Document dYExaYJDmd8G4XZbB1M2xV8LQ

POLYCHLORINATED BIPHENYLS IN GREAT LAKES FISH Toxicological Justification For Lowering The Acceptable Standard To 2 ppm Norman Zimmerman, Ph.D., J.D. Toxic Substance Control Commission 815 Washington Square Building Lansing, Michigan 48909 . March, 1982 * EX P-2696 Page 1 of 57 PCB-ARCH-EXT0371216 TABLE OF CONTENTS Executive Summary Introduction Distribution Rioaccumulation and Metabolism Metabolic Effects of PCBs Toxicity ; Carcinogenesis i Fetotoxic and Teratogenic Effects Synergistic Effects PCBs in Humans Epidemiology Analytical Techniques . Conclusion Tables 1-14 * Figures 1-6 References i 1 3 6 11 . 13 16 18 19 2J 22 23 24 25-38 39-44 45 EX P-2696 Page 2 of 57 PCB-ARCH-EXT0371217 EXECUTIVE SUMMARY 1. In.the 45-year period (1930-1975) approximately 1.4 billion pounds of PCBs were produced in the U.S. of which 1.253 billion went for domestic . usage. This is equivalent to the production of approximately 1 pound of PCS for every second during the 45-year period, (pp 2-3) j 2. PCBs may be contaminated by polychlorinated dibenzofurans (PCDF) and polychlorinated naphthalenes (PCH). These facts emphasize the need to consider possible synergistic effects in setting a standard, (p. ;3) 3. The high stability of PCBs suggest long-time persistence in the environ ment. (p. 2) 4. PCBs are present in many products currently in usage such as capacitors and transformers, (p. 3) ` 5. Distribution of PCBs is worldwide, (pp 4-6) 6. Lake Michigan contamination occurs both from air and surficial sediments, (p. 6 7. .Depending upon_tlie species, these pharmacokinetic parameters such as bioaccumuIation, excretion, and the potential to pass a given bodily . - ----- barrier may vary for different congeners, (pp 6-11) 8. -_ Data, from the Yusho_incideritysuggest;that ^otal PCB may not.be eliminated~\* even years after" exposure^: This mandates setting standardsrat levels to':?"` v ,, minimize exposure, (p. 10) 9. PCBs can affect DNA and carbohydrate, lipid, phospholipid, sterol and porphyrin metabolism.(PP 11-13) 10. The compounds can induce a variety of_enzymes includinq mixed function oxidases, enzymes ..involved in carcinogenesis and.mutagenesis, metabol ism of drugs,steroids and other xenobiotics. (p. 13) 11. PCBs are toxic to specific cells (e.g. lymphocytes, erythrocytes) and can cause histological changes, (pp 15-15) EX P-2696 Page 3 of 57 PCB-ARCH-EXT0371218 v ; 12. PCBs can cause severe pepatotoxic effects in jsertain jspecies^_ They have also been shdwrTto induce goiter and hypothyroidism. Coho salmon from the Great Lakes have an increased incidence of go'iter.(pp PCBs can cause significant problems in the inrnune system. This has shown to occur in humans, (p. 15) PCBs have been shown to be carcinogenic in animals. There is not enou.g evidence to determine carcinogenicity in man, (pp 16-17) 15. PCBs are tumor promoters. (p. 18) 16. PCBs can pass the placental barrier and induce fetotoxic effects, (pp 18-19) 17. PCBs may possibly induce additive or synergistic effects. This should be consiTTeTe'crTn standard setting, (pp 19-20) - 18. PCBs are wjdespread in the human population. All samples of Michigan nursing mothers tested in a study showed the presence of PCBs. This is in contrast to studies done in other areas where a much lower percentage of the polulation was found affected, (pp 21-22) 19. EPA found, in a recent study, that there is a _cgntinual increase in the percentage of the population having PCBs in the blood, (pp 21-22) 20. Epidemiology studies suggest that PCBs may cause various ^diseases in_^^^H humans, (p. 22) **-------~ ~~ 21. - Analytical techniques are of sufficient sophistication to allow application of a 2 ppm standard, (p. 23) EX P-2696 Page 4 of 57 PCB-ARCH-EXT0371219 INTRODUCTION The Toxic Substance Control Commission (TSCC) at its November 19, 1981, meeting recommended that the Michigan standard for polychlorinated bi phenyls (PCBs) in fish be set at 2 parts per million. The text of the motion reads: ' TSCC unanimously recommends that the PCB standard for fish, as regulated by MDA, be established at 2 ppm. TSCC requests MDA to begin the process of setting such a standard for Michigan. The samples of fish should be from skin-on fillets and the chemical analysis should be representative of the total PCB content using the best available technology. . There -has been much'controversy' generated as to whether such a standard is-"justifiedi It is recognized that in the