemically converted to Cl-DBFa can only be determined by further experimental studies. Analysis of the Yusho oil (the rlcs oil contaminated by PCB-contalnlng heat exchange fluid Implicated in the Japanese "Yusho" poisoning incident) by Nagayama at al., (1975) for Cl-DBF content, led to a value of 5 ppm Cl-DBFs. This value is about 300 times the Cl-DBF level expected in the Yusho oil if one simply assumes contamination of the oil by Cl-DBFs to be proportional to the Cl-DBF level found in unused Kenechlor 400 (the Japanese FCB mixture claimed to have been used as the heat exchange fluid) and to the level of PCB in the Yusho oil. This led Ruratsune at al., (1975) to suggest that Cl-DBF levels increased in the hast exchange fluid through use. Going a step further, these questions may be raised: Is Cl-DBF concentration increased, generally, in PCB-containing hast exchange fluids through use? Does such increase occur in PCB-containing trans former fluids or electrical capacitors through use? A critical review is needed of the published data upon which Kuratsune's suggestion is based (Nagayama at al., 1975). particularly 66 HONS 211914 Inc* th* results from a recent analysis of a portion of Yusho oil (Trotter, 1976) raises questions about the concentration of chlorinated biphenyls In the Yusho oil. Further laboratory studies suggested by such a review and analyses of selected, used PCB-contalnlng Industrial fluids night help to answer the above questions. Considering th* Cl-DBFs as th* coxlcologlcally nost signifi cant class of chaalcal Impurities In th* FCB mixtures, tha questions may ba asked: what portion of th* observed effects from animal and human expoaura to PCB mixtures can be attributed to the Cl-DBFs? It la doubtful thia question can ba answered in any quantitatively explicit way. Thar* are analytical problems in Identifying and measuring accurately tha amount* of chlorinated dlbenzofurans In varloua PCB mixture*. Evan though only a relatively few of tha 135 poaalbl* chlorinated dlbenzofurans may be identified In PCB mixture*, th* toxicity of th*** compound* can ba expected to vary with both mabar and ring poaltlon of chlorine atom* In the Cl-DBF molecule. Toxicity data on apeciflc, high purity Cl-DBFs Is very limited because the compounds themselves are not readily available. In addition to tbesa, essentially chemical, difficulties, toxico logical Judgements would ba difficult to make. To begin to evaluate tha toxicological "burden" to be placed on Cl-DBF contaminants In cosercial PCB mixtures, a practical approach would be to use analytical procedure* that can recover and quantitate specific Cl-DBF compounds known to ba highly toxic. For example, 2.3,7,8tetrachlorodibanzofuran, already reported by Bowes (1975b) a* O7 HOWS 211915 prune In PCBe, can serve as a measure of potential hazard until further toxicity data for Cl-DBFs become available. To dace, there have been no published, positive findings of Cl-DBFs in environmental samples or In foods. Analytical pro cedures to detect Cl-DBFs In such samples should be developed, tested and applied to appropriate samples. If chlorinated dibenro-p-dioxin residue analytical findings can be taken as an Indicator, procedures suitable for parts per trillion detection of Cl-DBFs will be required and quantitation will be difficult. No clear evidence for the presence of chlorinated dlbenzo-pdloxlna In cover dal PCB mixtures has been reported. A recent publication suggesting the presence of such contaminants in some PCBe and in a synthesized chlorinated biphenyl (Ax and Hansen, 1975) almost certainly misinterpreted the significance of the experimental finding related to the dioxins. III. Chemistry of Brominated Biphenyls A. Covarison of Brominated Biphenyls with Chlorinated Biphenyls Dnlika PCBe, the chemistry and stability of PVBs have not been mall studied and doc--ntad in the literature. It is difficult to assess the stability and the extent of possible chemical conversion of PBBa in the environment. PBBs can be compared chemically to the PCBs. Since both bromine and chlorine are 68 HONS 211916 halogens, PBB and PCB chemistry should be similar In some respects. Bromine, however. Is a better leaving group In chemical reactions than chlorine- The described laboratory experiments concerning PBB alkaline hydrolysis and photolysis show that bromine Is more labile than chlorine under comparable reaction parameters. PBBs may. therefore, be leas stable In the environment If PBBs were under the same physical reaction para meters In the environment as PCBs. The physical state and other physical reaction parameters of PBB and PCB residues In the environment can determine reactivity. Liquids and vapors often react more readily than solids. Aroclora are liquids, though some Aroclora are extremely viscous at room temperature. Soma PCBe, especially the lower chlorinated components, are volatile. PlreMaster BP-6, however, is a solid and has an extremely low vapor pressure. The production, distribution, and usage of PBBs has not been as wide spreed as PCBs. PBBs,unlike PCBs, may not be physically locstad In a position for chemical reaction. PlreMaster BP-6 has been extensively used, for example, in thermo plastics, such as typewriter and business machine housings (Karst. 1974). PlreMaster BP-6 has little tendency to migrate from the thermoplastic Into which it is Incorporated. The major portion of the products Into which PlreMaster BP-6 Is lncorporsted Is assumed to be burled eventually In refuse dusps. PCBs, on the other hand, are often used In transformers and capacitors and can be exposed to high temperatures which can accelerate possible chemical reactions. PCB residues are found In many diverse locations 69 HONS 211917 throughout moot of the world. Many of these locations may offer conducive settings for chemical and metabolic conversions. PCB residues In a body of water may react photolytlcally, whereas PBB residues burled In a refuse dump would not be In a position to absorb light. B. Non-metabolic Alteration of Bromlnated Biphenyls Oxidation and Hydrolysis the stability of PBBs to oxidation has not been studied and dociimantad in the literature. FireMaster BP-6 is unstable to alkaline hydrolysis. After refluxing FireMaster BP-6 with ZZ KOH In ethanol, CLC chromatograms show erratic degradation of hexabronoblphenyl (DT,DO,DCT,DCH,1976), the major component of FireMaster BP-6. The possible rata of PBB hydrolysis in the environment under milder conditions is not known. Photochemistry Recent analysis of FireMaster BP-6 by GCMS using an OV 101 coluK demonstrated 12 peaks whose mass spectrum corresponds to 2 pentas, 6 hexes, 4 heptaa and 2 octas. However, about 50 percent of the material is 1 haxa isomer with the next most abundant component being the hepta isomer at about 20-25 percent. Nuclear magnetic resonance and chemical studies have Identified the principal component of Firenastar BP-6 as 2,2'4,4'5,5'-hexabromobiphenyl (Andarson at al., 1974; Sundstrom et al., 1976). HONS 211918 The photolytic rates of reactivity of FlraMaster BP-6 and 2,2,4,4'5,5'-hexachloroblphenyl in methanol solutions Irradiated with 3000 A light were compared (Ruzo and Zablk, 1975). The rate of reaction of FlreMaster BP-6 was determined presumably by the decrease In the GLC peak of hexabromoblpenyl. Hexabromoblphenyl was found to be seven times as reactive ae the corres ponding PCB. Hexebromoblphenyl was found to undergo photolytlc reductive debrominetion in methanol yielding penta- and tetrabromoblphanyls and also > 12 methoxylated product. Dlmethoxy tetrabromoblphenyl was Identified as a product by mass spectrometry The highly efficient photoreactivity of PBB may be due to: 1. enhanced lntersyetem crossing to a triplet state due to vlbronlc coupling with the bromines, 2. sterlc Interference due to the bromines, and 3. the relatively low carbon-bromine bond energy. The aromatic carbon-bromine bond energy la 71 kcsl/mole vs the aromatic carbon-chlorine bond energy of 86 kcal/mole (Kerst, 1974). As with PCBs (Ruzo, et el., 1974), presumably ortho halogens pre ferentially cleave in PBBe upon photoexcitation. The poesible rates and extent of photolytlc reactivity of PBBe In the environment are not known. The photochemistry of PBBs has net bean studied in the vapor or solid states. Since FlraMaster BP-6 has an extremely low vapor pressure, vapor state photochemistry In tha environment would be extremely limited. The extent of thin film PBB photochemistry In the environment aey also be Halted. PBB contamination and resulting HONS 211*19 phocochnistry In river waters, except In the vicinity of PBB production facilities, nay ba limited. PBBa Incorporated Into thermoplastics, which are presumably burled eventually In a refuse dump, are not likely to absorb much light for photolytlc reaction. C. Bromlnated Biphenylsi Determination of Residues Polybromlnated biphenyl (PBB) residues ars analysed for In foods by methods quite similar to those used for organochlorlne pesticides and polychlorinated biphenyls (PCBs) (Pood and Drug Administration, 1968-1975; Fehrlnger, 1975; Pood and Drug Administration, 1976). Recoveries of PBB added to samples In vivo through these procedures Is over 80 percent. A method to Improve extraction efficiency of PBB residue from dry, high fat animal feeds has been reported (Fehrlnger, 1975). The mejor difference from methods for organochlorlne pesticides Is In the gae chromatographic decsrmlnation which is done at higher temperatures or on colusis with low liquid phase loads because of the lower volatility of FBBs. Methyl silicones or Sllar-lOC, the liquid phases most frequently used, separate the technical PBB, FlraKsster BF-6, into 3-9 peaks depending on the chromatographic conditions selected. PCBs and organochlorlne pesticides which would be recovered through extraction end cleanup procedures together with PBBa have earlier retention times under these conditions and ars separated from the MONS 211920 72 major PBB constituent, hexabromoblphenyl. Tha retention times of hexabromoblphenyl end deeachloroblphenyl are not greatly different on methyl silicone liquid phases but are widely separated on Sllar-lOC. Elution of the Florlsll column used for cleanup of extracts with petroleum ether rather than a mixture of petroleum ether and ethyl ether separates FBBs from most organochlorlne pesticides prior to gas chromatography end Improves cleanup for the PBB determination. PBB residues are detected and quantitated vlch the electron capture gas chromatography detector. Quantitation Is based on comparison of the size of the hexabromoblphenyl peak In the residue to the size of the hexabromoblphenyl peak In a known weight of the rsference material, PlreMaater BP-6. FlreMaater BP-6 Is a technlcsl mixture containing 60-70 percent hexabromobiphenyls (Sundstrom and Hutzinger, 1976). The accuracy of residue quantitation is affected by any change in the composition of the residue from that of the rsference technical PBB mixture. The identity of PBB residues can be confirmed by thin layer chroma tography, photochemical alteration, halogen-specific gas chromatographic detection, and by different retention times on Sliar-10C and mathyl silicon gas chromatographic colims (Pood and Drug Administration, 1976; Fehrlnger 1974; Fehringer, 1975; Erney, 1975). Mass spectrometry has also been used to confirm the Identity of the PBB residue (Ayers, 1975). 73 MONS 211921 The limit of quantitation for PBB residues, including capa bility for residue identity confirmation, Is about 0.05 ppm In fata and 0.01 ppm in non-fata (Food and Drug Adminlatration, 1976). Formalized Interlaboratory atudlea have not been reported for analytical methoda for determination of PBB realduea. D. Possibility of Bromlnatad Dlbenzofurana in Co--rcial PBB Mixtures Thera has been no report, so far, of the finding of brominated dibanzofuran (Br-DBP) co^ouads in ctwrclil PBB mixtures. Examination of FlreMastar BP-6 for possible contamination by Br-DBP is in progress in several laboratories. MONS 211922 74 METABOLISM AMD BIOCHEMICAL TOXICITY OF FCBs and FBBa I. Effacca of PCBa on Blocham'tfi 'F"ctiona One of the major biochemical effacca of PCBa la the Induction of mlcroaomal enzymea In che liver. Rlsebrough ec el., (I960) suggested chac PCBa had cha capability to lnduca the accivltiaa of mlcroaomal enzymea. Subsequently, Street at al., (1969) demonatrated che Induction of liver enzymea In rata by PCB'a. Since then, a great number of articlea have been publiahed on chia aubject. The enzyme ayatema atudied have included mainly hydroxylases, N- and O-dsaathylasee and nitroreductaaea, and to a lasaer extent nonapeciflc carboxyleateraae, bromoaulphophthalein-glutathlone conjugating enzyme, p-nitrophenol UDP-glucuronyl tranaferaae and EPN-detoxlficetion ayatema. The induction of mlcroaomal enzymes by coMercial PCBa haa been demonstrated perorslly In rabbits (Villenauve at al., (1971), rata (Littarst at al., 1972a) and primates Allan at al., 1974) and via lntraperitonaal injection (Bickers at al., 1972) and skin application In rata (Bickers at al., 1975). Values reported for the threshold of enzyme Induction by PCS'a vary between 0.5-25 ppm. (Littarst at al., 1972b; Villenauve at al., 1971; Turner at al., 1974). The time-course of mlcroaomal eyzyme induction was studied by Littarst at al., (1974) in rats. They found that significant lsvels of enzyme Induction occurred after 7 days of feeding FCBs 75 HONS 211923 la th( dl(t> A single oral dosa la rats raaulcad la enzyme activities at 24 hours. Blckacs at al., (1974); 1975) hava shown chat maximum induction occurrsd within 2-6 days la rats aftar cuta neous exposure to Aroclor 1254 or microscope iasLersion oils con taining 30-4SZ chlorine. In order to study the affect of chlorine content of PCB's on enzyme induction, different coMercial PCBs were investigated. Littarst at al., (1972b) studied PCBs with chlorine contents from 42 to 60Z. In rats the maximum activity of deaethylase was observed with the 54Z chlorine PCB's; activity for nitroreductase was obtained with the 60Z chlorine PCBs. Chen at al., (1973) reported that 54Z chlorinated PCBs gave isarlue response of denethylaae in rats. Similar results were reported by intraperi- toneal injection of various PCBd in rats (Scobichon et al., 1974), Goldstein et al., (1975) and Iverson et al., (1975) studied the effects of Aroclor 1016 and Aroclor 1242 on enzyme Induction in rats. Both PCBs have approximately 42Z chlorine, but the content of the penta-, haxa-, and hepta-ehlorlna isomers of Aroclor 1016 is reduced to 10Z of that in Aroclor 1242. Their results showed that Aroclor 1242 was a much more potent inducer than Aroclor 1016 in female rata. In comparing Aroclor 1232, Aroclor 1248 and Aroclor 1280, Sehmoldt et al., (1974) found that Aroclor 1260 was the moat potent enzyme Inducer in rats. The synthetic and iaomarically pure PCBs have been investi gated for enzyme Induction, Chen et al., 91973b) found that both 76 HONS 211924 2,3,4,5- and 3,5,3',5'-tetrschlorobiphenyls induced enzyme ayeteas in male rate, but no significant differences were found between the two iaoaere. Johnstone et al,, (1974); Ecobichon et el,, (1975); Ecobichon (1975) compared the effects of position of chlorine atoms on the ring. They found that enhanced induction of mono-oxygenases was observed for PCBs having chlorine stoma substituted at the 4and 4'- positions irrespective of chlorination at other positions. Substitution at the 2-positlon was next in importance followed by substitution at the 3-position, They concluded that the position of chlorination was as important as the dsgree of chlorination. There are two types of enzyme inducers being reportsd in the literature. One group to which phenoberbital belongs resulted in Increased cytochrome F-450 content, as well as increased benzpyrene hydroxylase and ethylnorphlne demethylase activities in the liver. The second group includes polycyclic hydrocarbons. This group stimulates the formation of cytochrome P-448 and an increase in hydroxylation but not demsthylation, Alvares et al., (1973) reported that rats treated with PCBs produced an increase in cytochrome P-448, hydroxylase, as well as demethylase. There fore, it appears that PCBs display induction behavior typical of both groups. Moat of the enzyme induction studies were evaluated in vitro. Several workers Indeed have observed enzyme induction affects in vivo by demonstrating shortened barbiturate sleeping times (Bickers et al., 1972; Johnstone et al,, 1974; Villeneuve et el-. 1972); 77 HONS 211925 CoMarclal PCS* are known Co contain mail amounts of dibenzofuran* (DBF). DBF* war* found co b* 170 tin** a* potent a* FOB* In Inducing enzymes. Alvales & Kappas (1975) r*port*d tha Induction of hydroxyla** and daaathylas* in placenta) as wall a* in th* fatua, when preg nant rats were treated with PCBs Valnio (1974) reported chat Clophen A50 enhanced hydroxylase 5-fold In the lung and 8-fold In the kidney sdcrosonas, whereas the adcrosomes fro* duodenal mucosa exhibited no enhancement. Benzpyrene hydroxylase activity In skin waa also Increased by PCBs (Bickers et al., 1975). Tos end Koeman (1970) found that several tissues of FCB-traated chickens were strongly fluorescent under ultra violet light. They suggest that PCBs could Induce ehamlcal porphyria In chickens. Later, they demonstrated that mitochondrial ALA Synthetase activity Increased 20-fold and fecal porphyria level* were significantly Increased in Japanese quail treated with PCBs (Toe, 1971; Tos sc al., (1972). They suggested that the porphyria caused by PCBs Is due to the increase of ALA synthetase, followed by overproduc tion of porphyrins. Slnllar result* In rat* were obtained by Goldstein et al., (1974), end Bruckner et al., (1974). Goldstein et al., (1974) suggested that the Induction of ALA synthetase Is probably not the primary cause of PCS-induced porphyria because of the delayed onset of porphyria as coapared with ALA synthetase induction. They suggest that the ALA synthetase In the liver say be related to the Increase of cytochrome P-450 content. This NQNS 211926 78 idea vu supported by Grot# et al., (1975), Aroclor 1242 vaa found to ba a mora potent Inducer of liver porphyria than Aroclor 1016 (Goldateln at al.