process^ofsetting-standards, the effect on the economy must be balanced against the benefit to the public that can be expected to acrue from the standard. The available scientific data must be evaluated to ascertain the minimum level-to be tolerated as an acceptable public risk. Even for compounds that have been extensively studied, there are usually significant gaps in the available data and some extrapolation is necessary. This report is intended to present a review of the current scientific 1iterature on PCSs. Justification of the position taken by the Com mission is indicated by the data found therein. PCBs are a class of compounds with the general formula shown in Figure 1. They are normally produced by the`chlorination of biphenyl by vaporized anhydrous chlorine with iron filings or ferric chloride catalyst. Toxicity is dependent upon the number of chlorine atoms located in positions 2 to 6 and/or 2' to 6'. The fact that commercial PCBs are a mixture of compounds necessitates consideration of potential synergistic and antagonistic effects. EX P-2696 Page 5 of 57 PCB-ARCH-EXT0371220 The molecular weights vary from 188.7 to 398.5 (Table 1) and there are 209 possible isomers although it has been estimated (1) that approxi mately 100 should be found in commercial products. The pure compounds are transparent, crystaline and solid, while the common commercial forms are liquid. Water solubilities range from 0.007 \ to 5.9 mg/1. Solubilities in most common organic solvents are high. PCBs are thermally and chemically stable, being resistant to oxidation and degradation by other chemical agents. PCBs are fairly stable to oxidation and hydrolysis under moderate con ditions. In contrast PCBs are quite easily photodegraded under various laboratory conditions. The main reaction is a reductive dechlorination, and chlorines in rnzia positions are preferentially lost. The rate of dechlorination is faster in polar solvents like methanol than in hydro carbon solvents. Environmental contamination problems are prevalent and disposal is con founded by PCB stability. Thermal degradation (burning, pyrolysis) seems to b* the most feasible method for the destruction of PCBs. This reaction has been studied by"* several researchers in laboratory experiments and in industrial processes. However, one problem is potential formation of chlorinated dibenzofurans. (Figure 1) . ' .'' '' ;v... . , One primary reason for the magnitude of the PCB problem is the massive t quantities of the compounds that have been produced since their intro ' duction in the late 19_20s by Monsanto Chemical Company. The production peak occurred in the U.S. in 1971 when approximately 85 million pounds of the chemicals were synthesized. Approximately 86% of the total yield was for domestic usage. Production fell until the last year of signifi cant production (1975) when 34.1 million pounds were synthesized of EX P-2696 Page 6 of 57 PCB-ARCH-EXT0371221 which approximately 94 was in^domestic sales. It is estimated (Brinkman and deKok in (2)that in the period 1930-1975 U.S. cumulative production was 1.4 billion pounds (0.63 x 10^ metric tons) with 1.253 being for domestic sales. Jmports were relatively light at approximately 3 million pounds. Monsanto ceased PCB production in mid-1977 and the compounds have not been produced in the United States since that time. The production of PCBs leads to the formation of polychlorinated dibenzofurans (PCDF) (figure 1) and polychlorinated naphthalenes (PCN) (figure as contaminants. Concentrations of up to 33 ppb PCDF have been detected as contaminants of commercial PCBs (Brinkman and deKok in (2),(3-6). The highly toxic 2,3,7,8 tetrachlorodibenzofuran has been found to be one of the contaminants. PCBs have been used for capacitors and transformers and may be components of inks, plasticizers, lubricants, extenders, adhesives, microscope im mersion oils, insulation and a variety of products. PCBs are components of many products currently in use (e.g. transformers, capacitors). DISTRIBUTION IN THE ENVIRONMENT - One methodology for evaluating the potential persistence of a chemical is the determination of the extent to which it has contaminated the environment. It is estimated that over the period 1930-1975 approximately 150 million pounds_cf PCBs were-exported from the United States. Between 1954 and 1972 Japan produced about 130 mill ion .pounds and exported about 11 million pounds. Lesser quantities of PCBs were produced from