> 1975). Both Aroclora have similar chlorine content, but Aroclor 1242 contains 9Z homo logs with five or more chlorines while Aroclor 1016 contains only 1Z of such homologs. Sinclair and Cranlck (1974) showed that cultured liver cells respond rapidly to PCBs In accumulation of uroporphyrin. Simon et al., (1974) suggested that porphyria may be due to the destruc tion of the phospholipid structure of cell membrane, resulting In Increased permeability of the substrate. Hirayama at al., (1974) observed hypo-blllrublnaeaia in Yusho patients and suggested that the induction of bilirubin UDP-glucuronyltrsnsferase may be respon sible for the low plasma bilirubin content. However, recently Bastomeky et al., (1975) shoved that the lowering of serum Bilirubin concentration by PCBe la due to the reduced binding of bilirubin to plasma protein rther then to the enzyme induction. Wit (1972) has shown that PBBe which are similar to PCBs are potent heptoporphy-rlnogealc chemicals. The chemical and structural similarity of DDT to PCBs led Jefferies et al. (1972) to study the effects of PCBs on thyroid functions. They found an increase in thyroid weight in FCBtreated black-backed gulls as compared to controls. Hurst et si., (1974) found no clear-cut effect of PCBs on thyroid gland size in <tualla. Byrne et al., (1975) reported that PCBs Increased HONS 211927 79 thyroxin* levels In the blood and also promoted peripheral degrada tion of :hyroxine In female minks, Bastomsky (1974) found a 4 to 5-fold Increase of biliary excretion of thyroxin* In rats treated with PCBs, PCBs also elevated iodin* uptake by th* thyroid and reduced serum PB1 concentration. Recently Bastomsky and Wise, (1975) shoved similar results In rats due to either lntraperitoneal injection or skin application of microscope inversion oil. Walker (1975) found that inclusion of Aroclor 1254 la fish food resulted in an increase in fish thyroid activity, Th* effects of PCBs on ATPase and oxidative phosphorylation have been examined by a number of investigators. Na'*Tf-ATPase and Mg44, ATPase from several fish tissues vers found to be inhibited by PCBs. (Tap *t al., 1971; Cutkomp et al., 1972; Desaiah et al., 1972; Koch et al., (1972). Further study showed thet in vitro data were not in agreement with the in vivo data (Tap et al., (1974). The inhibition of ATPase in fishes by PCBs was also reported by Kinter et al., (1972) and by Davis et al., (1972). Sivalingan et el., (1973) investigated the effects of PCB on the oxidative phosphorylation of rat liver mitochondria. They showed that the mode of inhibition appears to be different with PCBs of different chlorine content. PCBs with low chlorine content inhibited energy end electron transfer but PCBs with high chlorine content not only inhibited energy and electron transfer but also had an uncoupling effect. LaKocca et al.. HONS 211926 80 (1975) showed Chat th administration of comsrcial PCBs, as wall as purified isomers Co rats inhibited the total ATPaae activity in liver, kidney and brain tissues. Pardini (1971) has demonstrated that a marked inhibition of respira tory enxyae systems occurred when heavy beef heart mitochondria were exposed in vitro to numerous PCBs. Chesney and Allen (1974) showed that, the addition of PCBs to rat liver mitochondria in vitro caused an inhibition of oxidative phosphorylation and respira tion. Feeding rats with PCBs at the 1000 ppm level increased the oxidative phosphorylation in liver mitochondria, however no effect was seen at the 100 ppm level. Sharp et al., (1974) reported that the inhibition of beef brain and rabbit kidney Na+K+-ATPases can be reversed or prevented by phoaphatldylaerlne or phoephatidylinosltol, but not by phosphatldychlollne and phosphatldylethsnolamlne. They suggested that acidic phospholipids are required to stabilise the enzyme and thereby overcome the effect of PCBs. Risebrough et el., (1966) suggested that PCBs had the capacity to enhance aterold-hydroxylating enzyme activity, thus affecting estradiol metabolism. In addition, certain low chlorine containing PCBs have estrogenic activity which is reflected by an increase in glycogen content of the uterus of immature rats (Bltmsn and Cecil, 1970). Similar results were obtained by Fumlko (1972) in rats (Bltman and Cecil, 1970). Similar results were obtained by MOMS 211929 81 Fumiko (1972) 1& rats and by Orbarg, and Klhlstrom (197 ) 1& mice. Orberg at al., (1972) found that tha aitroua cycle of the mouse Increased In length after a single injection of PCBs. Linear and Peakall, (1973) reported an Increased rate of metabolism of estra diol in vitro in the liver of American kestrels after the feeding of PCBs. Similar results were also reported in white leghorn cockerels and pullets (Nowlckl and Norman 1972). Platonov and Funnell (1971) demonstrated that feeding PCBs to cockerels resulted in a decrease in testicular and comb growth. Decreased urinary excretions of estrogen and dehydroeplandroeterone were also observed in cockerels when PCBs were administered in high doses (Platonov at al., 1972). Ecobiehon and Mackenzie, (1974) studied the uterotropic activity of eoasrclil and lsomerically pure PCBs in rsts. They found significant changes with a number of PCB mixtures but experiments with the laomorically pure PCBs were inconclusive. Chsnges were observed with 2-chloro and 2,2chloroblphenyls. Nelson (1974) reported that PCBe are effective Inhibitors of the binding of ^H-eatradlol to the ret uterus cytosol fraction In vitro. Recent data showed that PCB feeding caused neither significant chromosomal damage nor arrest in the rate of spermatogenesis in male rats (Dlkshith at al., (1975). Flick at al., (1945) noted enlarged adrenals and small spleana in PCB-ereated white leghorn cockerels. Wassermann and Wsssermann (1972) and Wassermann at al., (1973) reported that rats recslvlng 200 ppm PCBs showed an increase in plasma corticosterone levels. HOMS 2119JO 82 These changes wars concimitant with morphological changes in the zona fasciculate of the adrenal gland which indicated increased activity. Sanders, et al., (1974) have shown similar increases in serum corticosterone in mice fed 60 to 1000 ppm PCBs. It is known that DDT decreases the utilization of dietary carotene and liver storage of vitamin A in rats and cattle (Phillips 1963; Phillips, and Hidiroglou, 1965). Villeneuve et al., 1971) reported that Aroclor 1254 but not Aroclor 1221 reduced the liver concentration of vitamin A in pregnant rabbits. Decreased liver vitamin A concentrations were found in male and female rats and also Japanese quail after PCB feeding (Cecil et al,, 1973). How ever, the laying female quail did not appear to be affected. The effects of PCBs on vitamin E have bees studied by Combs et al., (1975). Their results indicated that dietary PCBs increased the incidence of exudative diathesis in chicks and that this Increase can be overcome by increasing dietary vitamin E or selenium. The vitamin D mediated calclta metabolism was found to be altered by the oral administration of PCBs to chickens (Wong et al., 1974). It. The Comparative Biochemical Toxicity of Aroclor 1254 and Piremaster BP-6 PCB's have been reported to affect e large maber of bio chemical systems. To the extent that it has been reported, PBBs MONS 2U*M 83 affect aoat of these same systems in a similar fashion. However, quantitative and possibly qualitative differancas do not appaar to exist in the response of soma biological parameters. These include thyroid hyperplasia (Norris at al., 1973) and microsomal enayae induction (Farbar and Baker, 1970). A preliminary study was conducted to compare the relative effect of Arodor 1234 (Aro) with Firemaater BP-6 (FM) (Garthoff at al., 1975). This was set up as a broad survey to give an estimate of the relative biological activity of these chemicals in may different systems and pin point areas where further more definitive work should be done. Table 1 svamarlsse the time sequence of the various parameters studied. Adult male rats were fed either 0, 5, 50, or 500 ppm Aroclor 1254 or Flremaster BP-6, mixed in the diet, for 2, 3, or 5 weeks. Due to the large number of parameters examined and the time restraints placed on the study, the experimental design did not permit a totally unambiguous comparison between all of the effects caused by Aro and PM. Soma apparent differences in biochemical responses determined at sacrifice may have been partly due to the fact that, for the three-week study, rats were exposed to FM for 40 hours longer than rats were exposed to Aro. Although it is felt that-under the experimental conditions, this effect should be small, critical coapsrlson of Aro and FM toxicity should be 84 MONS 211*32 considered centetive until smaller experiments ere conducted which esa eliminate this difference in exposure time. The following evaluations were made based un the assumption that this two day differencs had no effect. Some parameters showed no consistent or doee related change with either chemical at any level. These included relative kidney weight and testes weight, hematology, SGPT, SCOT, plasma BUM, plasma corticosterone, the rate of liver protein synthesis per g tissue, liver dry weight, lyophlllsable kidney, liver, or testes, the incidence of chromosome abnormalities and the number of cells in mitosis from bone marrow of spermatogemlal cells. Growth and adipose tissue weight ware equally depressed by Aro and FM when fed over a three week period at 500 ppm. The growth effect appeared to be due primarily to decreased food efficiency. Three parameters were more sensitive to Aro than to Pm. These included the decreased liver protein and liver MU per g tissue and the decreased plasma glucose. Ob the other hand, six parameters were more sensitive to PM. These Included liver growth, increased liver lipid (total lipid, cholesterol, phospholipid, and neutral lipid), elevated plasma cholesterol. Increased microsomal ensyme activities, impaired mitochondrial function, and decreased liver MU synthesis. HONS 211933 85 Disruptions of ths redox and energy ststss of ths csll vsrs producsd by dietary exposure to Aro or FM. Thsss sffsct* were lndlcstsd by studios wit isolstod mitochondria snd frozen clamped liver. In the latter case the concentrations of adenine nucleotides, pyridine nucleotides, and glycolytic intermediates were determined. On a molar basis, hexabromoblphenyl was reported to be about five times more potent than Arochior 1254 in causing soma increased microsomal ensyme activities (Farber and Baker, 1974). A three-fold difference was observed between Flremaster BP-6 and Arochior 1254 (Table 3). A pathology team autopsled rats after both three and five weeks exposure to Aro and FM (Kasza et al., 1975). Pathological analysis of the liver of rata fed Aro or FM for three weeks showed no abnormalities on gross observation. No histopathologlcal lesions were seen in controls, Aro groups, or the 5 and 50 ppm FM groups, after three weeks. In the 500 ppm FM group, minimal vacuolstion and focal hepatitis were each seen in one rat. After five weeks exposure, gross observation of the liver indicated an increasing incidence of enlargement, friability, and prominent lobular pattern with Increasing dose of both chemicals. Micro scopically the 5 ppb Aro group appeared normal while 2/7 rats fed 5 ppm FM had cytoplasmic degeneration. All rats had this lesion at 50 and 500 ppm Aro or FM, with increasing cellulsr lipid. This lesion was recent, centrllobular and coalescing with no change in fibrous tissue or glycogen. The histopsthology was qualitatively similar but slightly more severe in rats fed FM HONS 211934 86 than in choc* fad Aro and ia consistent with biochemical findings. Ths biochemical findings wsrs all made after three weeks exposure at which tiae histopathologlcal changes were not evident. Even at five weeks the histopathology was not severe enough to alter most of the clinical chemistry in the rat. In most parameters evaluated, FM either caused an effect at a lower doae level or caused an effect In a shorter time or caused a greater effect at the same doae level, expressed as ppm in the diet, as Aro. When expressed on a molar basis FM becomes even more effective than PCS. Exceptions to this observation veie noted at the 500 ppm doae level where differential effects of Aro and Fm were often eliminated. A sumary of the major effects seen are found in Tables 2, 3, and 4, In all but one case the response of a parameter occurred et the highest doae level. Nlcrosomel enayme activities were, in genersl, greater et the 50 ppm level and the decline at 500 ppm relative to the 50 ppm level was more marked with FM. IZZ. Metabolism of PCB and PBB Mixture Generally, the tissues from animals and man containing PCB from environmental exposure have GLC patterns resembling those of PCB mixtures with more than 50Z chlorination. This ia in marked contrast to the major manufactured products that generally contain 422 or leas chlorine. While one cannot rule out the-poaaiblity that differential uses favored introduction of more highly *0NS 211*35 87 TABLE 1 parameters evaluated in study EXPOSURE TIME (WEEKS) TWO THREE FIVE PARAMETERS Body weight XXX Liver weight XXX Other tiseue weight* X Mltochondrlel reepiretlon X Liver composition X Liver protein eyntheele X Liver RNA synthesis X Liver Microsomal enxyme activities X Liver intermediary metabolites X Serum cholesterol X Serum enxymss XX Plasms cortlcostsrone X Plasma protela X BUN XX Gross pathology XX Hlstopathology XX Cytogenetic analysis X HONS 211*36 TABU 2 RELATIVE BIOCHEMICAL RESPONSE TO Aro AND PM AFTER TWO WEEKS DIETARY EXPOSURE PARAMETERS Liver vsltht Body vsifht Mitochondrial respiration LEL1 ppa Aro PM 500 50 500 500 MaxiMS Effect X Cootrol Aro PM 170 183 152 192 lowest effective level thet showed s doee-rsspoose relationship HONS 21193? TABLE 3 RELATIVE BIOCHEMICAL RESPONSE TO Aro AND FM AFTER THREE WEEKS DIETARY EXPOSURE LEI3- ppn Aro FM Maxinun Efface X Control Aro FM PARAMETERS Body walght gain SO 500 52 52 Livar waitht Body walght Livar dry vt. SO NE3 50 50 203 214 NE3 107 Livar lipid SO 5 138 156 Livar eholaatarol 500 Livar protaln 500 Livar RKA 5 Livar DMA S 50 NZ 50 5 226 232 84 NE 79 84 58 60 Livar RKA anythaala 500 5 78 86 MONS 211936 TABLE 3 (Cont'd) LEL2 ppa Aro FM PARAMETERS pera-Nltrobenaoate reductase activity 50 Plaeaa glucose 50 5 NE3 Please cholesterol 500 5 Hulna Effect X Control Aro FM 2882 5202 79 NE3 178 256 ^Lowest effective level thet shove e dose response reletionahlp. 2ttaxlauB effect occurred et 50 ppa dose, ^ - Mo effect MONS 211939 TABLE 4 RESPONSE OF RELATIVE LIVER WEIGHT TO Aro AND FM AFTER FIVE WEEKS DIETARY EXPOSURE* PARAMETER Liver veieht body weight LEL ppw Aro FM 500 50 Mejcimua Effect X Control Aro FM 171 216 Legend - see Teble 3. HONS 2i1940 chlorinated material* In the environment, the observation ha* led to the general belief that the leas chlorinated components are mors readily metabolized. The environmental observations have been confirmed in a large ntmber of experimental feeding studies in s number of maaulian and avian species. The abseacs or diminished concentration of the early eluting peaks have been reported in rsts (Grant at al., 1971a, Curley et al., 1971), rabbit (Grant at al., 1971b), cow (Fries et al., 1973a), quail (Bailey and Bunyan, 1972), and hens (Fries et si., 1973b). The peaks not present in tissues and products generally are the early eluting peaks that correspond to the PCBs with lower degree* of chlorination. The observations are conalstgnt with the belief that the rate of metabolic attack on PCBs decreases with Increasing chlorination. Studies of the single PCB homologs of various degrees of chlorination have shown that those with five or leas chlorine atone are more radily metabolized and excreted than the PCBs with higher chlorination (Berlin et al., 1975; Hutzinger et al., 1972; Helves and Brandt, 1973). The position of chlorine substitution also effects the retention and elimina tion of single homologs (de Freitas sad Norstrom, 1974) . Metabolism of Individual PCBs will be discussed more thoroughly in another section. MONS 211941 93 Among tha various food species thara appamra to ba oaa azcapclon to tha general rula that tha laaa chlorlnatad PCBa ara hydroxylated. Tha PCB raaiduaa In flah cloaaly resemble tha PCB to which tha flah vaa expoaad. Thara waa llttla change In tha ralatlva concantratlona of tha varioua homologs (Stalling and Mayar, 1972). Conalatant with thla obaarvatlon It haa baaa found that mono-, dl-, and tetrachloroblphanyla ara not metabolized by trout (Hutzlngar at al, 1972). Among tha food producing anlmala (cattla, chickens) concantra tlona of PCB raaiduaa in milk and agga raach a "*toady stata" aftar approxloataly 8 to 12 waaka of continuoua intaka (Prlaa at al., 1973a; Prlaa at al., 1973b). Tha ratio of raaldua lavala In food producta to lavala In tha animal*a dlat ara graataat with PCBa of 54Z chlorination. Tha raaldua lavala In milk ara approximately 4 tlmaa tha raaldua laval In tha cov*a dlat. Zn hana( raaldua lavala In agga ara approximately equal to tha dlat while raaldua laval in tha body tlaaua (fat baala) la approxlmataly 6 tlmaa tha dietary laval. Tha ratio of raaldua laval in product and tlaauaa to Intaka la lower with PCBa of graatar than 54Z chlorina tion. Slnca It la unlikely that thara la greater metabolism of tha moat highly chlorinated PCBa, tha raaulta auggaat that tha absorption for tha Cl tract la lower for tha more highly chlorlna tad PCBa (Smith at al., 1978). PCBa of four or fewer chlorines rarely occur In products whan tha anlmala ara fad environmen tally realistic lavala. MOHS 211942 94 Much lui is known about the metabolism of PBB than PCBa. How ever, the observation* that hava been reported tend to be consistent with the reporte on PCB metabolism. While the PBB of most lanadiate concern for human health (BP-6, Michigan Chemical Co.) is a multicomponent mixture like the PCBs, it is much simpler than the ordinary coMarclal PCB mixtures. The major component is a single hexabromobiphenyl accounting for over 60Z of the total material. The only other component occurring in significant amounts Is a single heptabromoblphenyl. In general, the measures of retention in edible tissues and excretion in edible products are similar for the hexabromobiphenyl and the hexschloroblphanyla (Pries at al., 1976; Pries and Marrow, 1975). Heptabromoblphenyl retention and excretion is only 1/10 of that of the hexabromobiphenyl. In cattle that have been heavily contaminated with BP-6, the heptabromoblphenyl was barely detec table (milk or tissues) 6 to 8 months after the time of Ingestion even though the hexabromobiphenyl level was over 1000 ppm (Pries at al., 1975). Since the heptabromoblphenyl le not more likely to be metabollxmd by the animal then the hexabromobiphenyl, it is reasonable to sssias that Its absorption is lower or its elimination through bile is greater than the hexabromobiphenyl component. Over 60Z of octabromobiphenyl is excreted in the fecea as the perent compound when it la fed to rats (Norris at al., 1974). This observation supports the conclusion that there Is less absorption with increasing halogenatlon. MONS 211943 95 IV. The Metabolism of Individual PCBs yuantitative studies of the distribution of FCBs indicate that most. If not all, FCBa which contain six or fawar chlorine atoms are efficiently abosrbad from the gut of higher animals (Albro and Fishbeln, 1972; Matthews and Anderson, 1975a; Van Miller et al., 1975). Furthermore, only about one-tenth of the absorbed dose Is excreted prior to metabolism to more polar compounds (Matthews and Anderson, 1975a; Van Miller et el., 1975; Balln et al., 1975). A number of the less chlorinated PCBs (l.e., mono-, di-, tri-, and tetrachloroblphenyls) are readily metabolized and excreted by birds and manals. There is little evidence that fish can metabo lize any PCD. It has bean adequately demonstrated that a number of manalian species and the pigeon can readily degrade and excrete monochloroblphenyls, primarily 4-chloroblphenyl (Matthews and Anderson, 1975a; Block and Cornish, 1959; Butzlnger at al., 1972; Safe et al., 1975a,b,c). However, the rate of metabolism drops sharply as the number of chlorines per molecule increases. For example, the rate of metabolism and excretion of several PCB isomers hee been shown to decrease as the number of chlorines Increases In the following order: 4-chloroblphenyl 4,4'-dichloroblphenyl 4,4'-dlchlorobiphenyl 2,5,2',5'-tetrachloroblphenyl 2,4,5, 2*,5'-pentachlorobiphenyl (Matthews and Anderson, 1975a; Van Miller et al., 1975; Matthews and Anderson, 1975b). The rate MO NS 211944 96 of Mtabolln *od excretion of 4,4'-dichlorobiphenyl 1* approxi mately one-half that of 4-chlorobiphenyl. in each casa th* PCBs Aro metabolized primarily by hydroxyletion And conjugation with glucluronic ACid (Matthews And Anderson, 1975a; BAlin et al., 1975). Tha importanca of cha routa of axcration, i.a., urina or feces, ia datarminad primarily by Cha dagraa of chlorination. Approximately 60, 30, and 7 parcant of a aingla doaa of a mono*, a di-, and a pantachlorobiphanyl, raapactivaly, wara axerated in Cha urina of rata. Tha remainder of Cha doaa waa axcratad in the fecaa (Matthew* and Anderson, 1975a). Once tha number of chlorine atone on tha biphenyl molecule raachea four, tha position of the chlorine atone becomes a more important factor in determining the rata of metabolism and excre tion (Matthew* and Anderson, 1975s; Da Freitas and Noretrom, 1974), As was observed with the chlorinated benzenes (Jondorf at al., 1955; Jondorf at al., 1959) those PCBs which have two adjacent unaubstltuted carbon atone era metabolized and axcratad more rapidly than PCBs having the same degree of chlorination, but which do not have two edjscent unsubstituted carbon atone (Matthews sad Anderson, 1975s). It is also likely that, as with the chlorinated benzenes, the absence of two adjacent unaubstltuted carbon atone has a greater affect as the dagraa of chlorination increases. For example, Che initial half-life of 3,5,5*,5*-tetrachloroblphenyl is only about two-fold greater M0N5 211945 in the rat (Matthews and Tuay, 1976) than that of 2,5,2',5'tatrachlorobiphany1 In tha saaa spaciaa (Van Millar at ml., 1975). On tha othar hand, tha initial half-Ufa of 2,4,5,2' ,4' ,5'-hexachlorobiphanyl appaars to ba infinite, vharaas tha Initial halflifa of 2,4,5,2',5'-pantachlorobiphanyl la only about two days (Matthews and Anderson, 1975a). This particular hexachlorobIphany1 has bean shown to ba in tha highast concentration of any PCB found in tha adipose tissue of tha Swedish population (Jansen at al., 1974). Tha importance of two adjacent unsubstituted carbon atoma to tha metabolism of PCBa nay ba derived fron Chair affect on the formation of arena oxide intermediates during tha metabolism of these compounds by tha hepatic mixed-function oxidases. Evidence for the formation of arena oxide Intermediates in PCI metabolism was first provided by Gardner at al., (Gardner at al., 1973) In their study of tha metabolism of 2,5,2',5'-tetrachloroblphenyl and In a later study of 4-chloroblphenyl metabolism by Safa at al., (1975c) which was specifically designed to provide evidence of an arena oxide Intermediate. Metabolism of PCBs which do not have two adjacent unsubstituted carbon atone by hepatic mixed-function oxidases, via direct hydroxylatlon of Che biphenyl molecule and/or the formation of an arena oxide between a chlorinated and an msubstltuted carbon atom could occur, but each of these mechanisms would ba expected to be slower than arena oxide forma tion between two unsubstituted carbon atone (Daly at al., 1972; Jerlna and Daly, 1974). Tha latter mechanism would be expected HONS 211946 98 to result In dechlorination and/or shifting of a chlorlna atom (Daly at al., 1972). Tha matabollsm of salactad hexachloroblphsnyls haa baan raportad (Jansen and Sundstrom, 1974; Hutzlngar at al., 1974; Hass, unpubllshad), and In tvo Incldants (Hutzlngar at al., 1974; McKinney at al., unpubllshad) dachlorlnatlon and/or chlorine shifting waa observed, but In all casaa tha rata of matabollsm was quits alow. Thus, thoaa PCBs which ara more readily metabolized and excreted may also more readily form arena oxides. Certain arena oxides have bean implicated sa potential carcinogens (Daly at al., 1972; Jerlna and Daly, 1974) as have certain PCB formulations (Ito at al., 1973; Kimbrough and Linder, 1973; Kimbrough et al., 1975), but there la no published evidence which indicates that tha less chlorinated PCB formulations have been ao rigorously tested. HONS aim7 99 animal toxicology Introduction A number of reviews on different aspects of the toxicology of the PCBs have been written recently end no effort has been made in this report to give a complete outline of the animal toxicology (Kimbrough. 