various sources in Europe (7). The widespread global contamination in many species that has occurred since . that time emphasizes the gravity of the situation. Examples of this contam ination are cited below (See Table 2). Austral i a Mussels (Mytilus edulis) from Port Philip Bay in Victoria, Australia were found to contain PCBs (8). In the Brisbane River estuary, 6 species of fish, 3 species of Crustacea, 1 species of mollusk, mixed species of the polychaetes (marine worms) and 1 species of bird was fc-'d to be contaminated with PCBs (9). EX P-2696 Page 7 of 57 PCB-ARCH-EXT0371222 'Fat tissues from porpoise contained 6.7 ppm PCB (10). Northv/est Atlantic Ocean, Gulf of Mexico Gag (Mycteroperca microlepis), black grouper (M bonaci), red grouper (Epinephelus .morio), red snapper (lutjanus campechanus), king , t mackerel (Scomberomorus cavella) and Spanish mackerel (S. maculatus) were found contaminated (13). Pacific Region - '/ Ten species of small cetaceans (e.g. porpoise) were examined for PCBs in blubber. The location of the sample ranged from the Eastern Pacific and along the coasts of California, Hawaii, Japan and Uruguay. All species tested were positive. One species (Tursiops truncatus) sampled off the California coast was found to have 2,695 ppm. This is believed to be one of the highest concentrations found in tissues of any popula tion of wild animals (14). Europe Sweden Seasonal variations in PCB levels in perch and roach were noted in waters near a nuclear power plant by Hamnefjarden, Sweden (11). It was also found (12) that PCB contamination existed in a variety of marine organisms off the coast of Sweden. .. ' . .. ~ : Finland In a lake area of eastern Finland, perch, roach, vendace, and rainbow trout were found to be contaminated (15). Seven aquatic bird species found in Lake Paijanne in Finland were also found to contain PCBs (16) Baltic herring (Ciupea harenqus) and cod (Gaddus morrhua) found in the Gulfs of Bothnia and Finland also showed positive results. On an archipeligo in southwest Finland arctic tern (Sterna paradisaea) showed PCBs in fat of females in concentrations up to 00.1 ppm. PCBs EX P'2?96 ' Page 8 of 57 PCB-ARCH-EXT0371223 ItalZ A variety of marine animals around the Bay of Naples was shown to be contaminated (18). England . Livers of grey seals taken off Fame Island showed positive results. The presence in the livers of mother/fetus pairs shows that the compounds can undergo transplacental movement (19). Spain Purple herons, spoon ducks, and heron eggs yielded positive results(20). Japan . Aerial samples of urban areas of large cities such as Tokyo may contain up to 2 ppb PCB (21,22). Tokyo Bay surface water was found to contain up to 320 ppt (23)-. Other studies too numerous to mention point out the seriousness of the problem in Japan. United States Boston, Massachusetts Air concentrations as great as 7.1 ppt PCB were found (24). t Columbia, South Carolina Air concentrations as great as 4.4 ppt PCB were found (24). Texas Seven species of wintering shorebirds off Corpus Christi were shown to be contaminated with PCBs (25). PCBs at concentrations up to 70 ppt have been found in Galveston Bay (26). EX P-2696 Page 9 of 57 PCB-ARCH-EXT0371224 Utah r ' PCBs were detected in western grebe (Aechmoporus occidental is) eggs in Bear Valley Migratory Bird Refuge (27). Midwest and Michigan As part of the National Pesticide Monitoring Program, it has been noted (28) that highest aerial PCB residues v/ere found in indus trialized areas of the northwest and midwest. In a recent study (29) it was found that average aerial PCB concentration over Lake Michigan was 1 ppt while that from surrounding urban areas was 5 ppt. Surficial sediments collected from the Lake in 1975 showed a mean residue of 9.7 ppt PCB (30). Figure 2 shows the distribution of PCBs in the Lake. The above survey (see Table 2) is not complete. Other studies show PCB concentrations in other species and areas (e.g. 31-33). BIOACCUMULATION AND METABOLISM . A corollary question to distribution is that of biouccvnulation. This . area includes such questions as how much is absorbed, metabolized, excreted, or "passes various body barriers? What is the biological half-life for various organs; the body as-a-whole? ^Unfortunately we do not have a complete pharmacokinetic picture for PCBs.