1974; Peakall snd Rlsebrough, 1975; Nelson et si.. 1972; Fishbein. 1974). The material summarized in this report consists predominantly of more recently published information on toxic effects of PCBs in maimala and experimental studies in birda. Since Nelson et al. (1972) discussed the toxic effects produced in fish by PCBs extensively, the reader la referred to this and the other reviews cited above for information on toxicity of PCBs in this species. The emphasis of the present review was placed on chronic toxic effects wherever possible. Unfortunately, very little information was available for halogenated dlbenzofurana and the bromlnated biphenyl mixtures. Acute Toxicity Polychlorinated Biphenyls (PCB) -The acute oral LDjq of the PCBs in rats, rabbits and mice ranges from 1 to 10 g/kg body weight (Tables 5,6). There is some indication that young animals may be more sensitive than adults. MONS 211**8 100 and famalas mors aanaltlva chan malaa (Tabla 5). According to tha claaalflcation uaad by tha American Industrial Hyglane Association (Hodga 6 Starnar, 1949), tha acuta toxicity of tha PCBs can bs classsd as slightly toxic (0.5 to 5 g/kg body waight) to practically non-toxic (5-15 g/kg body walght). Tha symptoms assocatlatad with administration of a toxic dosa of PCB (Aroclor 1242) In rats consist of dlarrhaa, dInin1shad axploratory bshsvior, dacraasad rasponsa to pain stinull, chronodacryarrhaa, adipsla, oliguria, anoraxia, arythama of limbs, followed by ataxia, cons and daath. Daath occurrad up to 14 days following administration of tha toxic dosa. 24 hours aftar administration of a slngla toxic dosa of Aroclor 1242, gross pathology indicstad all organs sppaarad normal axcapt tha livar and kldnays. No inflammation was obaarvad In tha abdominal and intastinal mucosa. Histopathological findings showad larga discrsta sudaaophilic vacuolas In tha hapatocytas. Wtdaly scattarad foci of tubular apithalisl calls wars prasant in tha kldnays (Brucknar, at al., 1973). Hovavar, Kimbrough (1974) raportad ulcarstion of tha gastric and duodsnal mucosa In rats sftar slngla oral dosaa of 3,000 mg/kg Aroclor 1254 or 1260. Only oma matabolita of PCB la known to hava baan tastad for tha scnta U>5q. Tha 5-OH matabolita of 2,4.3*4'-tetrschlorobiphanyl was known to ba mors toxic than tha parant compound (0.4-3 g/kgBU/2.15 g/kgBW (Tabla 5). Darmal toxicity of PCB is summarised in Tabla 7. HONS 211949 101 Po1ybrominated Biphenyls (PBBs) Polybrominated biphenyls show a low order of acuce toxicity to the rat (Table 8). In the case of chicks, a 37X mortality was reported when PBB was administered in the diet at a level of 400 ppm, for 15 days. Seven out of eight guinea pigs fed 50 ppm dietary PBB died (Vos and VanGerderen, 1973). fffacts noted Included cortical atrophy of the thymus, adrenal enlarge ment. marked depletion of the follicles and periarteriolar lymphocytes shesth of the spleen. Chlorinated Dlbensofurans See subacute toxicity. Chlorinated Wapthalenes See end of chapter. HONS 211950 T.MLZ 5 CoooOuftd Toiled Si>*el tout* |/C b*d-v wc. LO.is Mfaranc* Araclor 1254 " 1260 " USA H 1240 " 12 54 " 1221 H ia|2 " 1240 ` " 1254 a 1254 " Ui4 '* 1254 AjnaClor-400 KAtl4Clor*400 KiAielar-400 Kjajc1[*400 JCAnoclor-300 K*ntcior*100 If`200 (bU>HMyl of jlCLorlda and fcolON 2,^' -dicrilorolipfeonyl Cf icnloro61ph<6)f 1 bi^uvnyl oJ trichloride and boltw 2,4,3', 4' -cocracir.lorodlphMyl 5-ON doriMtivo of I,**}'*'- cAtrachiorooipheorl 2,3,4,3 * 4 ' -pMAachlorotipJtaayl ILat(Aduit-Sh*rm*n Strain) lUe (Adult-ShorOAA ScrlA> lC {)M|t7 Iftft *' ") l*i(w#jnllrti hC{f4B4l* btlfiuii " bcffiMl* "> "' ) ) ") !( lot (WLalar-JQ-dr-old N-F) fcaefVUtar-40-d4y-old H-n l*c(Wlie*r*120-d*f-old N) l*t(Ul*e*f*120-d*f-old F) -Cfwtcf Strain ' Ptela) BotUttotar Strain - Fm*41o) Rlc(CFl Strain PtoUl He(CFl Strati FomIo) l4t(4ij[r Strain Ril<) Le(Ul*tf Strain * null) MlC*<dd~tT4A-f*MU) oral oral orl oral L*tr*v*4ua oral oral oral oral oral oral oral oral oral oral oral oral oral oral Hie* (dd~ac rain- foaolo) 7tU(dd-'-acrain-f*Mlo) M'.co(dd-atraia-(oa*l} oral oral oral Hlc*(Cf1 strain) Hlcfttcri itnla) loetaporlcoMal " Klco(CFI *trt<) 4-10 4-10 1.245 1.315 0, 331 4 06 11.3 4.25 l.) 1.4 2.0 2.5 1.30 (!/!*> 1.14 <ol/l*} 1.175 (ml/tt) l.57 (Bl/Kf 1.15 1.05 1.31 Under. t 1 fJ9741 TT* Panoi on Haxardoua Traca Subtcantaa Iruckaor. e al. fi9?3) Craat 4 rhiUipa U974) Ikodo, *t al. (1970) lltoaur* (1972) 7.14 3.04-4.73 3.27 2.13 0.43 Yaaaooco 4 Yoahlo^ra (1973) 0.43 HONS 211951 TABLE 6 Or*l_JLpjQ (1UC) (Pnl on H*trdou SubCncti. 19721 Conpound Tttttd Aroclor 1221 Aroclor 1232 Aroclor 1242 Aroclor 1248 Aroclor 1260 Aroclor 1262 Aroclor 1268 (undlluCtd) (undlluctd) (undlluctd) (undlluctd) (SOX aoln In corn oil) (BOX soln In corn oil) (SOX coin in corn oil) g/Kg body wt. 3.93 4.47 8.65 11.3 10.3 11.3 10.9 mm 2 HONS 11952 TABU 7 Skin HLD<n Rebblte (Panel on Hererdoue Subecancel. 1972) Conpound tented Aroclor 1221 Acoclor 1232 Aroclor 1242 Aroclor 1248 Aroclor 1260 Aroclor 1262 Aroclor 1268 (utdlluted) (undiluted) (undiluted) (undiluted) (501 solo in corn oil) (50Z eoln In corn oil) (33.3X eoln in corn oil) t/Kg body wc. >2.000 >1.26 > .79* > .794 >1.26 >1.26 >2.5 <3.169 <2.0 <1.269 <1.269 <2.0 <3.16 HONS 211953 Coaoound Flraaaacar IPS Oc c*brMDblph*ay 1 flceabr twblphany 1 Maxabroaob lpbany 1 Octtbroabblpl.anyL SpQClot lUc h( Sobvhlca Quail babble babble TJLELE 9 LDgii Polvbroolnafd tlohonyla Roue* Oral Oral Oral Danal Danal tjo 21. J */M Hot* Chao 2 l/Kb Hot* eban 12.5 IIU Hot* ebaa J i/M Hot* Cbao 10 b/M R*fr*nc Kill Top Ri**rcK, Inc. II9T0) Michigan Ch*icl Cap4ny Aftooalt, it *1., 19?2 HONS 2L1954 SUB-ACUTE AND REPRODUCTIVE EFFECTS OF PCB'S, PBB'S AND CHLORINATED DIBENZOFURANS a. Mammalian Sub-acute Effaces of PCBa In contrast to the acuta toxicity (oral and dermal) of the PCBa which la of relatively low order when the aubataacaa are adminlatered aa a alngle doae, aapecta of aub-acute toxicity of both the PCBa and individual chlorinated blphenyla appear to be of far greeter concern exhibiting apeclaa aenaltlvity and cumulative toxic affacta following continuoua expoaure at low levela. When groupa of mice were given toxic rice bran oil containing 1,600 ppm PCBa or 5000 ppm of PCB mixture with ASX chlorine, the toxicity reaponae waa ateller in the two groupa (Nlahlzumi 1970). In a group of rata fed 2000 ppm of Phenochlor DP-6, deatha occurred between the 12th and 26th daya with enlarged llvera, atrophy of the apleena and a progreaaively Induced hepatic porphyria waa noted (Voa and Koeaen, 1970). Administration of 1000 ppm of Arochlor 1254 In the diet of rata reaulted in deatha between the 28th day and 53rd day (100X mortality) of feeding (Tucker and Crabtree 1970), SOX mortality at the name doeege in an 8-month atudy waa reported by Kimbrough at al. (1972). MOMS 211955 107 Two groups of 52 male Sherman strain weanling rats were fed 100 ppm Aroclor 1242 (6.6-3.89 mg Aroclor/kg body weight) and 100 ppm Aroclor 1016 (6.9-3.5 mg Aroclor/kg body weight) respectively for 6 months and 54 rats were fed ground chow. Groups of 4 rats were sacrificed at intervals. Microscopic examination of the liver showed evidence of lipid accumulation, enlarged liver cells, inclusions in a number of livers and hemorrhage and necrosis in one animal per experimental group each. No difference in effect between the two compounds was noted (Burse et al., 1974). Rats given 100 mg Aroclor 1242/kg body weight by stomach tube every other day for 3 weeks showed hlstopathologlcal changes only in the liver and kidneys as foci of sudanophllic vacuoletion but no overt sign of toxicity (Bruckner et al., 1973). Rats (Sprague-Dawley) given 100 ppm of 2,5,2', S'-tetrachloroblphenyl in the diet for 3 weeks showed less liver hypertrophy and fewer biochemicel changes than rats given a similar dose of Aroclor 1248 (Allen et si., 1975). Administration of 100 mg/kg body weight of Aroclors 1242, or 1254 orally to rabbits once a week for 14 weeks resulted in enlarged livers with epperent destruction of the rough endoplasmic reticulum and- atrophy of the uteri, while Aroclor 1221 did not cause this effect. (Koller and Zinkl, 1973). HONS 21lt$6 108 Th unusual sensitivity of mink to PCBs was reported by Aulerlch et al., (1973) who noted 100Z mortality within 6 months following administration of 30 ppm PCB (10 ppm each of Aroclor 1262, 1248 and 1254} to adult mink. Feeding of Aroclor 1256 at levels of 5 and 10 ppm in the diet for 4 months led to a dosedependent retardation of weight gain of growing female mink. Platonow and Karstad (1973) produced 100Z mortality in mink when they fed this species 3.6 ppm Aroclor 1254 in the diet for 105 days. Rhesus monkeys are also very sensitive to PCBs (McNulty, 1975)For example, adult female rhesus monksys (Macaca Mulatta) have been fed diets containing 2.5. 5.0 and 25.0 ppm of Aroclor 1248 over periods ranging from 2 months for the higher PCB levels to 1 year for the two lower doses (Allen et al.t 1974; Bersottl and Allen, 1975). Animals on the 25 ppm dose developed facial edema, alopecia and acne within 1 month, and 1 of 6 animals had expired as a result of PCB Intoxication 2 months after removal from the diet. The total Intake of PCBs ranged from 250 to 400 mg/animal. As was the case with the higher doses, these animals developed anemia, hypoprotelammla, bone marrow atrophy and severe hypertrophic hyperplastic gastritis. PCB concentrations in subcutaneous adipose tissue averaged 127 pp* at termination of experimental diet and 34 ppm 8 months later. Surviving animals continued to show clinical signs and. lesions of PCB intoxication 2 years following PCB exposure (Allen, 1975). MQNS 211957 109 Whan adult rhesus monkeys were fed diets containing 2.5 and 5.0 ppm Aroclor 1248 for 1 year, the females of the group developed periorbital edema, alopecia, erythema and acne from lesions that involved the face and neck within 1 to 2 months (Allen et al., 1974; Allen, 1975). Although males consumed more PCB, they exhibited only moderate periorbital edema and erythema. An abnormal dysplastlc growth pattern of the gastric mucoaa was also noted In male rhesus monkeys fed 300 ppm Aroclor 1248 for 3 months (Allen and Norback, 1973). Cygomegalous or squirrel monkeys fed different dietary levels of Kanechlor 400 showed weight loss, palpebral edema, enlargement of the liver and pneumonia. The lowest effective total dose was 300 mg/kg bv given over 20-32 weeks (Nlshlsuml, 1970). A rhesus monkey fed 3 ppm Aroclor 1242 for 8 months died spontaneously and on autopsy was found to possess a gastric lesion manifested by extensive down growth of the epithelium deep into the subserosa and into the muscular wall of the sto mach itself (HcMulty, 1975). One monkey fed 2,4,4'-trichloroblphenyl and another fed 2,4,5,4'5,-pentachloroblphenyl at 10 ppm for 90 days were reported clinically well and autopsies showed no positive findings (Oregon State.University, 1975; McNulty, 1975). 21l95g 110 Third-litter sows received feed containing Aroclor 1242 at a concentration of 20 ppm throughout gestation and nursing. Trastad sows differed significantly from control sows In the number of live pigs farrowed. Treated sowe alao had more muonlfled fetuses. Performance of live-born pigs was not affected by feeding PCB to the sows during nursing. Pathologic changes In the sows included hypertrophy of the liver and erosion of the stomach. Slight atrophy of the spleen and thyroid gland were observed in pigs from treated sows (Hansen at al., 1975). Dermal toxicity studies in rabbits of techinial PCB samples that contain an average of 60S chlorine (Phenoclor DP6, Clophen A60, and Aroclor 1260) as well as fractions containing tetra and pentachlorodlbensofuran have been recently described by Vos and Beene (1971). PCB-induced skin lesions were hyperplasia and hyperkeratosis of the epidermal and follicular epithelium fol lowing application of 11B mg of the three PCBa (5 times per week for 3B days) in the back skin of adult female New Zealand rabbits. Hlstopathology of the liver Included centrolobular degeneration, centrolobular liver cell atrophy, focal necrosis, and cytoplasmic hyalin degeneration. PCB-induced kidney lesions were hydropic degeneration of the convoluted tubules and tubular dilation with the presence of casts. Definitive hyperplasia ana hyperkeratosis of the follicular epithelium of the ear skin were seen after the topical application of fractions of Phenoclor and Clophan, while the fraction from Aroclor caused a minimal hyperplasia and hyper- HONS 211959 111 keratosis of the follicular epithelium. Other efface* elicited by the dermal application of the PCBa including thymua atrophy and lymphopenia as well aa elevated excretion of fecal copro porphyria and protoporphyria. From the raaponae of the back akin and the liver of the rabbit to the three PCB mixture*, and from the reaponaa of che ear to the 25X diethyl ether-hexane fraction* it waa concluded that there were definite quantitative difference* in toxicity, at laaat batween the aamplea used in the above study and prior studies. The extent to which these samples era representative of the normal commercial output has not been established and emphasis** the difficulty in the evaluation of toxicity data of PCBa In which the samples may differ in the amount and nature of toxic impurities. Vos & Notenboom-Ram (1972) compared tha toxicity of Aroclor 1260 with a single isomer 2,4,5,2',4',S'-haxachlorobiphenyl in New Zealand rabbits. Dermal applications of a total 120 mg Aroclor 1260 (5 times/week for 28 days) resulted in early macro scopic skin lesions. The lesions in a 2,4,5,2*,4',5'-hexachlorobiphenyl group of rabbits treated similarly appeared later and were less severe. When groups of 5 male mica wars fed dietary levels of 10, 30, 100 and 300 ppm of the PCB isomers 3,4,5,3',4',5'-hexaehloroblphenyl, 2,4,5,2' ,4',5'-hex*chlorobiphenyl and Z^.b.l'^'.b'-hexachlorobiphenyl HONS 211960 112 for 4 vtcki it was noted that the 3,4,5,3',4',5', isomer of hexachlorobiphanyl caused death in mice at a dietary intake of 10 ppm (2*1 mg/kg bv) within 36-47 days of exposure while consumption of the other two isomers resulted only in deach at a dietary level of 300 ppm (64.3 mg/kg bw). Body weight was reduced at the lowest dose for the 3,4,5,3',4',5', isomer and only at the highest dose for the other two Isomers. Atrophy of the thymus and hepatomegaly was noted at the lowest dose in the mice receiving the 3,4,5,3',4',5', isomer and only this isomer produced experimental porphyria. B. Avian subacute toxic effect of PCBa. Chickens fed 10 ppn Aroclor 1242, or 100 ppm Aroclor 1254 showed diminished growth but e dietary level of 100 ppm Aroclor 1260 did not produce this effect (Kaplinger, et el., 1971). Sy^>toaa of FCB poisoning in birds consist of tremor, ataxia, ruffling, lose of feathers. Edema of the subcutaneous tissues, and fluid acctaulation of the abdominal end thoraxic cavities are characteristic signs at autopsy. These findings are responsible for this diseese being designated chick edema disease. Several outbreaks of chick edema disease caused by PCBe have been reported (Firestone 1973, Kohanowa 1969a, 1969b). Chicks (one day old white leghorn cockerels) ware fed the following FCB isomers; 2,3,6,2',3',6'- haxachlorobiphenyl 2,4,6,2',4*,6'.haxachlorobiphenyl, 2,3,4,2'3',4hexachloroblpheny1, MOWS 211961 113 2,3,6,2',3*,6*, haxachloroblphenyl and 2,4,5,2',4*,5' hexachlorobiphenyl for 21 days at a level of 400 ppm. The Isomer 2,4.5,2',4',5' haxachloroblphenyl was also fed at che dietary level of 100 ppm. The 3,4,5,3',4',5' Isomer was the most toxic causing death of all chicks within 11 days after onset of exposure. These chicks had chick edema disease, atrophy of the thymus and loss of subcutaneous and visceral adipose tissue. The other isomers also caused weight loss the 2,4,6,2*,4',6' isomer being the least effective. Liver enlargement was noted for all groups the 2,3,6,2',3',6' being the least affective and the 2,4,6,2',4',6* the most effective. The microscopic findings made in the liver were most pronounced in the group fed the 2,4,6,2', 4',6' isomer. {Since the chicks fed the 3,4,5,3*,4',5' isomer died relatively early in the experiment pathological findings in the livers of these chicks are not comparable to those made for the other isomers.) When chicks were fad 5X rice oil which had produced Yusho they developed chick edema in 17 days. Similar symptoms were produced when chicks were fed 400 ppm Kanador 400 (Goto at al., 1969). C. H--p-a Reproductive Effects of PCBs Oral administration of 0.025 mg/day of Clophan A60 in peanut oil to tMtl-mice for 10 weeks lengthened the estrus cycle from 6.6 -2.5 days to 8.7 + 4.3 days (Orberg and Kihlstrom, 1973; Kihlstrom at al., 1973). Females treated analogously for 62 days HONS 211962 ii*. and utid with untreated males exhibited a significant decrease In the Implantation rate from 87. OZ In the controls to 79.5Z. Male castrated mice were given 0.25 gm PCB (Clophen A 60) In peanut oll/day by stomach tube for 28 days and 70 gtg testosterone proplonete dally on days 19-28. The dry weight of the seminal vesclcles after this treatment period was compared to castrated male mice given testosterone but not PCB and was found to be signlflcanlty reduced. Groups of 10 KMR I female mice were Injected subcutaneously with 50 mg/kg body weight of Clophen A 60 In peanut oil or only the vehicle on the day of parturition and thereafter once weekly for 3 weeks. The 209 offspring were bred when reaching sexual maturity by mating them in pairs as follows: 17 pairs of controls, 19 pairs where the males had received Clophen A 60 as a suckling, 23 pairs where the female had received Clophen A 60 as sucklings and 23 pairs were both sexes had received Clophen A 60 as sucklings. The controls had an average of 11.5 offspring per litter while the parents where both nates received PCBs as sucklings had 8.8 offspring per litter. The resorption rste for both groups was the sane but the number of implantations were reduced. Reproduction was not affected where only one mate received the PCB (Klhlstrom at al.,1975). Keplinger at al,,(1972) reported low mating Indices and decreased survival of pups for rats receiving Aroclor 1242 at 100 ppm, and decreased survival of pups receiving Aroclor 1254 HONS 211963 115 at 100 ppm. No reproductive effects were found with Aroclor 1260 at 1..10, or 100 ppm, or with Aroclor 1242 or 1254 at 1 or 10 ppm. These studies suggest that In mammals reproductive effects decrease with Increasing chlorination. Sherman rats were exposed to polychlorinated biphenyls, Aroclor 1254 and Aroclor 1260. Rats exposed to Aroclor 1254 at dietary levels of 20 ppm or more had fewer pups per litter chan the controls In the Fib and F2 generations {Linder et al., 1974). The 100 ppm exposure level of Aroclor 1254 Increased mortality In the Fib offspring and markedly decreased mating performance of the Fib adults. The 500 ppm dietary level of Aroclor 1260 reduced litter size and decreased survival In the FI litters. Dietary levels of 5 ppm Aroclor 1254 and 100 ppm Aroclor 1260 had no effect on reproduction in rats exposed through two generations. Liver weights were increased In 21-day-old FI male weanlings at the 1 ppm level of Aroclor 1254 and In either sex of FI and F2 weanlings at 5 ppm or higher levels of both Aroclor 1254 and 1260. Histological changes In the liver and increased liver weights were observed In sdult rats exposed to the higher levels. Pregnant rets given Aroclor 1254 at the rate of 100 mg/kg/day on days 7-15 of gestation produced grossly normal litters, but only 30.lt of the pups survived to weaning. Reproduction and pup survival were not affected at dosage rates of 50 mg Aroclor 1254/kg bw/day or 100 mg Aroclor 1260/kg bw/day. HONS 211964 116 Rabbits have bean found to be nor* sensitive to PCBs than rata in regard to fatotoxic and reproductive effecta (Vllleneuve et al., 1971). In rabblta. no effecta were reported at doaagea of 6.25 or 10 mg Aroclor 1254 /kg/day, but at doaagea of 12.5 co 50 mg/kg/day, fetotoxlc effecta were noted. Abortions, maternal death and atillborne occurred, but no conalatent akeletal abnor mal It lea were found. Ringer et al., (1972) found that 1 ppm of Aroclor 1254 caused a alight reduction In reproductive aucceaa in mink, but at 5 ppm Aroclor 1254 complete reproductive failure waa noted. In the same study 12 mink were fed a diet containing 30X Coho Salmon from Lake Michigan. None of the mink whelped while 11 of 12 controls whelped. Female adult rheaua monkeys fed Aroclor 1248 at a level of 25 ppm for 2 months noted at beginning of the 5th month experienced either fetal resorption or abortion during their second month of pregnancy (evidenced by regression In uterine size and reestablishment of menstruation (Allen et al., 1974). A third salami carried her fetus to term. This infant's weight was considerably lower than the average rhesus Infant (375g vs 544 + lOlg) and had 25 >ug FC?