--.Significant. . . . problems arise in interpreting the data avilable. Fo'r example, it was ' found (17) that in arctic terns average concentrations of PCBs in fat for females and males, respectively, were 38.7 and 80.1 ppm. To evaluate the meaning of these results there are a variety of questions that must be answered. For example: Do females have a tendency to bioaccumulate one half as much as males or is some other factor involved? (e.g. Do the females lose PCBs to the eggs and offspring?) Can the figures be extrapolated to the_h.ujE.art_ popul ation?^ Do females Fiave more body fat thereby "diluting the PCBs? Do female terns consume the same diet as males, etc. EX P-2696 Page 10 of 57 PCB-ARCH-FXTn37199c;  . 1 k.., V. , -7- There is significant evidence for species differences. For example, different~bioaccumu1atTorTTa'ctors are indicated for perch, roach, vendace and rainbow trout (15). However, in another report (34) on the specific congener, 2,2',5,5' tetrachlorobiphenyl, it was found that whole body elimination was similar for yellow perch (Ferca flavescens) and rainbow, trout (Salmo Gairdneiri). The tissue distribution was quite different, however. In perch the viscera and carcass were the main distribution sites. Skeletal muscle, skin and scales were minor sites. In the trout, skeletal muscle and carcass were major sites while viscera and skin were minor sites. This study points up the need for standard edible portion to be based upon fillet plus skin-on sampling. In another study (35) ~~ 3,3' ,4,4'tetrachlorobiphenyl was examined for excretion rates in rats and rhesus monkeys. Distribution and clearance was similar in male'and female rats (contra (17) in terns.) Monkeys clear the compound more slowly. The difficulty of the problem associated with absorption of different congeners is pointed up by a recent study (36). Six hexachlorobiphenyl congeners were fed to rats, guinea pigs, rabbits, Japanese quail, and trout. Total levels for fat after 29 days were, in descending order found in rat, rabbit, guinea pig, trout and Japanese quan. The results suggest that the extent of ortho substitution affects the level of retention. Rabbits and guinea pigs have greater retention of *PCBs with 0, 1 and 2 ortho chloro substituents. Quail retain only the 3,4,5,3',4',5'- iscmer while no retention preferences appear to exist for rat and trout. The authors' results are shown in.Table 3. Other authors (37) have reported contradictory results (in rats) claiming that absorption effi ciencies decreased as the number of chlorines substituted in biphenyls increased. They claim that absorption depends upon molecular size. Others (38) tested absorption of polychlorinated dibenzcfurans in monkey and rat livers. They found that livers of both species favored absorption of the 2,3,4,7,8 compound. This is contrary to the previous authors' assertations regarding absorption and molecular size. "" EX P-2696 Page 11 of 57 PCB-ARCH-EXT0371 Still other authors (39) contrasted elimination of 4,4' dichlorobiphen.yl in beagle dogs and cynomolous monkeys (Macaca fascicularis). The elimina tion rate in the dog_is_9 times greater than in the monkey. In anes thetized dogs 33% of the dose was excreted into the bile in 2 hours. In the monkey the amount of excretion is 0.4%. A better understanding of PCB metabolism should help answer significant questions on health effects of the compounds. Three key questions are of interest: 1. Since routes of metabolism depend upon species, v/hat are the differences in metabolism in species such as dog, rat, monkey and man? 2. What structure-activity relationships exist for metabolism? 3. What congeners are present in man. Lake Michigan fish? While a thorough discussion of these problems is beyond the scope of this report, several recent findings are presented. Humans In a recent study (40) on humans exposed to PCBs, the following was discovered with respect to PCBs found in plasma and adipose tissue. -: . a) the major congeners hacL.chlorines in the 4 and..4.' positions.. b) congeners with chlorines on the 2,4, and 2',4' and/or 3,4 and ; 3,4' positions were present in higher concentrations than in commercial mixtures. c) congeners with unsubstituted 3,4 positions were found in lower concentrations. EX P-2696 Page 12 of 57 PCB-ARCH-EXT0371227 v. O -9- It should be noted that other authors found that the simplest inducer of cytochrome P-448 was 3,4,3',4: tetrachlorobiphenyl (41, 96). Chlorines at both ortho positions (e.g. 2,4,5,2',4' ,5'hexachlorobiphenyl) causes induction of cytochrome P-450. These cytochromes can also be induced by 3,4 substitution on