/g/(ppm) of adrenal and adipose tissue. Adult female rh jus monkeys fed 2.5 and 5.0 ppm Aroclor 1248 were bred to control males following the establishment of a relati''*lv steady tissue level of FCB at 6 months. Following 3 mating, 12.5Z of the 5 ppm group and 37.5X of the 2.5 ppm group was pregnant HONS 211965 117 compared co 9015 in the control group (Allen, 1975). the conception rate of females bred to PCB-fed males was equally as great as that in females bred to control males. D. Mammalian Sub-Acute and Reproductive Effects of PBBs Male Sprague Dawlay rats maintained on diets containing 0. 0.1, 0.01 or 1Z octobromobiphenyl for 30 days did not exhibit overt toxicity during this period. Gross pathological studies revealed enlarged livers and some kidney changes (petechial hemorrhage, enlargement), while histopathologlcal changes in the 1Z group included liver lesions consisting of centrilobular cytoplasmic enlargement and vaculatlon and kidney lesions con sisting of hyaline degenerative cytoplasmic changes (Aftosmis et al., 1972). Thyroid hyperplasia was observed in all rats in all test groups. Rats were fad 100 and 1000 ppm octabromoblphanyl in their diets and then a number were sacrificed after 2 and 4 weeks of feeding and after 2,6 and IS weeks of recovery on an octabromobiphenyl-free diet* The hepatocytes were markedly enlarged in rats fed 100 ppm and 1000 ppm of the PBB after 2 weeks of treatment. Histopathologic changes were characterized by hepatocellular hyper trophy with the pathologic changes localized mostly in the centrslobular zone (Aftomis et al,,1972b). The cytoplasmic inclusions ware more pronounced in the livers of rats fed 1000 ppm than 100 ppm of the PBB. There was some evidence of recovery since the livers of the 100 ppm level animals returned almost to normal 2 weeks after HONS 2119*6 118 withdrawal of the compound. Preliminary 28-day feeding studios in rats with octabroaobiphenyl at 100 sad 1000 ppa demonstrated liver enlargaaant, hepatocellular alterations with cytoplasaic inclusions end eccuauletive affects of broaina in the fat, liver end auscle (Aftosais at el., 1972). Norrie at el., (1974) demonstrated heaatological chengas, liver anlergaaent end liver end kidney lesions et ell levels of octabromobiphenyl (10. 000, 1000 end 100 ppa) in the diet of rets. When three groups of priaagravld rets fed diets containing 100, 1000 or 10,000 ppa of octabroaobipheuyl (BB-8) from day 6 through 15 of pregnancy and the pups delivered on day 20, anasarca was observed in one fetus at each of the two highest levels end geatroschlsis was observed in another fetus at each of these levels. No other gross effects were observed in either the aothers or the pups and dose-related levels of broaina wars found in the fetuses (Aftoais at el., 1972a). Whan 1 g/kg of octobroaobiphenyl was applied in corn oil under occuluded conditions to rabbits for 5 hrs/day for 10 days over a 2-wash period, liver anlargaaent was found while et O.lg/kg no enlargnent was found (Aftosala, et al.. 1972b). Haxabroaobiphaayl (Firaaaater BP-6) fed to pregnant SpragueDewley rets and Swiss/ICB nice (100 and 1000 ppa) resulted in a dose dependant decrease in aaan fecal weight. (Ntaber of pregnant HONS ill**7 119 mice: 16 control*, 9 received 100 ppm and 12 recalved 1000 ppm. Number of pregnant rats: 12 controls, 7 received 100 ppm and 6 received 1000 ppm). Exencephaly was produced In the offspring of the mice that received both 100 (3/121)and 1,000 ppm (2/174 dosages while cleft palate (4/87) and defective kidneys (2/87) were not*d in offspring at the 1,000 ppm level. Nonpregnant mice fed 1,000 ppm PBBs for 11 days showed marked increase in liver size and weight (Corbett et al.t 1975), Cows from the Halbert herd received feed containing PBB at a level of 2914 ppm at a consumption rate of approximately 15 lbs/head/day (200 cows) or 8 lb*/head/day (200 cows) for a period of approximately 16 days. Early signs of toxicity included anorexia, decreased milk production, increased frequency of urination, lacrlaation, soma lameness, and shrinking of the udder of recently freshened cows. Upon removal of the contaminated feed appetite i^roved. Cows bred 4-6 weeks prior to onset came back in heat suggesting early resorptions. Later signs exhibited by the cows included hematomas, abscesses, weight loss particularly in high producing cows, abnormal hoof growth, rough hair coat, alopecia, thickening of skim, dystocia, lack of udder development at freshening, metritis,- and nag igibla milk production in cows freshening 3-4 months post-exposure. Cows also exhibited a lack of wound healing, general weakness and were highly susceptible to stress. Six months post-exposure non-lactating cows showed depression of HONS 211968 -20 mice: 16 controls, 9 received 100 ppm and 12 received 1000 ppm. Number of pregnant rats: 12 controls, 7 received 100 ppm and 6 received 1000 ppm). Exencephaly was produced In the offspring of the mice that received both 100 (3/121)and 1,000 ppm (2/174 dosages while cleft palate (4/87) and defective kidneys (2/87) were notd In offspring at the 1,000 ppm level. Nonpregnant mice fed 1,000 ppm PBBs for 11 days showed marked Increase In liver size and weight (Corbett et al., 1975). Cows from the Halbert herd received feed containing PBB at a level of 2914 ppm at a consumption rate of approximately 15 lbs/head/day (200 cows) or 8 lbs/head/day (200 cows) for a period of approximately 16 days. Early signs of toxicity Included anorexia, decreased milk production. Increased frequency of urination, lacrlmation, some lameness, and shrinking of the udder of recently freshened cows. Upon removal of the contaminated feed appetite i^roved. Cows bred 4-6 weeks prior to onset came back in heat suggesting early resorptions. Later signs exhibited by the cows Included hematomas, abscesses, weight loss particularly In high producing cows, abnormal hoof growth, rough hair coat, alopecia, thickening of skim, dystocia, lack of udder development at freshening, metritis, and neg igible milk production In cows freshening 3-4 months post-exposure. Cows also exhibited a lack of wound healing, general weakness and ware highly susceptible to stress. Six months post-exposure non-lactating cows showed depression of MOHS 211966 -20 appetite, preparti* vukntif, failed co develop signs of lsbor end died without calving. Gross pathology included liver end rensl degeneration, end hemstomes end sbscesses in the peritonesl end thorscic csvities. Twelve 6 to IS month-old heifers end bulls were offered the contsminsted feed sd libitum, five of the snlmsls were deed within 6 weeks. Younger snimsle became prostrate, went through a short period of coma and died. Signs and lesions included testicular atrophy, abdominal adhesions, and massive liver sbscesses. At 5 months only 2 animals remained alive. These became anorexic, would consume only milk and developed signs of hyperkeratosis over the entire body (Jackson and Halbert, 1974; Uelborn, 1975). The health status of 16 herds of dairy cattle exposed to low levels of polybronlnated biphenyl (FBB) was compared with that of 15 control herds. Milk production of the contaminated herds was not significangly changed In 1972, 1973. and 1974, and was not significantly different from that of control herds in the same years. Mortality of adult cows and calves, the percentages of cows culled from the herds because of old age and low production, disease, or sterility, and the general health conditions were similar in the two groups. Scrum concentrations of calcium, glucose, and cholesterol in contaminated herds were significantly different from those of the control herds, but the relationship to PBB exposure needs further investigation (Mercer at si., 1976, In Press). HONS 121 A study consisting of sn Investigation of records and herd history of herds accidentally exposed to PBBs was also done. Significant findings Included a decrease In production of 20 to 50Z; severe weight loss at calving tine; sterility; decreased growth rate in young aniaals; evidence of soreness and stiffness; poor response to therepy of coanon diseases; prolonged wound healing; abnormal hoof growth; calf losses; and malformed calves. (Dealng, 1975). Polybroainatad biphenyl was administered to four cows at a dose rate of 10 ng/haad/day for 60 days. Although the study wss not designed ss a toxicology study, retrospective evaluation of the data provided no evidence of clinical probleaa aaong test aniaals during the period of exposure to PBB nor for s year thereafter (Fries and Harrow, 1975). E. Chlorinated Dibanxofurans - *lian Toxicity It is presently assuaed because of our experience with the chlorinated dlbeasodloxins, that the 2,3,7,8-tatrachlorodibanzofuran la the aoat toxic conpound of this group of chealcsls, while the dlbeaaofuran Itself or the octadibenzofuren have little inherent toxicity on so acuta basis (Kimbrough, 1974), Bauer at al , in 1961 daaonstrstad the toxicity of mixtures of tri and tetrachlorodibenxofursns. A single dose of 1 ag or 0.5 ag/kgbody weight given orally to rabbits caused severe and often fatal liver necrosis. Application of this aatarial to the rabbit HONS 211*70 122 ear caused ssvscs hypscksrstosls at ths application sits. Ths single oral LDjg foe tetrachlorodibenzofuran foe guinea pigs is between 5 and 10 /ug/kg. Symptoms of toxicity weee seveee weight loss, ateophy of the thyaus and splean. Hemorrhages were observed in the adrenals, urinary bladder and single cell necrosis was noted in the liver- C. D. Strain rats given as much as 1000 ug/kg bw of the TCDF failed co show any symptoms of toxicity and microscopic examination of their tissues did not reveal any abnormalities. Similarly oral single doses of up to 6000 ug/kg body weight of TCDF failed to elicit any toxic effect in C57B1 mica. A subcutaneous dose of 6000 ug/kg bw given to these mica produced weight loss, hepatomegaly and atrophy of the thumus but no fatalities (Moore at al., 1976). . While these findings indicate that in mica TCDF is 30 fold less toxic than TCDD, ths overall estimated difference in toxicity between TCDD and TCDF is about tenfold (Bauer at si., 1961; Kimbrough, 1974; J. G. Vos, personal cowLication). F. Chlorinated Dlbanxofurans - Avian Toxicity Wham 1 day old chicks were started on daily oral doses of 5 /ug/kg TCDF daily most of them died within 11.S days. One of 6 chicks died g. .en dally oral doses of 1 /ug/kg bw. Symptoms of toxicity st the low as well as the high doses consisted primarily of weight loss, decreased food conswption and general unthrifty- nass. At autopsy the most striking findings were subcutaneous edema, ascites and hydroparicsrdlum and atrophy of the thymus. MOMS *11971 123 An inflammatory exudate was noticed on microscopic examination in the lung and the pericardial surface of the heart showed inflanatory changes. Congestion and hemorrhage into the gastrointestinal tract was also observed (Moore et al.. 1976). C, Brominated Dlbenzofurans - Mammalian Toxicity When dally doses of 4 /ug/rabbit of 2,3.7,8-tetrsbromodlbanzofurans were applied to the rabbit ear for 5 days giving a total dose of 20/ug/rabbit or about 5-6/ug/kg bv, the rabbits (4) developed hyperkeratosis of the treated ear and aesas of liver cell necrosis (Kimbrough, personal comwication). No other information is presently available on the toxicity of brominated dibensofurans. H. Longterm Toxicity Including Tumoroganeais Chlorinated biohenvls Mice When male BALB c/J mice were fed Arodor 1254 for 11 months at a dietary level of 0 or 300 ppm (49-9 mg/kg body weight) 22 of 50 mice in the experimental group end 24 of 50 in the control group survived. The livers of the test animals were markedly Increased in else, and showed a qualitative increase in porphyrin. A total of 9 experimental mice showed 10 neoplaetlc nodules (hematomas, hyperplastic nodules) in their liver ranging in size from 0.1 - 0.5 cm in diameter. Other morphological changes observed In the livers of the experimental group including pleomorphism, HOMS 211**2 124 irui of ntcroili, ind foci of adano fib cools. (Kirtrough and Linder, 1974). Neither tumors nor tha othac morphological changaa wara notad in tha controls. Only ona of 24 surviving mala BALfl c/J mica fad Aroclor 1254 for 6 Booths followed by a control diat for 5 months had a hapatoma (naoplaatic nodula) measuring 0.3 cm in diameter. Ito at al-, (1973) fad mala dd mica 500, 250, 100 and 0 ppm Kanechlor 500, 400 and 300 in tha diet respectively. After ona year, hepatocellular carcinomas wara observed in 5/12 mica fad Kanechlor 500 at a dietary level of 500 ppm, tha other 7 mica in this group had nodular hyperplasia (neoplastic nodules?). No metastases ware observed. Feeding a, 5, or / haxachlorocyclohexana (BMC) isommre with or without Kanechlor 500 at a dietary level of 250 ppm to mice for 24 weeks did not produce any tumors in tha liver of animals receiving only the PCB (Ito at al., 1973). Liver tumors were observed in mice receiving 250 ppm a BBC alone or 250 ppm PCB and either 100 or 50 ppm a BBC or 250 or 100 ppm 8 BBC. Thus PCBs enhanced the tumor development of liver tumors of a or 8 BBC isomers. lets Dietary exposure of male Sherman strain rats to levels of 500 ppm Aroclor 1254 (34.4 mg/kg body weight) for 6 months resulted in pronounced lipid aceiaulation in the liver which persisted for a 10 months observation period following the discontinuation of HONS 211973 125 the dietary exposure to PCBs. Adanofibrosis (cholanglofibrosls) which was observed in the livers of rats ingesting the PCB for 6 months was still present 10 oonths later. Whether this lesion is persistant could not definitely be daterained since high levels of the higher chlorinated biphenyl isomers were still present in adipose tissue and liver (Kimbrough, at el., 1973). Adenofibrosis is a focal proliferation of glandular epithelium forming ducts often surrounded by extensive fibrosis. This lesion usually occurs concomitantly in rat livers with hepato cellular carcinomas and has been produced by many known hepatocarclnogens. It was first described by Edwards and White (1941). The extremely long retention of some PCB isomars in rats was again noted when male Sherman strain rats ware fed 100 ppm Aroclor 1254 for 6 months and then allowed to recover for 16 months. 4.4 ppm of PCB derived material was present in the liver and 152 ppm in adipose tissue on s wet weight basis (Kiabrough, 1976). Continued dietary exposure to comerclal PCB mixtures in aaaals results in their storage in sdlpoee tissue and over an extended period high levels nay bs attained (Curley, at al.t 1971; Basse, at el., 1974). Blood levels are more a function of the #ose, the rate of metabolism, availability of binding sites in blood, chsmical characteristics of the coi^ound given, rather than the amount stored in adipose tissue (Smrek, at si., in preparation). MONS 211974 126 In another study, 200 experimental fmli Sherman strain rats were fad 100 ag/kg Acoclor 1260 In tha diat (11.6 - 4,3 mg/kg body vaight) foe approxiaately 21 nontha and 200 faasle rata warn kapt aa controla. A total of 184 experimental rata and 173 controls aurvlvod to aacrlflca at 23 nontha. Hapatocallular carclnonaa vara obaarvad in 26/184 axparlaantal rata and In 1/173 controla. Nona of tha controla but 146/184 axparlaantal rata had naeplaatlc nodules within tha livar (hyperplastic nodulaa). Areas of hapatocallular altaratlon warn notad in 28/173 controla and 182/184 axparlaantal rata. Tha incidence of tuaora in other organa was not altered by FCB ingestion and aatataaaa from tha livar tuaora ware not obaarvad (Kiabrough at al., 1975). Adenofibrosls of tha liver was also notad in this study. In reporting thaaa livar tuaora tha classi fication developed in a rat livar tuaor workshop was followed (Squire and LeVitt, 1975). Liver tuaora (aultiple adanoaatous nodulaa) ware also induced in 6/10 faaala Donryu rata with Kanachlor 400 but not in aala rats. Tha dietary exposure varied throughout tha study from 38-616 ppa (Kiaura and Baba, 1973). Tha induction of livar tuaora by known carcinogens In Sprague Dawlsy rata such aa 3'aathy1-4-dlmathylaninoaaobaniana, K-2-fluorenyl acetaalde sod diethylnitrosaaine was inhibited by Kanachlor 500 (Makiura et al.,1974). Groups of 50 sale and faaala Charles River rata ware fad dietary levels of 0, 1, 10 and 100 ppa of Arodor 1242, 1254, and 1260 raapactively (Calandra, 1976). At 3, 6, and 12 aonchs after MOHS 211*75 127 onset of the experiment, 5 rets per esch group end aex vere ecrificed. The surviving rats vere sacrificied 24 months after onset of the experiment. The only body weight depression noted was observed in the female rats fad Aroclor 1254. The author reported that Important histopathologic changes ware only noted in the rats fed the different Aroclors for 24 months. The liver weights were increased in the mala and female rats fad 100 ppm of Aroclor 1260 and 1254 and only in females fed Aroclor 1242 at a dietary level of 100 ppm. 1c was noted that of a total of 20 surviving male and female rats (10 mala and 10 female rats) fed 100 ppm Aroclor 1242 in the diet for 24 aonthe 8/20 rats showed liver leeions called nodular hyper plasia, 2/20 had hepatomas and 1/20 bad a cholangiohepatoma. A total of 27 male and female rats (13 mala and 14 female rats) survived to 24 aonthe in the groups of rats fed 100 ppa Aroclor 1254 of these 13/27 had nodular hyperplasia of the liver 4/27 showed hepatomas end 2/27 showed cholangiohepatoaas. Similar findings were aade in the total 27 surviving male and female rata fed 100 ppm Aroclor 1260 for 24 months. In these groups 7/27 aale and female rats had nodular hyperplasia of the liver. 5/27 had hepatomas, and 2/27 bad cholengiohepatomas. Croupe of 4 male and 4 female dogs were fed dietary levels of 0,* 1, 10. and 100 ppa Aroclor 1242, 1254, and 1260 respectively for 2 years, 1 female fed 100 ppm Aroclor 1242, 1 aale fed 10 ppa Aroclor 1242, 1 female dog fed 100 ppm Aroclor 1254, 1 female HONS 211976 128 fed 10 ppm and 1 mala and 1 female fad 100 ppm Aroclor 1260 diad. At tha dlatary laval of 100 ppm of alehar Aroclor 1254 or Aroclor 1260 tha body weight gain vaa slightly dacraasad for both aaxae- Aroclor 1260 alao raducad tha body valght gain In tha famalaa at tha dlatary laval of 10 ppa. In tha baaglas fad Aroclor 1260 at a dlatary laval of 100 ppm, tha llvar valght vaa lncraaaad and tha aina alkallna phoaphataaa vaa alavatad. All but 1 dog that dlad had althar chronic parltlonltls or chronic pneuaoaltls. Many of tha doga fad 10 or 100 ppa of dlffarant PCBs ahowad chronic Inflanatlon of a variety of organa. Mala doga fad Aroclor 1254 ahowad dlffuae hyparplaala of tha Intaratltlal calla of tha taataa and aaparaatoganaala (Menaaato, 1971). Dacachloroblohanvl No long tan atudlaa ara available. Polybromlnatad Biphenyls No long tan atudlaa ara available. Chlorinated Pibamaofurana and Chlorinated Naphthalanaa No loaf tan atudlaa ara available. tiuoouppreaalvm Effects of PCB and PBB Polychlorinated blphanyla (PCB) have bean Implicated aa lnuaosupprassants. Friend and Trainer (1970) foiad a dacraasad resistance to duck hepatitis virus la PCB exposed ducklings. HONS 211977 129 PCI ha* baan shown to produce lyuphold atrophy in rabbits (Strsst sad Sharaa. 1975; Vos and Beans, 1971), chickens (Flick st al., 1975; Vos and Koeaan, 1970) and gulnaa pigs (Vos and DeRoiJ, 1972; Vo* and vm Gandarsn, 1973). In addition, PCBs wars found to supprsaa huaoral lanEuns raaponsas to asvaral antigens In rabbits (Kollar and Thigpsn, 1973, Street and Sharaa, 1975) and gulnaa pigs (Vos and OslalJ, 1972, Vos and van Gendaren, 1973) and to suppress call aadistsd loauns raactlona In guins* pig* (Vos and van Gsndsran, 1973). In all of thsss studias tha PCBs studlsd h*va baan conarclal mixtures; ths rola of contaalnants such as chlorinated dibsnsofurans nasds to bs considered. Tha offset of HBB on ths 1sauna syatsa was studlsd.In chlcksns and gulnaa pigs (Vos and von Gandarsn). In chlckana ths atrophy of ths lysphold tissue was observed In bursal follicles and In ths spleen. It suppressed ths huasral lasuns response In guinea pigs. MDTACBMICITT BMP TBUIOLPCT Polychlorinated Blohenvls The aucaganlcley sad cytotoxicity of PCBs. Aroclor 1212 and 1254, were studied by Crxea st *1., 1975, a, b. The lack of eutagealc actlvii, as asssured by the doalnsat lsthsl test vss reported for eel* Osborne-Msndel rats subjected to four different dosage reglaeat: (1) single oral Intubations of 425, 1250 or 2500 ag/kg (for Aroclor 1242); (2) five dally dosas of 125 or 250 ag/kg (for Aroclor 1242), 73, 150 or 300 ag/kg (for Aroclor M0NS 211971 130 1254); (3) five daily doses of Arocloc 1254 at 150 mg/kg, than atarvad ovarnlght prior to mating; (4) chronic faadlng Cor 70 days at a dintary laval oC 25 or 100 ppm prior to mating (Grata at al., 1975a). In a aacond study, bona marrow and aparaatogonlal ctlla wara cytoganatically lnvaatlgstad from Aroclor-traatad Oaborna-Mandal rata. Aroclor 1242 waa glvan orally at alngla doaaa of 1250, 2500, or 5000 at/kS or aa four dally doaaa of 500 mg/kg/day; Aroclor 1254 vaa admlnlatarad orally aa flva dally doaaa of 75, 150, or 300 mg/kg/day. No atatlatlcally algnlfleant incraaaaa In chroaoaomal abarations ware found for the Aroclor-traatad rata. Aa indicated by Graen at al , (1975 b), while theaa atudiea wara negative In regard to chroaoaomal mutationa, they do not entirely rule out the poaaibillty that PCBa may induce point mutationa. To data, there are no known raporta in the literature concerning the Induction of point mutationa by PCBa in laboratory modal ayataaa. The obaarvationa of Nilsson and Kaval, 1974, in Drosophila where no chroaoaomal breakage waa found among Clophen A30- or A50-treated fliaa, are fully conalatant with thoaa of Green at al., (1975, a, b). TOtATOLOGT Female NHBX mica, orally admlnlatarad 0.025 mg/day of Clophen 60, a'..owed evidence of an lncraaaad aatrua cycle from 6-6 + 2.5 daya to 8.7 +4.3 daya and a dacraaaad ova implantation rata from 87.0 to 79.5Z (Orbarg and Kihlstrfta, 1973). KihlatrSm at al., (1974 a), alao demonstrated a decrease In the frequency of Implanted ova HONS 211979 131 among young animals nursed on milk containing PCB (Clophen 60). Both the male and tha female contributed to the magnitude of the reduction of implantations. The mechanism by which exposure of the developing male to milk containing PCB leads to a reduction in the precenc of ova Implanted is unknown but need not necessarily involve dominant lethal events. No reports of frank terete were found, but a number of studies indicated the facility of PCBa for placental transport. Grant et al., (1971 b), showed that the concentration of PCB in fetal tissue from oral administration of Aroclor 1254 was doee-dependent and that accumulation occurred in fetal liver where higher concentre* tions were found chan in maternal liver. Placental transport of PCB has been reported for the rabbit, rst, mouse, and cow (Grant et al., 1971; Platonov and Chen, 1972; Berlin et al., 1975; Halvas and Brandt, 1973; Brandt and Ullberg, 1973). Evidence of placental transfer was also observed among Yusho patients. Wnila PCB has no known or clearly defined teratogenic effect in mammals, their easy passage across tha placenta suggests the possibility of potential for soma form of fetal toxicity. Chlorinated Plbensofursns No Information available. Brominated Biphenyls 15 pregnant rats were force fed 100 mg hexabromoblphenyl/kg HONS 132 body woight on days 6, S, 10, 12, 14 and 16 of pregnancy In corn oil. 14 control rata were given corn oil. The papa were obtained by ceaarian on day 19 of pregnancy. No terata were observed and no difference between control and experimental rats was noted in any of the other parameters studied5 control and 6 PBB treated animals were treated with colchicine and bone marrow waa studied cytogenetically. Animala treated with PBB and colchicine had higher metaphase and mitotic indices than non-treated animals. Chromosome aberrations were not noted (Fiscor and Marts, 1976). Toxicity of Chlorinated Naphthalenes The following information la derived from a review published by the EPA, namely Environmental Hazard Assessment Report 'Chlorinated Naphthalenes', EPA-560/8-75-001*. For literature references please refer to this report. Animal Studies The following tables summarize the available studies in laboratory animals. Three routes of exposure are reported. This dociaent 's available through the National Technical Information Service, Springfield, VA 22151. 'Preliminary Environmental Hazard Assessment of Chlorinated Naphthalenes, Silicones, Fluorocarbons, Ben*snspolycarboxylates and Chlorophenols'. (NTIS accession number PB-238-074/AS) MONS 211981 133 Compound MonoOi- TriTrlTri/Tatra Fiou Panta/Hau- Panta/Haxa Pant*- HaxaHau* HauHapcaehloroOct aOcca Soaclai lac Mouae Rat Rabble Rac Rat Rac Cul&it pig Rat Rat Rat Rabble Rat TABLE 9 (a) Dietary Expoaure Doaa Laval Oavi Expoaad Non* raportad 5g/Kg feed 15 2.5 at/day 300 a*/day IS af/Kg body we. SO f day 20 9-136 60 63 300 af/day 33 100 at/day 55 2.S at/Kt 1 dll at 20 */Kf 63 at/Rt 200 at/Kt 1t 0.5, 2 or St/Kt 8* 8* 86 Nona raportad Slnfla doaa 22 Efface Re do re Incraaaa In llvar weight, growth lapaltad, rough coat. Nona* Facey laflltreclon af liver. None. Jaundica/fatty degeneration of llvar. Daach with yallow livers and axtraaa fatty degeneration. Daath with yellow llvara and axtraaa faccy daganaraclon. Death-Severe weight loa* fatty degeneration of liver. Halfbc lota. Haight loaa. Daath. Daath In savan daya. Decrease In llvar but not plasaa Vie. A. *(! HONS 211982 TABLE 10 (b) Inhalsdon Studies Compound TrlTrlTrl- Pasta- Sptelaa Lac Rat lac tat Dose Level 0.01-0.2 sg/L 1.31 ag/L 10.98 ag/L 1.16 sg/L Days Exposed Effete Reported 2 hr/day for 20 daya L6hr/day for DA deye I6hr/day for 102 daya 16hr/day tor 12 daya No toxic effect. No toxic effect. Slight livar dlicoloratlon. It rata ahowad incraaaad farcy daganaration Jaundice, enlarged Liver end 69t fatality. Mo atudla* hava baas raportad tor cha othar chlorlsatad oaphthalanaa. (c) Daraal Studlaa Coaeouad Mono* (tons- TrlTrlPastaPasta- Uau- Ssaelaa Kabblt(aar) Rabbit(aar) Lac(akia) Mica (akin) Svlna (akin) Rabbit(aar) Rabbit(aar) Cone Aonllad 90 ag/g Acatona 100 ag/g Acatosa Mot givas Mot glvan M sg/LtH. 30 ag/g acatooa/day 10 sg/g acatosa Daya of Asollcatlon 1-7 5-7 2 hr/day/*0-60 daya 2 hr/day/*0-60 days 6 daya/mak-* maka 1 1 f fact Mild reddening. Severe reddening, no decreeee of eebeceoue glands. Nona raportad Nona raportad Slight hyparkaratoala. Mild daraatitla, hair follcular attanuatlon. Decreeee in eebeceoue glands SCI mons 211983 TOXICITT IH BIRDS Turkay Poulti Panta/Haxachloronaphthalanaa In tha diet. (a) S ppm for AO daya cauaad 23X raducclon la valght tala with 6.5X mortality. (b) 100 ppm - all poulta diad within 33 daya. Hlato- pathologic examination ahowad aalargad and darkanad 11vara. Tha LCjq vaa 20 ppm with an avaraga dacraaaa la waight of 51X. Chickana Panta/Haxachloronapthalene in tha dint. (a) No affacta raportad at diaenry lavala of 4 and 20 ppm (33 daya). (b) 100 ppm pravantad agg production (33 days). (c) 2330 ppm cauamd 100X mortality (14 daya). Effacte raportad lacludad anlargad flbroua 11vara, lack of faathar pl^amtatlon and paricardlal and paritonaal adama. A fourfold incraaam In Vitamin A in tha diat dacraaaad tonic affacta. 136 H0NS 2119*4 TOXICITY IN DOMESTIC ANIMALS Ingestion Is the most important route for exposure of domestic animal*. Cattle Highly chlorinated naphthalene poisoning in catcla is known as bovine hyperkeratosis or X-disease. Detailed information on toxic doses is lacking. Fenta/haxa mixture at a dose level of 5.5 mg/Kg body vt (orally) for 5 days caused a sharp drop in plasma Vit. A by the end of the third day and depressed plasma Vit. A for over 30 days. A single oral dose of 1 mg/Kg body wt of hexachloronaphthalene caueed mortality within two weeks. The first effect of chloronaphthalene poisoning is an interference with the biotransformation of carotene to Vit. A. The Vit. A. deficiency is quickly followed by inflammation of the oral mucosa, lacrimation, excessive salivation and irregular food consumption. Gross physical effects that develop during the course of disease include a general thickening of the skin caused by over development of the skin's hairy layer with loaa of hair (hyperkeratosis). There may be degeneration or irregular growth of the horns. Continued exposure results in anemia, dehydration, loss of weight, fever and death. There nay be sr ere liver damage. Swine No toxic effects were observed at levels of 100 mg/Kg HONS 211985 137 body vt of the chlorinated naphthalenes (degree of chlorination was not seated). 150 mg/kg body vt caused a marked depression of plasma Vlt. A, and death occurred at a dose level of 198 mg/ Kg body wt. Sheep Toxic effects are observed at doses of chlorinated napthalene 10 X that required to produce toxicity in cattle. Sheep do not show cutaneous hyperkeratosis or an excessive drop in plasma Vlt. A as observed in cattle. Effects reported include nasal discharge, weakness, loss of weight, loss of appetite, ascites, necrosis and clrrohsis of the liver and cardiovascular effects. Toxicity In Man The most important route for mens occupational exposure to chlorinated naphthalene is via the inhalation route. Dermal absorption route nay also occur. Penta/haxachloronaphthalene at a concentration of 30 mg/g acetous, applied to tha backs of human volunteers for a period of eiz weeks caused typical chloracne, -Daily topical application of mono-dichlorlnatad naphthalene to the human ear in mineral oil suspension for 30 days caused no observable effect. Two clinically distinct but oftern concurrent A syndromes have bean described, namely, liver necrosis, and chloracne MONS 211986 x j8 & with Itching. A number of deaths Involving acute atrophy of the liver have been reported: with the course of the disease similar to other forms of severe liver damage. Autopsy of fatally exposed workers show severe yellow atrophy of the liver. Definite data relating to dose response effects are In general lacking. However, two daraal studies with man have been reported. SUMMARY The degree of toxicity of chlorinated napthalenes Increases with the degree of chlorination. The higher chlorinated naphthalenes (especially hexa and penta) have been associated with chloracne and liver damage In man. Dose response effects have not been estab lished. Cattle ere more susceptible to the effects of chlorinated naphthalene then other domestic anlmels. The effects observed Include development of Vit. A deficiency, hyperkeratosis and liver damage. HONS 211907 139 HUMAN EXPOSURE TO POLYCHLORINATED BIPHENYLS I. Introduction Although the polychlorinated biphenyl* (PCB*) have been used in a wide variety of industrial application* in the U.S. from 1930 until 1970, when their distribution was voluntarily restricted to closed systaas, there is scant information on human exposure to adverse huaan health affects in the U.S. In the U.S., minimal huaan exposure of the population to PCBs is limited to food, air and water, while significant huaan exposure appears to ba United to sports fishermen consuming fresh water fish froa contaminated straaas and lakes: and to occupational exposure in industrial workers II. Dietary Exposure to PCBa The May 1972 Report of the Interdepartmental Task Force on PCBs suaaarisad the findings in food sampling progress con ducted by the Food and Drug Administration and the Department of Agriculture. Jslinek et al., (1975X have provided an update to that summary and evaluated trends which Is presented below. The 1972 Task Force report outlined three broad sources of food contsalnation. These are restated here in order to show which sources have been effectively controlled and which sources appar ently persist, as reflected by results obtained in continued food saapling. HONS 211988 140 1. Environmental contamination - background levels of PCBs In fish from lakes and streams. 2. Industrial accidents - isolated Incidents involving direct leakage and spillage or contact of PCB fluids and other PCB containing materials on animal feeds, feed Ingredients or food. 3. Food packaging materials - PCB migration to foods packaged in PCB contaminated paper products. As discussed in the 1972 Task Force report, there were a num ber of other positive findings in foods which could not be traced to one of these broad sources prior to 1972 and the source of such occasional findings remain speculative. However, in more recent years, essentially all positive findings of PCBs in the food sup ply have been attributable to one or more of these three sources. Jelinek and Corneliussen (1975) reported that for the 19*9-1975 period, there has been a significant decrease in PCB levels in all foods with the exception of fish, where no particular trand has bean noted. From that observation they concluded that procedures instituted to exclude the use of PCBs in "open" applications have been affective, but that more needs to be done to prevent Chair entry into the acquatic environment. Jelinek and Corneliussen (1975) presented data on FDA's sampling program showing a vary conclusive decline in the FT* 1973-1975 MOWS 211989 1-1 period in uni of occurrence rate of PCBe and maximm levels found in Bilk, eggs, chaese, animal feeds and components, processed fruits, and baby foods. Except for fish, occurrence rates in FY'75 had dropped to zero or less than 12 for all categories, and there were no findings in excess of the existing temporary tolerances. Similar data were presented covering USDA's sampling of meat and poultry which also showed a decline, to 0.32 positive and only 0.62 above the temporary tolerance. Jelinek and Comeliussen (1975) also reviewed FDA Total Diet Study findings (FY'71-75) which showed that all food classes of the total diet have declined to no PCI occurrence and no calculable daily intake of FCBs, except in the maat-fish-poultry composites. About 402 of these composites continue to contain detactable PCBs, although only traces have been detected in the latter years. The limit of quantitative detection in the Total Diet Study is about 0.05 ppm; positive findings below that level are generally reported a* traces since they are not reliably quantitated. Occurrence of PCBs ended in 1973 in all other categories, notably in dairy product composites and the composites of grain and cereal products. This probably reflects the control measures effected on animal feeds and on the control of ingredients in recycled paper v sd for food packaging. What remains, based on the total diet study, is a continued low level occurrence (about 402 positive and only at trace levels) in the meat-fish-poultry *ON$ 211990 142 composites. The fact chat lavala la chaaa coapoaitaa hava declined to only.traces further supports the inference that the neat and poultry and eggs no longer contain detectable PCBs and that the low level findings are primarily due to the fish in those composites. This would imply chat the PCB levels in the diet may have "bottomed out" and may remain static until such time as there is a change in the levels for fish. Estimates of dietary intake of PCBs based on the Total Diet Study must be taken as gross estimates because only traces ere currently found and some type of eatimeted value must be assigned to those traces to calculate microgram per day intakes of PCBs. Jelinek and Corneliussen used a sat of consistent assumptions for those traces in computing the daily Intakes for the FY-1971-75 period. Their calculations showed a steady decline from IS meg per day down to 8.7 meg/day, with all of this coning from the meat-fishpoultry group in FY'74-75 (minor and declining portions attributable to other food classes in earlier years.) Review of this information implies that, although the various assumptions for trace residue reportings snd varying food consump tion dats have a significant effect on dally intake calculations, an overall estimate in the order of 5-E meg/day PCB dietary intake of PCBs for th general public seems reasonable. This figure would not be applicable to diets which may include regular consumption of fish from certain locations with high PCB levels. HONS 211991 143 III. PCB Residues in Fish Review of the data gatharad by various agencies on fish resi dues indicates a serious incompatibility of sampling programs for obtaining data which reliably define trends in PCB levels. The Food and Drug Administration's primary concern is the levels which exist in the edible portion of commercially imported fish which are marketed interstate. Therefore, in most cases, the heads, entrails, skin and fins are excluded. Furthermore, FDA sampling has not included fish which are caught and consumed locally (freshwater fish) and the sampling has dealt with the most impor tant consumption species which are primarily of marine origin and which ganerelly do not contain PCB levels as high as freshwater species from certain locations. In order to determine the human exposure to PCBs through dietary fish, one must know the levels of PCBs residues in the edible portions of fish, and the amounts of the various kinds of fish consumed by the population at large, and by special subpopula tions. During the past few weeks, information has bean compiled by National Marine Fisheries Service-NOAA on the PCBs levels of these residues in marine and freshwater fish and shellfish, and on dietary habits. Table 11 lists the 20 most important types of fish eaten in the U.S. today, and the mean daily amount of each type eatsn by #ONS 211992 144 TABLE 11 Fish and Shellfish Contrition in the United States (September 1973-August 1974) Total Tuna (saialy canned) Unclassified* (mainly breaded, incl. flah sticks) Shrinp Ocean Perch* Flounder Crabs/Lobatera Salmon Oysters/Scallope Trout** Cod* Baaa** Catfish** Haddock* Pollock* Herrin*/ Smelt/ Sardines Pike** Halibut* Snapper Whiting All other tlassified Amount (mill.) ifeiZlli Pertant of total br vt. Nuaber of Mean amount Actual Users per user (millions) (araas/dav 2957. 1 634 100. 21.4 197. 130. 18.7 6.1 2 542. 18.4 68. 10.0 3 301. 4 149. 5 144. 6 113. 7 110. 8 101. 9 88. 9 88. 11 78. 12 73. 12 73. 12 73. 15 60. 16 54. 17 33. 18 32. 18 32. 20 25. 152. 10.2 3.0 4.9 3.8 3.7 3.4 3.0 3.0 2.7 2.5 2.5 2.5 2.0 1.8 1.2 1.1 1.1 0.9 5.1 45. 19. 31. 18. 13. 19. 14. 9. 12. 7.6 7.5 11. 11. 10. 2.5 5.0 4.3 3.2 n.a. 8.3 9.7 8.6 7.6 10.6 6.7 7.8 12.3 8.1 12.0 12.1 8.6 6.8 6.7 17.4 8.0 9.3 9.7 a*t* * Mainly iverts ** Fresh water HONS 211993 the subpopulations of actual uaara. Tha Tabla lncludaa all dietary fiah, fro* aport fishing and fro* doaaatic and imported coaurclal flailing. Thaaa 20 ltama comprise 951 of tha fiah products aatan. Tha raaulta ara takan from information in a recant Saafood Conauaptlon Study by tha National Purchaaa Diary of Schaumburg, Illinois, sponaorad by tha Tuna Raaaarch Foundation, and National Marina Fiahariaa Sarvica data on tha amount of fish and shallflsh procasaad and distributad in tha U.S. Savaral items in tha Table ara worth noting for purposes of tha present analysis. (1) Although ninety-three percent of tha U.S. population (197 million) eat fish, tha average annual par capital consumption of fish is small: 15.0 lbs/yaar--fish aster; (2) a large "Unclassified" fish fraction exists in tha U.S. diet, ranking just below tuna in importance; (3) tha major portion of many of our moat familiar types of saa food is imported; and (A) freshwater spades, lad by trout, bass, and catfish, comprise about 9X of our total fish diet. A compilation of FCB Data, representing tha results of all tha measurements known to NMFS of FCBs in fish used in the U.S. diet is presented in tha Compendium of PCB data. It includes dsta from government reports, and state and private publications. As effort was made to include current, unpublished data from marine laboratories. The compilation includes the results of a comprehensive literature search by the NQAA-Envlronmental Data Service (EDS). An EDS-EKDEX dsta search for unpublished dsta HONS 211994 146 and a EDS-OASIS March wara made, crossing marina food spacias with PCBs. two significant sourcas of data--tha 1975 FDA-Market Basket results, and the results of tha extensive Fall 1975 New York State Department of Environment analyses vara not available to NMFS at the time Compendium was published. However, FDA has indicated that the 1975 Market Basket results do not differ dramatically from 1973/4 data. Preliminary results of the New York Study are included in Table 13. Table 12 summarises the PCB information assembled in tha Compendium relating to the 20 most important kinds of dietary fish identified in Tablell. Although at one time or another, some PCB measurements have been made on many fish, Tablell points out the inadequacy of information on PCB residues in the most important items in the fish diet. Sampling and analysis have been sporadic and not designed to monitor treads in human exposure. Systematic surveys of neither the important items nor the species most likely to be contaminated have been undertaken. Very few recent (1975) meaenramante have bean reported. However fragmentary the data on PCB levels may be, they do not include a single measurement exceeding 1 ppm in any of the ten most important fish foods, except for Dover sole and crab taken in the lMediate vicinity of Los Angeles sewerage outfalls, and crab living on contaminated sediments in Upper Chesapeake Bay. MONS 211995 147 Rank# 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. TABU 12 Maximum Reported PCB Residue Levels In Fish Muscle (White Meet) Kind Range and Max. Level Reported (ppm) (see Compendium) Tuna Unclassified (mainly breaded) Shrimp Ocean Perch Flounder/Sole Clams Crabs/Lobeter Salmon Oysters, Scallops Trout (axel. Lake Trout) Cod Bass Catfish Haddock Pollock Harring/Snalt/ Sardinas (narina Pika Halibut Snappar Whiting 0-0.49 (1973) no data 0-0.167 (1971) 0-0.25 (1974) 0,6.3* (1972) 0-0.819 (1974) 0-4.9b (1972) 0-0.5 (1974) 0-0.43 (1974) 0-0,56 0-0.4 0.3-8.4* 0-4.4d 0-1.16 0.0 (1972) (1974) (1974) (1974) (1974) (1974) 0-1.6 0-0.65 0-0.149 0-0.1 0.0 (1974) (1974) (1974) (1974) (1974) Mean Level in the L'.S. Diet Unknown* Unknown Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* Unknown* # Thass 20 itaas conposa 93Z of the fish in the U.S. diet. Probably wall balow 1 ppn *Single Dover Sola taken near Los Angelas nunicipal sewerage outfall. The mean value of Sole samples taken in this vicinity was 1.3 ppm in 1973. Mean values of other saaplss taken away from outfalls were leas than 1 ppn In 1973. In any case, although Dover sola la an important food fish in northern California, it ie not taken comercially in Southern California water*. bSingle yellowneck crab taken near Loa Angelas outfell-mean of group including this crab was less than 1 ppn. A conpoeita of Upper Chesapeake Bay crab contained 1.2 ppn in 1974. cTaken In 1974 fron St. Croix River, Wlacon*In--naan of 4 White Base was 6.2 ppn. dTaken fron Mississippi River, Minnesota-mean of a saarpla of 3 catfish was 2.3 ppn. MONS 211996 148 No more than 1.6 ppm has baan raportad In items 11 through 20 in importance, with tha excaption of frashvatar bass and catfish living in river and lake areas known to ba contaminated with PCBs. None theless, as emphasized in tha table, there have been no PCB measuremanes on tha important unclassified dietary fish items, and the analytical information is in no Instance systematic enough to permit an estimate of the actual mean PCB values, hence human exposure. The strongest statement that can be made, based on the Incomplete data summarized in the Compendium, is that most of the Important fish mast Items eaten in the U.S. probably contain mean PCB residue levels well below 1 ppm. Fragmentary evidence also leada to the conclusion that the PCB levels in fish liver, roe and oil are approximately tan times that in the edible muscle and the PCB concentrations ere greatest in the older larter oily fish, especially those that spend part or all of their lives in contaminated lakes, rivers or estuaries. Table 13 lists the location of the species where high PCB levels, exceeding the PDA 5 ppm guideline, have been reported. Although the trout, salmon and chub in lakes Michigan and Ontario and the Hudson klver baas, perch and eel constitute only a few percent at meet of the national fish diet, in some areas they are a significant nsrt of the fish diets. In order to discover what is presently known about U.S. fish consumption habits, a wide range of government and private sources 149 HONS 211997 TABLE 13 Areas Where at Least One Example of High* PCB Residue Levels Have Been Reported in the Listed Species Species Striped Bess Chub Carp Chain Pickerel, Aleulfe Coho Salmon Chinook Salmon Steelhesd Trout Lake Trout Bass, Smallmouth White Perch Alevife White Sucker Walleye Base, Largamouth Yellow Perch American Eel, Craba, Sturgeon Rock Bass, Catfish Bluefiah Location Hudson River, New Jersey Lake Michigan Mississippi River, Minn., Lake Onandaga. .'iY Hudson River Lake Michigan, Lake Ontario, Hudson River Lake Michigan, Lake Ontario, Hudson River Lake Michigan, Lake Ontario Lake Michigan, Lake Ontario, Lake George . NY Lake Onandaga, Genaasaa River, NY Hudson River, Mohawk River, St. Lawrence River Lake Onandaga, Hudson River Hudson River Hudson River, Mohawk River Hudson River, Mohawk River, Black River, NY Hudson River, Mohawk River Hudson River Hudson River . Hudson River Long Island Sound (muscle from one large oily fish contained 8 ppm) This Tsble includes recent data from New York State Department of Environmental Conservation not included in the NMFS Compilation. The Hudson River above Tort Edwsrd, New York, is not contaminated with PCBs and the residue levels in fish sre quite low. *Above the current 5 ppm FDA Guideline. MOWS 211998 150 vara consultad. A numbar of pra-1970 studios *rs tvsilsbls, including s 1969 NMFS Survsy of Fish Purchssss snd a 1968-70 HEW Isa Stats Nutrition Survsy. Ia addition, a fsv specialized rsports havs bssa publishod, including annual FDA Regional Basket Survsys that do not induda fish sstan in rsstaurants and inatltutioaa or by sport fishermen, and survays by tha Sport Fishing Xostituta and a 1974 Tsxas AIM Analysis of Seafood Con sumption Pattarns in Texas. Howavar, tha most up to data and coaprohansiva data ara svailabla in tha pravlously aantlonad 1973 74 Saafood Consumption Study by tha National Purchaaa Diary. Tabla 14 suaaarizas rasults of fish consumption daaographic information that dafinaa tha nuabar of U.S. conauaars who would axcaad an arbitrarily sat dlatary micro-constituant laval. To data, this kind of information is svailabla only for total dlatary fish and ona aubcoaponant, tuna. Tabla 14 lndicstas that 75 million U.S. conauaars aat aora than tha national svarsga of 18.7 grans of fish par day and 29 million would axcaad tha prasant dlatary intaka Halt of PCB (175ncg/day) if tha 35 or aora g/dsy of fish aaton was contaninatad on svarsga to 5 ppm, tha currant FDA guldallna laval. Thara is, of couraa, no avldanca that tha mama Pd laval in fish is anywhara naar 5 ppm. Nonathalaaa, should it in tha futura ranch 1 ppm, than by an axtrspolatlon from Tabla 14, an astlaata can ba aada that about 200,000 U.S, conauaars would ba axcaading tha 200 mcg/day FDA Halt. As HONS 211999 151 TABLE 14 Number of I'.S. Consumers Who Eet More Then X X (/dav> 0 18.7 (national avg.) 35 50 100 150 Number (thousands of consumers) Total Fish Tuna 197,000 130.000 75,000 5,700 29,000 1,300 14,000 400 2,000 43 400 10 * Derived from graphs on pp. 21 end 23 of the Seafood Consumption Studyi September 1973--August 1974 by the National Purchase Diary, Shsumburg, Illinois, 1975, Sponsored by the Tuns Research Foundation. HONS 212000 152 another hypothetical cue, if the PCB level in tuna should ever reach l ppm, then about 5,000 tune fish consuaers, those who est in excess of 175 g/day, would exceed the ZOOmcg/day limit from tuna alone. Survey data do show that in general U.S. fish esters include s wide variety of fish items in their diet. Thus, even if certain items contain toxic micro-constituents st or near FDA guideline level, the great majority of consumers would receive only a smell fraction of their average dietary intake from these sources. Clearly, there is s need for demographic information, similar to that available on total fish and tuna, on the other principal foodfish items in the U.S. diet; and a need to extend and better measure the high-conamptlon tall of the distributions, to get a correct picture of the number and type of consumers eating in excus of lOOg/day of any single fish product. IV. Conclusions At pruant, it is difficult to estimate human exposure to PCB from eating fish, either for the population as s whole or for subgreopa at higher risk. Fragmentary evidence suggests that the exposure to the population as s whole from PCB residues in ingested fish .s probably well below 19 meg/day-conauner, based on PCB levels which sre probably well below 1 ppm and 19g fish consumed/day. HONS 212001 153 Although It It Important to know the total population exposure, the firat priority should ba given to defining the dietary Intake of pCBs in special subpopulations st high risk by reason of high con sumption of fatty fish, or high sensitivity to the toxic action of PCBs, Such populations might include pregnant woman and young children, dieters, low Income groups and sport fishermen living in coastal and lake regions where there is known PCB con tamination. Primate studies suggest thst pregnant women and young children will be particularly sensitive to PCBs, and dieters, and in soma areas low income groups may rely heavily on fish for food. For each subpopulation, the amount and kind of fish in the diet and the PCB residue levels in the fish eaten oust be obtained in order to determine axpoaure. None of the necessary information to define the human PCB exposure in subpopulations is presently known. The first priority in systematically monitoring and analyzing for PCB residues in fish should therefore be among the oily fish such as salmon, trout, mackerel, shad, sardines, herring, bluefin tuna, sable fish, white fish, striped baas, and blueflsh, that have been identified as significant food items in the diets of special subpopulations of conataers. Although onl, i dietary exposure to PCBs may occur through food, there is evidence of potentially significant exposure MOWS 212002 154 in those sub-groups of ths population who rsgularly consume fresh water fish from waters which are contaminated with PCBs at levels which exceed the FDA temporary tolerances. These sub-groups of the population would include sports fishermen and others who consume locally caught freshwater fish. In surveys conducted by the State of Mew York and FDA, samples of 17 species of fish were collected from 10 points on the Hudson River between Glens Falls and Alpine, 13 samples were collected at Glens Falls and 68 samples below Glens Falls. The data from these surveys Indicate that 53 out of 68 samples of fish collected at or below Fort Edward (approximately 7 miles below Glens Falls) con tained PCBs in excess of 5 ppm. The average level of PCB contamina tion in these 68 samples was 31.3 ppm. In contrast, only trace levels were found in the 13 samples obtained at Gians Falls. A high proportion of finfish and eels from all sampling points below Glens Falls contained PCB levels in excess of 5 ppm. The Fort Edward sampling point had the highest levels for the species examined, with individual samples ranging from 20.1 to 178 ppm. Twenty-eight of the 33 samples of coho and Chinook salmon from Imka Ontario exceeded 5 ppm, with a range from 21 ppm to 24.6 ppm. Of the 4 eample# of lake trout tested, results were in the range of lh to 15 pt .. All other spades of fish sampled from Lake Ontario were at 5 ppm or less PCBs. The Great Lakes represent another area of the nation contamina ted with PCBs. The Michigan Water Resources Commission has reported HONS 212003 155 chat stay surface water samples contained FCBa at concentrations above the detaction limit of 10 ppt. (10 significant level* of PCBa have bean found in rivers and streams discharging into the Great Lakes. Effluents from wastewater treatment plants servicing Industrialized communities have been found to be highly contaminated. Sampling surveys of Great Lakes fish have shorn that most species tasted contained detectable levels of PCBa and that residue levels were generally proportional to fish size (age) and highest in the predator species. Except for whitefish, the species of coMarcial or sport interest (trouts and salmons) from Lake Michigan were found to be highly contaminated with PCBs. Data obtained from lake trout collected from various areas of Lake Michigan show mean PCB levels ranging from 3.06 ppm to 11.93 ppm. The Michigan Department of Public Health has recently completed a study (Humphrey at al., 1976) which attempted to assess some of the consequences of human exposure to PCBs from the consumption of sportsfish caught in different areas of Lake Michigan. The study included exposed and control subjects from five areas of Michigan bordering on Lake Michigan. Exposed study subjects were those individuals who consumed st least 24 to 26 pounds of Croat Lakes fish per year. Control subjects were thoee individuals who consumed lass than six pour's of Great Lakes fish per year. A preliminary assessment of Lite findings in the study indlcsts that the most frequently recorded quantity of fish consumed by the MONS 212004 156 study participants was in the 24-25 pounds par year range. The highest recorded fish consumption over the cvo year period of the study was 180 pounds per year and the highest single season consumption was 260 pounds. Mean PCB levels in whole Lake Trout are reported as 18.93 ppm in 1973 and 22-91 ppm in 1974, and 12.17 ppm in Coho Salmon in 1973 and 10.45 ppm in 1974. However, comparisons of PCB levels in raw vs. cooked fish indicate that actual human exposure to PCB from fish consumption is less than might be expected from the raw fish data. This is not unexpected since preparation (trlsing awsy fatty tissue) and cooking have been shown to decrease the concentrstlon of PCBs. For example, the PCB lsvel in cooked Lake Trout consumed by the study participants ranged from 1.03 ppm to 4.67 ppm; in cooked salmon from 0.48 ppm to 5.38 ppm; and in other cooked fish from 0.36 ppm to 2.06 ppm. These levels are decidedly lower than the level of PCB contamination reported in raw trout and salmon. PCBs were found in 501 blood and breast milk specimens col lected from study participants during the study. The vslues ranged from a low of 0.007 ppm in blood in the control group to s high of 0.366 ppm In the exposed group. Although there was a wide range of blood values for stch quantity of fish consumed, there was a highlv aignif. ant correlation between the reported quantity of Lake Michigan fish consumed and the concentration of PCB in the blood of study participants. Using a natural log transformation HONS 212005 157 of PCB values, the correlation between amount of fish conaumad and PCB bio Of- lavala was significant (t-6.24, p 0.0001), with higher reported fish consumption being associated with higher blood PCB levels. No annual variation in PCB blood levels in humans could be demonstrated. The mean PCB blood values for the control and exposed groups did not appear to change markedly from 1973 to 1974. In addition, abstinence from Lake Michigan fish consumption for s period of 90 days or more did not change the PCB blood levels sig nificantly. PCB blood levels over the abstaining period shows variation but no steady decline In PCB. In fact, more subjects showed no change or a rise than showed a decline In PCB blood levels over time. The calculated quantity of PCB Ingested by eating Lake Michigan fish averaged 46.5 mg/year and ranged from 14.17 to 114.31 mg/year. PCB ingestion for each Individual was determined by proportioning the reported 1 fish consumption by frequency of species satsn and the cooked fish PCB levels for those fish. The cosunlty average for cooked fish was used in Instances were cooked fish determination were not available for a study participant. Because fish consumption was found to vary from year to year the average annual consumption for each individual for the two baseline years of study was used In each case. HONS 212006 158 Results froa this study Indlcats ths strong likelihood that the maximum allowable Ingestion of 1 meg/leg body veight/day recommended for protracted exposure to PCB Is being exceeded by the majority of the exposed group participants, and by inference, by a substan tial number of individuals participating in sport fishing and who consume quantities of containated fish. The calculated mean daily dose received by the exposed group is 1.7 aeg/kg/dsy and ranges froa 0.49 meg to 3.94 aeg/kg/day. If the average annual race of PCB ingestion froa fish indicated by these study results were con tinued over the years, and if net accumulation occurs, the average sports fisherman consuming contaminated fish could receive a total PCB dose equal to the 200 og safety limit in approximately 4.3 years. Under the same set of aasiMptiona, individuals consuming greatet than average eaounts of contaminated fish would reach the total dose level sooner. Mo adverse health effects or group of symptoms could be iden tified in the exposed group that were clearly related to PCB exposure. This implies that exposure to PCBs froa eating contamina ted fish at the levels observed and the presence of PCB* in these exposed persooa has not caused any observable adverse health effects at the time of tha study. However this does not exclude the possibility that effects too subtle for detection are occurring or the possibility of long-term adverse health effects. HONS 212007 159 Considerable scientific Interest has centered on the Yusho Incident in Japan where in 1968 human intoxication vich Kanechlor 400, a PCB manufactured in Japan, was noted when a heat exchange leaked this PCB into rice oil ("Yusho" oil) which was consumed by Japanese families. The typical clinical findings Included chloracne increased pigmentation of the skin, increased eye discharge, transient visual disturbances, feeling of weakness, numbness in limbs, headaches and disturbances in liver function. Most of the babies bom to mothers with Yusho had skin discoloration which slowly regressed as the children grew In size. Adult "Yusho" patients had protracted clini cal disease with a slow regression of symptoms and signs, suggesting a slow metabolism and excretion of the PCB in humans, probably resulting from a long biological half-life. A review of the literature extent in 1972 led to the following facts with respect to this tragic Incident in Japan: 1. The average PCB content of ths rice oil in the doseresponse epidemiologic study wes 2,500 ppm. 2. In this dose-response epidemiologic study, the average emulative intake of PCBs, leading to overt symptomatology, was 2000 -g. In this same study, the lowest dose leading to overt symptomatology was 500 mg. MOWS 212008 160 3. The toxicity seen was the result of the ingestion of the PCS* contaminant in tha rice oil. A swmnary updating of tha "Yusho" incident was presented by Professor Kurataune at the National Conference on Polychlorinated Biphenyls, Chicago, Illinois, November 19, 1975 (Kurataune et al., 1976). This report affects the original conclusions enumerated above. 1. PCBs Content of ''Yusho1' Oil. The first reported PCB determination was made on a canned contaminated rice oil produced on February 5, 1968, and whose consumption was associated with overt symptomology. One group of Yusho patients was associated with ingestion of this contaminated canned rice oil produced or shipped on February 5 and 6, 1968 (Kuratsuna at* al., 1972). The detailed epidemiologic studies are given in the papers by Kuratsuna et al., (1969) and Yoshimura (1971) and sum marized in English by Kuratsuna (1972), Kuratsuna et al., (1971) and Kuratsuna et al., (1972). Of 325 patients queried, 166 of 170 used cawed rice oil produced or shipped on February 5 and 6, 1968, and 143 of 155 used bottled rice oil shipped from February 5 to 15, 1968. Tsukamoto et al., (1969) determined that the organic chlorine . ntant of canned rice oils produced or shipped on February 5, 1968, was 1000 to 1500 ppm, by chemical and activation analysis. Since Kanechlor 400 had a 48 per MONS 212009 161 cent chlorine content, if one attributed all the organic chlorine to this specific chlorinated biphenyl, then this canned'rice oil contained, as an average, 2500 ppm PCBs. Kuratsune et al., (1969) reported that a qualitative GLC survey of 109 samples of bottled rice oil shipped between October, 1967, and October, 1968, shoved ''significant1' PCBe content only in those samples between February 7 to 10, 1968. The largest amount was reported in the sample of February 7, 1968, but the quantity was not given. No bottled rice oil samples were available for February 5 and 6, 1968. However, in the paper by Kuratsune et al., (1971) the most marked contamination of bottled rice oil was for that shipped on February 7, 1968; and the maximum chlorine content was 462 ppm. One may assusie that this number applies to the sample of bottled rice oil of February 7, 1968, giving this sample a PCBs content of 924 ppm. Kuratsune et al., (1976), reporting the work of Nagaysma et al., (1975b), gives the PCBs analyses for three samples of toxic "Yusho" oil, used by three independent families with "Yusho", as approximately 800 to 1000 ppm, These were GLC/MS measurements. Nagayema, et_ al., (1975b), described these samples as follows: "Klee oil used by patients with Yuso ("Yusho oil").* 3 samples of ric^ oil used by 3 Independent families with Yusho in 1968. These samples had been kept in glass bottles with glass stoppers at room temperature in our laboratory until 1974 when the analysis was made.'1 MOWS 212010 Kuratsuna (1976) pointed out that thus three samples wars canned oil and considered to have been produced and shipped on February 5 or 6, 1968. This is also suggested in footnote a, table 5, in the paper by Kuratsuna et al., (1976). It was reported previously that, based on organic chlorine determinations, Tsukesioto et al., (1969) reported an average PCB content of the canned oil of February 5, 1968, to be 2500 ppm. Now, based on BLC/MS, we have a PCB content of ce 1000 ppm for canned oil sample of February 5 or 6, 1968. What ere the possible explanations for this discrepancy in values? One possibility is based on the assumptions associated with the method of measurement by Tsukamoto et el., (1969); namely, determination of total organic chlorine. Thus, any orgnic chlorine preeent in the rice oil, which wee not a chlorinated biphenyl, would be included in the PCB analysis. Some possibilities include: chlorinated dibeneofurena, chlorinated naphthalenes, chlorinated paraffine, other chlorinated aliphatics and non-biphaayl aromatics. Another possibility is related to the distribution, with time, of thi contaminated rice oil samples. February 5, 1968, ccurred on a Monday. The first rice oil samples reported to contain PCBs were produced on this date (Tsukamoto at al.; 1969). If the leak first occurred sometime over the weekend (February 3 and 4, 1968), and MONS 212011 163 production was resumed Monday morning, Ftbruary 5, 1968, than ona would expect tha samples of this data to be higher In PCB content. Therefore, ona would axpact the first batches Monday morning to be highest with a diminution of PCB content with time. Tsulcastoto et_ el., (1969) reported the organic chlorine content of six canned oils produced Monday, February 5, 1968, as follows: 1020, 1170, 1070, 1080, 1030, and 1500 ppm. In the survey of bottled oils, Ruratsune at al., (1969) reported that for those samples where PCBa were found, the greatest amount was In the samples of February 7 (Wednesday); and after February 11. It wee inferred from the paper by Ruratsune at al., (1971) that the PCB content of the bottled rice oil samples of February 7, 1968, was 924 ppm. In addition, in Table 5, Ruratsune at al., (1976) gave a value of 134 ppm PCB for a rice oil produced on February 10, 1968. Thus, the available data does confirm a diminution of PCB contamination of the rice oil with time. This is sumsarized below: Date Source Feb. 5, 1968 canned Feb. 3 or 6,1968 canned (?) Feb. 7, 1968 bottled Feb. 10. 1968 bottled After Feb. 11 1968 bottled PCB content (ppm) Method 2000-3000 organic Cl ca 1000 GLC/MS 924 organic Cl 134 ? just detectable GLC (7) HONS 212012 164 The resolution of eh* varying PCB valuaa for tha canned oil cannot ba made. However, Kuratsune (1976) confirmed that tha production of rica oil had baan suapandad for an unknown pariod of time prior to February 6, 1968, when it resumed. 2. Dose-Raeponae Thera vara 1291 "Yusho" patianta aa of April 30, 1975 Oeme, 1975). Of thia total group, tha doaa-raaponaa ralationahip vaa davalopad from a datailad analyaia of 146 patianta (Yoshlmura 1971; Kuratauna at al., 1972). Thaaa 146 patianta all contused contaminated rica oil produced on Fabruary 5, 1968 contained 2000 to 3000 ppm PCBa aa Kanaehlor 400 (Taukamota at al., (1972) aatimatad that tha lowest doaa producing symptoaa waa 0.5 gn. If tha PCB content of thia cannad oil vaa ca 1000 ppm, aa dlaeuaaad in tha previoua section, then tha average doaa producing an overt affect vaa 0.8 gn. and tha lowaat doaa producing an overt affect was 0.2 gn. 3. Cauaatlva Frctora. Originally, rica oil contaminated vith a heat exchanger, Kanaehlor 400, a polychlorinated biphenyl, vaa aasociatad with "Yuaho" symptomatology (saa discussion in section HONS 212013 its 5 1 above), PCB* ware Identified In the contaminated rice oil consumed and in cha blood and tissues of Yusho patients. Therefore, the effects sees were attributed to PCBs. In the review by Kuratsuae at al.t (1976), a new factor was introduced into the system; namely, the caa&ad rice oil was also contaminated with chlorinated dibenzofurans to the extent of 5 ppm. In addition. In this same paper by Kuratsuae at al., (1976), they presented data of Bagayasa at al>, showing polychlorinated dlbenzofurans In the liver and adipose tissue of Yusho patients, while none was fouad la that of a control group. The ratio of PCBa to PQ)Fs In Kanechlor 400, a Yusho oil of February 5 or 6, 1968, in adipose tissue and liver from a Yusho patient were 50,000; 200; 144; sad 4 respectively. Thus, relative to Yusho oil, the liver with a PCB/PCD? ratio of 4 to 1, appears to selectively concentrate PCDFs relative to PCBs. Relative to PCBa, PCDFs are about 250 times higher in the rice oil than in Kanechlor 400 (Nagayama at al, 1975b; Kurataura at al., 1976).* They also stated that . . PCDFs &, be a factor la Yusho disease. In another *^It should be noted that we are comparing chlorinated dlbensofuran content of an unused Kanechlor 400 to that after prolonged use in a heat exchanger. MOWS 212014 166 paper Nagayaaa ec al., (,1975a) reported that It "is eald that the toxicity of PCDF la said to be from 200 to 500 time that of PCB". At the lower factor (200), con taminated rice oil would be expected to be twice as toxic as Kanechlor 400. Also, In this same paper they stated that th% symptoms seen in Yuaho patlente eeemed to be more severe than would be expected just from the PCB Intake associated with the oil. If PCDF is 200 to 500 tlaea more toxic than PCB (Nagayaaa at al., 1975a), then contaminated rice oil would be 2 to 3.5 tlaea more toxic than that expected from its PCB con tent, alone. Kuratsune at al., (1972), estimated that if the average amount of contaminated rice oil ingeated by a "Yuaho" exposed person, is related solely in terms of its PCB equivalent, then the amount of oil needed to be ingested, on that basis, would be 1600 al to 2800 al. At a PCB content of 2500 ppm, this la an Ingestion of 4 to 7 gm of PCBa equivalent; whereas at a PCB content of 1000 ppm, this is an ingestion of 1.6 to 2.8 gm of PCBa equivalent. Conclusion The c. plexlties and uncertainties associated with the most recent reports of the "Yuaho" incident in Japan, make it extremely difficult to quantify possible human health effects resulting from exposure to PCBa alone. MONS 212015 167 V. Occupational Exposure to PCBs The earnest reports of adverse health effects due to exposure of workers to PCBs in this country are probably those of Schwartz (1936) who described skin lesions and symptoms of systemic poisoning among workers who were said to have inhaled chloro diphenyls; their complaints Included digestive disturbances, burning of the eyes, impotence, and hematuria. Patch tests with the chlorodlphenyls were negative, and Schwartz speculated that mechanical plugging of the follicles of the skin as the fumes solidified on it were respon sible for the skin lesions; the chlorine present in the products were thought to then exert an irritating effect on the plugged fol licles, and to thus cause suppuration. No quantitative data were reported, but a number of preventive practices were recoonsnded. There have been numerous reports over the ensuing years (Schwartz and Bartow, 1942), (Graenburg et el., 1939), (Drlnkas at al,, 1937), (Good et si., 1943), (Schwartz, 1943) and (Haigs, at al., 1954) of cutaneous eruptions and of systemic manifesta tions (sometimes fatal) as wall, among marine electricians, machinists, capacitor and transformer manufacturing workers, and others oceupatlonally exposed to PCBs. However, in many of these reports the exposures are described as having been to mixtures of chlorinated hy** ~ocarbons, quite often of chlorinated naphthalenes and PCBs. MONS 212016 168 The skin lsslons dsscclbsd by Schwartz (1936) hava cons to be designated ganarally aa "chloracna." Chloracnc can ba produced by 4 nuabar of cha&lcal compound*, Including chlorinatad dlbanzofurans and carcaln laomara of tha chlorinatad dlbanzodloxlna (Kimbrough, 1972). Oily akin and larga pore* aaam to pradlspose to tha dlacasc, while tha oppoalta la tha caaa for smooth, taadar akin (Kimbrough, 1974). Chloracna also has occurrad among workers engagsd In the production of 2,4,5-T (Poland at al., 1971). Part of tha chloracna laslon rssaablas adolescent acna, but It la ganarally more sewers, and lesion distribution la Inconsis tent with, although It may ba superimposed upon, adolescent acna. It Is known that chloracna can be produced by either tha systemic absorption of chlorinated biphenyls or tha direct application of chloracnegenlc compounds to tha skin. Systemic affects sometimes result after occupational chloracna has manifested Itself; these may Include loss of appetite, nausea, edema of tha face and hands, abdominal pain, vomiting, and burning and soreness of the eyes. No fully satisfactory explanations have been made of tha develop ment of chloracna. Chloracna Is generally vary persistent, and there la the preferred control measure. An excellent review of this subject la that by Crow (1970). Tha U.S. .ccupatlonal standards for PCBs are: 8-hour timeweighted average exposure limits of 1 mg/m3 (skin) for tha 42Z chlorinated product, and 0.5 mg/m3 (skin) for the 542 chlorinated product (CPU, Title 29, Part 1910.93). The National Institute for Occupational Safety and Health has under way the preparation of 169 MONS 212017 criteria for a recommanded standard for occupational exposure to polychlorinated biphenyls. In addition to an environmental exposure limit, this document will provide comprehensive recommendations for medical surveillance, safe work practices, and engineering controls-to ensure employee protection during occupational expo sure over a working lifetime; It 1s scheduled for transmittal to the Department of Labor In late 1976 or early 1977. NIOSH also Is undertaking certain studies of employee populations (mainly In U.S. capacitor factories) In order to better define the nature and extent of any chronic and/or life-shortening effects of exposure to PCB. Practically all of the currently available Information on worker health and PCB exposure Is found in foreign literature. For example, Karppahan and Kolho (1974) reported on relationships between the concentration of PCB's in the blood of all, and In the adlpoee tissues of some, of 29 persons in "good health"* from Kolho has stated that the capacitor plant workers received quar terly health examinations, and, In addition to the clinical examina tion made at the time of the investigation, serum alkaline phos phatase, GOT, sad GPT activities were determined; the 6 employees with the highest blood concentrations of PCBe also had BSP excre tion tests performed. All results were normal. In view of r)sfms based on n<mi experiments that PCBa can induce liver microsomal enzyme activity, the authors also determined the half-time of ant'*>yrin before and after phenobarbltal Induction (1 mg phenobarbltal/kg body weight/day/3 days) for 6 capacitor plant employees and 6 controls; no enzyme induction was observed. Further, .because of claime that TCB's have effects on steroid metabolism, 4 capacitor plant workers were tested for ACTH (serum, presumably); all results were reported to be within the normal range (letter from S. Hemberg to A. C. Kolbye, Jr., 12/23/75. NONS 212018 170 three different employee groups In Finland. None of the employees hed no history of occupational exposure to PCBs, six of them hed handled PCB samples in an analytical laboratory, and the other eleven of them had been employed for 6 years in a capacitor fac tory where Aroclor 1242 ves used as the Impregnating fluid. It ves stated that average PCB concentrations In the air of the capa citor factory had not exceeded "Internationally accepted limits**," and that special attention had been given to skin protection. Table IS shows the observed tissue concentratins of PCB's; the authors were unable to detect any biological effects of the approxi mately 50-fold larger PCB concentrations in the blood of the capa citor plant employees, compared to the "unexpoaed" control group. Ow et si., (1974) conducted a survey to determine the degree of absorption and the health effects of exposure to "electrical grade" Arodor 1242 (that "did not contain any impurities1') for varying periods of tima. PCB concentrations in the air of a capa citor factory in New South Wales, Australia*, were measured, and 34 occupationally axpoaed employees (IS males, 19 females, ranging in age from 33 to 55 years) were examined and compared with volun teer controls (23 males, 7 females ranging in age from 20 to 50 *Tha Joint 11/ 'WHO Committee on Occupational Health, in its sixth report (World Health. Organisation Technical Report Series No. 415, 1969), reeomnded for international adoption a "safe concentra tion none" of 1 ag/rnr for chlorinated derivatives of diphenyl. **Aroclor 1242 la not manufactured in Australia (Ow et al., 1974). HONS 212019 171 TABLE 15 Concentration of PCB's Subjects In blood Pat basis m*/k Average Range In adipose tissue Pat basis ng/kt Saaple no Workers In capacitor factory 313 100-700 i 200 2 11 3 160 4 285 5 635 Persona handling PCS'a In analy tical laboratory Persona without any special exposure Co PCB's 53 5.4 33-71 3.6-9.9 not analyzed 1 2.3 2 1.5 From Larppanen and Kolho (1974) HONS 212020 172 yurt) having no history of occupational axpoaura to PCB'a. Study parameters included occupational and madical histories, PCB-In blood concentration estimates, and liver function Cserum bilirubin, alkaline phosphatase, total protein, and GPT, and BSP excretion) tests. The authors noted that exposed workers tended to complain of eye, face, and skin "burning," that the PCB "'ftae' ... has a pungent smell which often causae persistent body odour," and that the employees with higher blood concentrations of PCBs complained most often of the skin lesions Cone case of chloracna, five cases of "an eczematous rash on" the legs and hands), although there apparently was poor correlation of blood PCB's with the severity of the complaints. No significant health effects were observed among those of the 34 workers whose blood PCB concentrations were below 200 ppb. There was a statistically significant difference between the blood PCB concentrations of the exposed group end the control group CP lees than 0.01). Table 16 shows the concentrations of Aroclor 1242 In the capacitor plant air prior to and after exhaust ventila tion eyetarn "Improvements" had been made. (The Australian National Health end Medical Research Council recommended (Atmospheric Contaminants, 1970) exposure limit valuee of 1.00 mg/m for PCBs of 42Z and 34Z chlorine content, respectively). Table 17 shows that there was no lowering of blood PCB concentration among those workers tested two months after the installation of a.more MONS 212021 173 TABLE 16 Arodoro 1242 concentrations in the air inside capacitor plant before and after Improvement of exhaust ventilation system Aroclor concentration No. Areas In the Impregnation Room in ms/m^ Before After 1 Area in the unloading tank in front of exhaust register from operator's breathing zone 1.44 0.75 2 Area in the unloading tank not in front of exhaust register 2.22 0.7 3 General atmosphere near tank 1.08 0.18 4 Soldering area 0.32 0.08 From Ow at al (1974) HONS 212022 TABLE 17 Mean blood Aroclor 1242 levels bfor and after laprovenent of exhaust ventilation and the ncowo* datloo to use "suitable lnparvioua" gloves. Croup Before After Statistical differences Mean blood Aroclor levels In ppb. Retention tines relative to Aldrir l. 0.69 1.31 1.41 281.6 133.1 38.41 477.2 225.4 324.7 Not significant difference (P 0.01) Not significant difference 0 0.01) Significant 'difference (P 0.03) Era Ous et el (1974) MOMS 212023 officiant exhaust ventilation tyitn ud the concomitant recom mendation to veer "suitable impervious gloves" In order to reducs PCB absorption enrough the akin. The authora suggested that a failure to adhere strictly to the glove racowendation night explain the continued elevation blood PCB levela. Another poaalbillty night be that PCBa were continually aoblllaed fron atorage In adlpoae tiaauea. The Japanese Hlnlatry of Labor flagniarion for the Prevention of Diaturbancea due to Specified Chenical Subatancea. under the Act for Safety and Health of Workers, of April 28, 1971) eatabllahed an occupational esvlroaanatal ezpoaure Halt for PCBa of 0.5 n$/m3 at 25 C, one ata (Ad Hoc Coanlttea, 197&). Japaneae laportatlon of PCBa eosenced around 1990, and early uaea were aa dlalactrlca in capaeltora and tranafonars. In 1994, Kanegafuchl Chenical Induetry Co., Ltd. etarted PCI production la Japan and it waa at about thla tlan that chloracne eruptioua aaong workera were first reported. Slightly earlier (1993) incidents, however, were reported by Hare (1969), of changes attributable to PCBa. in blood and urine findings aaong capacitor factory workers. According to an Ad Hoc Coanlttea sponsored by the Japanese Pawl ton--iral Agency (1974), PCB coneantretlona in the sir of e Japanese capacltot factory (yhere the major PCB constituent wee "diphenyl pantachloride") were found to range fron 0.37 to 6-79 ag/a3; this waa batvean the years 1993 and 1997, and prior to tho 176 HONS 212024 data whan th aforementioned occupational axposura limit was promul gated. Tha hlghaat concantratlon waa measured vhara PCBa vara haatad to drlva out antcalnad/dlsaolvad air. Admittedly, tha analytical mathodology for PCBa in thoaa yaara waa not aa raflnad aa that now in uaa, but It la thought likaly (authora) that some employees vara exposed continually at concantrationa of several milligrams par cubic matar. Tha Janapaaa lltaratura ia aaid to have contalnad no furthar raporta on PCI-in-air measurements until tha apring of 1972, juat prior to tha auapanaion of thair uaaga. Tha production of PCBa waa dlacontlnuad in Japan in Juna 1972, and thair importation waa diacontlnuad in tha following Saptaabar. At that tlaa Haaagawa at al., (1973) raportad PCB measurements in tha air of ana PCB (Kanaehlora 200-600) production plant (0.005-0.02 ppm), four capacitor manufacturing plants (Kanachlor 300 uaad) (0.01-0.05 ppm), and ana plant vhara PCB s (Kanachlor 300) wara uaad aa a hast exchange medimi (0.002 ppm). Paak valuaa of 0.17 ppm and 0.67 ppm, raapactivaly, vara found in a capacitor plant "air-riddance" procaas araa and in a capacitor Impregnating araa vhara tank leak age had occurred. Estimated (presumably 8-hour workday) timeweighted average expoaurea of affected amployaaa wara 0.02-0.03 ppm. Airborne PCB a m tha capacitor plants wara raportad to con-slst of 70-80X vapor for material corresponding to Kanachlor 200, and of particulates larger th*., 0.1 micron for Kanachlor 300. Tha authors attributed tha 3/1 vapor/partlculate mix to selective evaporation of lov Bolling components of Kanachlor 300. 177 "ONS 212025 Although no measurements were available, Hasegava at al.. (1973) estimated that environmental conditions In carbonlaas copy papar manufacturing facilities (using Kanechlor 300) vara roughly similar to thoas found In capacitor factories. Beginning In February 1971, PCBs vara phased out as the mlcrocapsular solvent in this process, being raplacad by "SAS" or JOtC-oll;" however, during the period from November 1972 to January 1973, when occupa tional health surveys were being performed relative to SAS or KMCoil toxicity, PCB concentrations of 0.013-0.4 ppb were measured In the environments of these facilities- General environmental con centrations of PCBs at that time were lees than 0.0005 ppb, and outdoor PCB concentrations around the facilities vers 0.0043 ppb (Hasegava et al., 1973) and 0>009 ppb (Sato and Hasegavs, 1974), In an office room where carbonless copy paper was used, an air concentration 1.1 meg PCB/m^ was measured, and in the carbonless copy paper storage ares of a post office, PCB concentrations of 8.7-21.1ncg/m3 were detected (Nlshlyama et al., 1973). The Japanese Ministry, in its "Regulations for Physical Examinations for the Prevention of Disturbances due to Specific Chemical Substances," promulgated on October 14, 1971, advised PCB uaere to employees for dermal andhapatic "symptoms" and thmlr anamnesis, "subjective symptoms" such aa anorexia and asthenia, and urine urobilinogen, in the first (pre-employment?) physical examination, and to "survey" working conditions, blood tests and liver function tests in the "secondary" (periodic re-?) 178 MONS 212026 examination. A 1973 notice adivsed those users of PCB's ss heat exchange fluids to perform comprehensive health surveillance (Environmental Agency of Japan 1974), Yamamoto (1974) reported "positive" findings in 37 employees of 323 (total employment 706 in SI establishments) who vers given "special" physical examina tions in 1971. Hara (1969) reported a 20-30X incidence of dermel "symptoms," consisting of "distinctive hair follicles" in exposed arses such as the face, neck, and forearm, and pimple-like skin eruptions of the face and neck, among employees in a capacitor factory that had been in operation between 1953 and 1963 (pentachloroblphenyls) 1953-1957; Kanechlor 300, 1958-onward). No other, e.g., hepatic, dysfunction was observed, and no quantitative Information was reported in the reference from which the preceding report was obtained; however, in a follow-up survey of workers in capacitor factories, Hara at al (1974) reported that by one year after the suspension of PCB usage skin findings had become milder. Hasegava et al., (1973) reported on s 1972 survey of capacitor factories, in which they noted that persons working in environments that contained 0.2-0.3 mg PCB's/m^ (Q.02-0.03 ppm) showed dermatolpglc allmanta that included "brown chromodermatosls" of the dorsal joints of the hands . d fingers and nail had, and "acnaform exanthema," Several cases of comedo or acneform exanthema of the jaw, back, and thighs were seen also. These signs were no longer observed one month after the cessation of the handling of PCB's. HONS 21*027 179 In a factory where Itanechlor 300 vet employed a heat exchange medium, the environmental concentration of PCB's wee reported to be 0.02/mg/m^, and no dermal manifestations were observed among its employees (Hasegawa et al., 1973). Nor did Hashimoto (1974) observe any abnormalities among 236 workers in such a facility, to whom he administered "examination centered around liver function tests." In 1972, when Hasegswa et al., (1973) performed a health sur vey of workers in carbonless copy paper factories, i.e., 2 years after the use of PCB# in such processes had ceased, they noted no dermal effects, and no liver function, blood, or urine test abnormalities were seen, with the exception of "slight abnormalities In lipid metabolism". PCB levels in the blood of these workers ware reported as "approximately 0.01-0.02 ppm," or what amounted to a decrease to 10Z of the levels during the period of PCB usage. The authors' data indicated that PCB's collected from the work atmosphere had apparently degraded to a product containing one less chlorine atom than the average number of chlorine atoms in the PCB's handled in the factory, and that, conversely, the PCB's found in the workers' blood contained one more chlorine atom than did the PCB's handled in the factory, i.e., if the PCB's found in the workers' blood were Inhaled at the workplace, it can be.surmised that the dl- and trlchloro- biphenyls disappeared rapidly from the body, whereas the tetrachlorobiphenyl was metabolized slowly. 180 HONS 212028 Hasegava at al., 0.973) concluded that blood analysis is a feasible method for determining body burdens of PCBs, and "pro bably more useful than the technique presently used by extracting adipose tissue and using it for PCB;" urinalysis for "PCBs did not appeal to them as a viable monitoring method. They noted, how ever , that there was no correlation between length of employment (and presumably, duration of exposure) and the amount of PCBs accumulated in the blood, l.e., with continued exposure, PCB levels in the blood did not increase linearly; In light of the demonstrated alow excretion rate of PCBs from the body, the authors conjectured that soma different blotrensformative mechanism comas into play after blood PCB levels exceed approximately 1 ppm (perhaps fat storage). This would seem to belle their confidence In blood analysis as a reliable Index of exposure. Kltamura et si., (1973) reported that the mean PCB level In the blood of 10 capacitor factory workers was 0.82 ppm (0.32 2.1 ppm) lsadlately after the cessation of PCB usage, and 0.31 ppm 3 nontha later, when almost all of the subject were observed to have akin slgna. It seems significant that the blood PCB con centrations observed in this study were several fold higher than these seen by Hasegawa at al., (19731. Kltamura at al., (1973) attributed the skin effects to PCB'e accumulated in the workers' bodies during the period of PCB usage. No other abnormalities were reported. 181 MONS 212029 Si(Ol et al>, (1973) caporead the caaa of a houaawlfa in whoa they obsarvad akin effects tavaral yaart after aha had left 6 yaara of amployaant in a capacitor factory. Sha vaa found to hava PCB concantrationa of up co 0.13 ppa in har blood and of 42 ppm In samples har aubcutanaoua adipoaa tiaaua; thia contra*tad aharply with tha findinga of Hara at al (1974) that tha blood PCB lavala of 3 vorkara who raaainad at tha aama factory had decreased to 0.03 ppa and balow, ona yaar aftar tha caaaation of PCB uaaga (froa lavala of 0.05-0.3 ppa during tha time of PCB uaaga (Haaegawa at al., 1973). N.B. Soaa capacitor aanufacturars la tha U.S. uaa epoxide-type alkylating aganta aa additivaa to prolong tha aarvlca Ufa of PCBn. EPA aac. 308 raaponaaa froa tha General Elactric Co., probably contain identifying data. VI. Air and Water Exposure to PCBa Aa indicated previously, nominal hunaa exposure to PCB'a in the U.S. population aay occur froa air and water. Seaplea of aahlaat air ware collected in auburban areaa of Mini, Florida; Jackson, Mlaalaelppl; and Fort Colllna, Colorado. Preliminary reaulta (Kata and Tang, 1975) for aaaplea taken in April, May and June of 1975 ahow that PCBa were preaent at all locatlona. Although the data varied, the average concantrationa at each of tha three locatlona waa approximately 100 nsnograme per cubic aetar. Initial 182 HONS 212030 identification of tha PCBs indicated that they were most com parable to the Aroclor 1254 standard. Dennia (1975) has reported that data gathered from monitoring activities of surface waters and bottom sediments of tha major drainage basins of the United States indicate the widespread occurrence of PCBs in both surface water and bottom deposits. A preliminary assessment of PCI levels shows mean residue levels in water ranged from 0.01 to 0.05 mcg/llter. The 0.05 mcg/llter were found in the South Atlantic Slope and Eastern Gulf of Mexico drainage basin. In general, the lowest PCI residue levels were found in drainage basins west of the Mississippi. Kleinert C1975) has simmerised the work completed by the Wisconsin Department of Natural Resources to identify some PCB sources in the environment in Wisconsin. Effluents from cooling water in foundries contained PCBs ranging from 11.5 to 315 ppb. Investigations revealed the esaon source to be leaking hydraulic fluids containing PCBs which, were used in die cast machines. Effluents from paper mills that recycle wastspapers has maasursable discharges ranging from 0.01 to 25 ppb. Snow samples Odeinert, 1975) were collected early in 1975. Analysis of tha snow melt water from Racine, Kenosha, Madison, and Milwaukee revealed concentrations from 0.17 to 0.24 ppb. The author concludes that these values suggest that fallout of PCBs from the air may be a principal source of PCBs entering the waters HONS 212031 183 of the Stmt*. Basse 0.975) Indicates that similar to the studies In Wisconsin, not all Industrial effluents tasted In Michigan contain aeasureable PCS levels. In testing over 900 Industrial samples, approximately 40 percent contained PCBs above tha 0.1 mcg/liter laboratory sensitivity limit. Twenty-thraa percent of the Indus tries tastad had greater than 0.5 mcg/litar, 18 percent greater than 1 mcg/litar, 6 percant greatar than 10 mcg/liter and 2 percent greatar than 100 mcg/litar. Although much of the PCBs entering municipal waste treatment facilities are removed and become incorporated into tha waste sludge, a sampling of 58 municipal wastewater treatment plant effluents throughout Michigan in 1973 showed on avarsge concentra tion of 0.52 mcg/liter. Concentrations of PCBs are much higher In tha sludges. The avaraga for all plants was 15.6 mg/kg with individual values as high as 350 mg/kg. Sines sewage sludges are cotMonly disposed of by incineration, spreading on agricultural land, or placing in landfills, tha addition of PCBs to tha environ ment is obvious. 711. tesidues of PCB's in Human Tissue and Milk Tabs C1972) u reported that 31.1 parcent of 637 somplas of human adipose tissue collacted from the general population as part of the Monitoring Survey during 1971 vers positive for PCBs in measuraahla amounts. These samples were collacted in 18 states HONS 212032 184 and tha Diatrict of Columbia and positive aaaplaa wcra obtainad from aach of tha sampling stataa and District. Tha parcantaga of PCB lavala rangad from 34.2 parcant non-detected, 33.3 parcant lass than 1 ppm, 27.3 parcant 1-2 ppm and 3.2 parcant greater than 3.2 ppm. Kutz and Straaaman (1973) have described tha raaulta of PCB monitoring during fiscal years 1973 and 1974 in which 33.1 parcant and 40.3 percent, respectively, of tha tissues collected contained levels of 1 ppm or mors of PCBs on a net-weight basis. Analysis of tha tissue revealed that tha compounds found in adipose tissue ware most comparable to those prevalent in Aroclor 1234 and Aroclor 1260. Additional analysis Indiestad that tha most frequently encountered PCB residues ware petsn-, haxa-, and heptachlorobiphenyl compounds. Residues of PCBs have also been detected in a study of human milk collected in Colorado where 8 or 40 samples contained residues ranging from 40 to 100 ppb (Savage at al., 1973). A study of adipose tissue samples collected at autopsy from (grant at al., 1973) indicates that tha majority of rmmmAimmm have adipose tissue residues of 1 to 2 mg/kg of PCB. All adipose tissues had detectable lavala of PCBa and 30 percent of tha samples had PCB residues greater than 1 mg/kg with a range MOHS 212033 185 of 0.11 to 6.60 mg/kg. PCB residues in htonan milk fro* Ontario ruidtnti were found to be approximately 1 mg/kg on * fet basis. MONS 212034 186 HUMAN EXPOSURE TO THE POLYBROKINATED BIPHENYLS I. Introduction and Background The polybrominated biphenyls (PBBa) in this report refers to either Firewater BP-6 or hexabromoblphenyl manufactured by Michigan Chemical Corporation for uae aa a flaae retardant for thermoplastics. Thla product ia a mixture of bromlnated biphenyls with an average bromine content equivalent to about six bromine atoaa per biphenyl molecule. Piremaater BP-6 ia a mixture of the following brominated biphenyls (-1): Tetrabroaobiphenyl 2.OX Pentabromobiphenyl 10.6X Hsxabromobiphenyl 62.8X Heptabromobiphenyl 13.81 Other broaobiphenyla 11.4X The Michigan Chemical Corporation haa atated that to their knowledge Piremaater BP-6 ia the only polybromlnated biphenyl produced in commercial quantity in the U.S. Their production eatimetea are: Tear 1970 1971 1972 1973 1974 (projected) Pounds 20,000 200,000 2.300.000 3.900.000 4.800.000 MOWS 212035 1S7 Flremaster BP-6 hu been used as m flsns retardant in the mmnnfieturs of typewriter, cslculstor and microfilm rsadsr hous ings, radio and TV parts, miscallanaoua small automotive parts and small parts for electrical applications. The use of Flremaster BF-6 has been restricted to those applications where the end-use product la not exposed to either animal feed or food and there is no known use in flame retarding fabrics where human exposure would occur Cl) The ultimate disposition of Flremaster BF-6 upon burial is uncertain. The Michigan Chemical Corporation has stated that in their opinion this material will eventually undergo oxidative/ biological degradation forming carbon dioxide, water end bromide ion CD. II. Human Exposure In October of 1973, adverse health effects were obeerved in cattle in several dairy herd* in the State of Michigan. At that time, the cattle refused to eat manufactured feed; milk production decreased; there was a loss in body weight and the cattle developed abnormal hoof growth with lameness; cattle and swine aborted; and farmers reported the inability to bread heifers after they consumed feed manufactured by Farm Bureau Sarvlcea. A herd of some 100 head of cattle sent to slaughter during this time period, exhibited enlarged livers. HONS 212036 1SS Analysis of staples of the suspected feed by laborstorles of the U.S. Department of Agriculture at Beltsville, Maryland, revealed thet the feed was contaminated with a flame retardant chemical, hexabrominated biphenyl. Subsequent investigation revealed thet the Michigan Chemical Corporation manufactured magnesium oxide, e dairy feed supplement sold under the tradename Nutrlmaster, end a flame retardant, hexabrominated biphenyl, sold under the tradename Firemaster BP-6. Both of these products were distributed In brown paper bags with either the name Nutrlmaster or Firemaster stenciled across the top of the beg. When the top of the bag was torn off and discarded. Identification was essen tially lost. As the result of a mix-up In bags, Firemaster BP-6 was mixed with animal feed In place of ths Nutrlmaster, apparently in the same proportion of use for the Nutrlmaster. It appears that three kinds of feed were initially Involved in this episode with PBB levels as followst Feed No. PBB ' 405 2.4 410 1,790 407 4,300 Semples of milk collected from individual farms soon after the PBB was identified as the contaminant ranged from 2.6 ppm on MOWS 212037 189 m fmt buia to 270.S ppa on a fat basis. Othar products seized and destroyed included: Butter Cheese Canned milk Range 1-2 ppm 1.4 - 15.0 ppm 1.15 - 1.62 ppa It has been estimated that between the onset of contamination in the fall of 1973 and the establishment of the quarantine of affected herds and flocks in the spring of 1974 over 10,000 Michigan residents have been exposed to PBB through the consumption of contaminated milk, meat and other dairy products. A considerable amount of variation in exposure has proabably occurred in both length of exposure and levels of exposure. As a group, the farm family members have been at greatest risk followed by those indi viduals who purchased dairy products from contaminated farms on a regular basis. In order to determine whether or not persona exposed to PBBcontamlnated products had suffered eny acute adverse health effects, the Michigan Department of Public Health undertook a series of studies in the suemer and fall of 1974. Study participants for the exposed group were dairy farm residents from farms which had been quarantined by the Michigan Department of Agriculture. The exposed subjects were limited to thoee who had lived or worked on the quarantined dairy farms for more than six months since May MONS 21203a 190 of 1973. Non-expoaed subjects wara randomly salactad from a list of dmlry producers in tha sane gaographical araa where farms had not baaa quarantinad. A total of 298 parsons wars intarviavad in tha study and physical examinations and/or blood samples wara obtainad for 110 parsons in tha exposed group and 104 parsons from tha control group. Tha Michigan Dapartaant of Public Haalth has raportad (1975) that rcaponsas to a aat of 24 apaciflc madical conditions ravaalad that none of tha haalth complaints occurrad couaiatantly in aithar of tha study groups. Statistical analysis shoved that none of tha llatad complaints was significantly mora fraquant in thoaa parsons with tha hlghast PBB larval* whan compared to other study subjects. Physical examinations of adults and children showed no unusual abnormalities of tha heart, liver, spleen or nervous syatam. Urinalyses and complete blood counts did not reveal a significant excesa of unusual abnormalities related to exposure or PBB levels. These studies showed that blood levels of PBB wars significantly hlg&ar in the study subjects from quarantinad farms as compared to these from the nom-quarantlnad farms; although some subjects from the farms shewed low PBB blood levels (Table 18). Several exposed females delivered normal babies without complication. Tests showed concentrations of PBB in breast milk to be considerably higher than chat found in paired Blood plasma (Table 19). mons 212039 Table 18 Distribution of PBB Blood Levels, Michigan. 1974 ?BB Blood Levels (ppm) Quarantined Farms Adults Children No. z No. % 0 0.002 - 0.019 0.020 - 0.090 0.100 - 0.490 0.500 - 2.260 3 3.7 43 52.4 8 19 23.2 10 11 13.4 3 6 7.3 7 28.6 35.7 10.7 25.0 Nonquarantined Farms Adults Children No. 7. No. 7. 21 28.4 - 52 70.3 29 1 1.4 1 00 0 00 0 96.7 3.3 0 0 TOTAL 82 100.0 28 100.0 74 100.1 30 100.0 Table 19 Comparison of PBB Concentrations In Human Breast Milk and Blood Plasma Data PBB Lavels (ppm) Breast Milk Blood Plasma 12/74 10.800 .082 6/74 22.700 .252 10/74 1.800 .014 3/75 .210 .003 3/75 92.660 1.068 Paired samples of adipose tissue and blood were collected in a group of 13 Individuals entering the hospital for surgical procedures. The concentration of PBB in adipose tissue ranged from 61 to 370 times the PBB value found in blood plasma with an average ratio of 175 to 1 (Table 20). MOHS 212040 192 ifeft.-.. TABLE 20 Compariaon of PBB Concentration* In Human Adipoa* Tlaau* and Blood Plaana Data 11/4 6/74 - 1/75 3/75 3/75 PBB Lavala (ppn) Adipoae Tisaua Blood Plaana .410 .002 1.400 .005 3.000 .012 174.000 1.068 3/75 .248 .002 3/75 .274 .003 4/75 .177 .003 4/75 .152 .003 4/75 1.140 .004 4/75 .808 .004 4/75 .530 .007 4/75 .210 .003 5/75 1.110 .003 Report* of health conplainta auch aa msbneaa, atoaach pain, haadacha, fatigue and anxiety continue to be reported In the varloua neoapepera In Michigan. Report* have aleo appeared In the preaa that aeveral phyalciana In Michigan have reported abnornal liver function teat* In patianta expoaed to PBB. Attaapta to verify the** report* with phyalclana have been unaucceaaful. MOHS 212041 bibliography AD HOC COMMITTEE OF JAPAN PUBLIC HEALTH ASSOCIATION. (1974). Environmental Health Criteria for Polychlorinated Bl- and Terphenyls. Environmental Agency of Japan/WHO Environmental Health Criteria Programe. Aftoamla. J. G., Cullk. 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Allen, J. R., Caratens, L. A., and Barsotti, D. A. (1974), Residual Effects of Short-term, Lov-level Exposure of Non human Primates to Polychlorinated Biphenyls, Toxicol Appl Pharmacol 30:440-431. Allen, J. R., Norbeck, D. H., and 3au, I. C. (1974), Tissue Modifications in Monkeys as Related to Absorption, Distribu tion, and Excretion of Polychlorinated Biphenyls, Arch Environ Contan Toxicol _2:86-93. Allen, J. R. (1975), Response of the Nonhuman Primate to Poly chlorinated Biphenyl Exposure, Fed Proc 34:1673-1679. Allen, J. R., Carstens, L. A., Abrahamson, L. J., and Marlar, R. J. (1975), Responses of Rata and Nonhuman Primates to 2,5,2*,5'-Tetrechloroblphenyl, Env Res 9:265-273. MONS 212042 Alvares, A. P., Bickers, D. R., and Kappaa, A. (1973), Polychlor inated Biphenyls. New 'type of Inducer of Cytochrome P 448 In the Liver, Proc Nat Acad Scl 70:1321-1325. Alvarea, A. P., and Kappaa, A. (1975), Induction of Aryl Hydro carbon Hydroxylass by Polychlorinated Biphenyls In the Ratroplacental Unit and Neonatal Llvera During Lactation, FEBS Lett 50:172-174. Anderbaon, K., Norstrom, A., Rappe, C., Rasmuson, B., and Swahlen, H. (1974), Photochemical Degradation of PCB, PBB Other Flame Retardants, 34th International Congress of Pesticide Chemistry, Helsinki, Finland. Arakl, Y. (1974), Comparison of Hepatic Enzyme-Inducing Activities of Chlorinated Dlbenaofuran and Chlorinated Dlbensodioxln, Fukuoka-Izaku-Zasah1 65:61-64. Armour, J. A. and Burke, J. A, (1970), Method for Separating Polychlorinated Biphenyls from DDT and lta Analogs, JAOAC 53:761-768. Armour, J. A. (1972), Gas Chromatographic Data for Polychlorinated Biphenyl Components in Six Aroclors, J Chromatoar 72:275-282. Armour, J. A. (1973), Quantitative Perchlorlaation of Polychlori nated Biphenyls as a Method for Confirmatory Residue Measure ment and Identification, JAOAC 56:987-993. Armour, J. A. and Burke, J. A. (1971), Behavior of Chlorinated Naphthalenes in Analytical Methods for Organochlorlne Pesti cides and Polychlorinated Biphenyls, JAOAC 54:175-177. Aulerich, R. J., Ringer, R. K., and Iwamoto, S. (1973), Reproductive Failure and Mortality in Mink Fed on Great Lakes Fish, J_Reroda_Fert_Sul 19:363-376. AUSTRALIAN NATIONAL HEALTH AND MEDICAL RESEARCH COUNCIL. (1970), Atmospheric Contaminants NHJMRC, Canberra. Ax, R. L. and Hansen, L. G. (1975), Effects of Purified PCB Analogs on Chicken Reproduction, Poultry Science 34(3);895-90Q. Ax, R. L., Miller, V. L., and Hansen, L. G. (1976), Toxicity Comparisons of TVo Pentachlorobiphenyls. Abstract for presen tation at American Society of Animal Science - Midwestern Section. MONS 212043 Bach*, C. A. and Link, D. J. (1973), A Qualitative Method for Detecting Hydroxylated Metabolite* of Polychlorinated Biphenyls, Bull Environ Contam Toxicol J):315-317. Bailey, S., and Bunyan, P. 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Bull Environ Contam Toxicol 12:318-321. Zltko, V., Butstngor, 0., and Choi, P. M. K, (1972), Contamination of tha lay of Fuady - Gulf of Maine Area with PCBa, PCTe, Chlorinated DIP and DID, Environ Health ParaPact 1;47-30. Zltko, V., Nutsinfer, o., and Safa, S. (1971), Ratantlon Tlmea end Electro* Capture Detector fteaponaea of Some Individual Chlorohlphoayla, frvtroa Cooten and Toxicol 6:160-163. Zltko, lotsinfer, 0., end Safa. S. (1971), Ratantlon Tlmea and Electron-capture Detector Raaponeea of Some Individual Chlorobiphanyla, Bull Environ Contam Toxicol 6:160-163. Zltko, ?., Hutsinger, 0., and Choi, P. M. R. (1974), Determination of Pentachlorophenol and Chlorobiphenylola in Bloloflcal Semplaa, Bull Environ Contam Toxicol 1: 649-633. MOMS 212070 HAtt Chlorinated biphenyl* Appendix A CHU2RIH4TTD ASCT-ATIC CMPOONIS KDTKRCO TO IN CHEJUSW.T RtPOftT STWCTWtt EXTENT 0? CJOOSIHATION ------- TO3TSC--------- x- 1.10 NIB IBIS OP C.'.'Vnr TgaiTOtva ti&T 209 W 7 5 Chlorinated dibenaofunn* Chlorinated dibea*o-p-dlein pentaehlorophenol 135 1-0 IS HONS 212071