cne-ring (42, 92) (e.g. 2,4,5,3',4'-pentachlorobiphenyl).. Figures 3 and 4 and Table 4 (from 40) show the differences in an adipose sample of a PCB-exposed worker and a commercial Aroclor. The signifinance of this work is that, man appears to preferentially biocon cent rat ePCBstliatcaninduce cytochromes. Dog It has been shown (43) that the dog can excrete 70" of administered 2,3,6,2',3,,6' hexachl orobiphenyl in 3 days. The study on humans does not show the presence of this congener in plasma or adipose tissue. Monkey . In this same study it was shown that after 15 days, monkeys were found to have excreted only 61" of 2,3,6,2',3',6' hexachlorobiphenyl. The elimination rate constants for dog were three to four times greater than those obtained for the monkey. ' A further discussion of these problems is given in (2). The results of pharmacokinetic and structure activity relationships known to date is sum marized in Tables 5 and 6. The data to date suggest the following conclusions: 1._ The literature available is insufficimt-ta.. define pharmaco kinetic parameters either for different species or congeners. ` 2. Depending upon the species, these parameters may vary for dif ferent congeners. * 3. The parameters for a given congener may vary for different species. 4. With this paucity of data, a worst^case analysis must be taken. While the bioaccumulation studies performed to date on animals cannot provide enough information to define the extent of the problem in man, EX P-2696 Page 13 of 57 PCB-ARCH-EXT0371228 several studies based upon one incident suggest the need <or a "worst case" analysis. , In western Japan in 1968 a mass poisoning occurred due to the ingestion of rice oil contaminated by PCBs that had been a component of heating exchange oil that had leaked into the food. Hayabuchi et al (44) measured blood levels of PCBs 5-8 years after consumption of the oil (Table 7). The authors claim a positive correlation between amount of oil consumed and PCB in the blood. Levels as high as 39 ppb were found and the mean appears to increase as the time from the date of the incident progresses (e.g. The mean for males and females studied in 1976 is 10.5 ppb as contrasted to a mean in 1973 of 7.1 ppb. This represents an increase of approximately 48%.) Considering the fat:blood ratio of lipid-soluble compounds, one would expect PCB concentrations to be considerably higher in adipose tissue. In another article (45), the authors measured PCBs in Yusho,victims, unexposed individuals and a PCB worker. The. results are shown in Table 8. Several inferences may be made from examining this data: 1. Using PCB in liver as a measure (since it is the only parameter provided for all patients) there appears to be some positive correlation between the severity of Yusho observed (and probably the amount of exposure to the PCBs) and the PCB tissue level (Firure 5.) 2. So-called unexposed individuals may have PCB concentrations as high as those exposed. ~ ` 3. No individuals tested were completely free of PCBs.. This suggests that the body cannot totally eliminate PCBs or * .. individuals are constantly being exposed from some external source.-. The data is clearly incomplete and there may be distinct differences in bioaccumulation exposure to high versus low doses. However, the data does prove one point. Since PCB levels were found to be high in patients for as long as nine years after exposure, it i_s_clear tha t_ the_body is less than 100% efficient in removing the pollutants. This maridates standards minimizing human exposure. EX P-2696 Page 14 of 57 PCB-ARCH-EXT0371229 -11- Figure 6 shows the type of metabolites that may be produced by PCBs. It must be emphasized that the type of metabolite(s) produced (if any) depends upon the structure of the given congener and the species tested. Different species can yield different metabolites and there are variations in which congeners can be metabolized. It has been found (46) for various hexachlorobiphenyls that the dog can eliminate the PCBs more rapidly than the mouse, rat or monkey. Species differences have also been shown for 4,4'dichlorobiphenyl (47). Other studies on various congeners emphasize these species differences (43-50). PCBs can undergo -- - * n n i ---- . -- . i ,. --i hydroxylation, dechlorination, and rearrangements. They can form di- hydrodiols, phenols and phenol acetates, methyl ethers and sulfur con taining metabolites. Arene oxides are proposed intermediates in the formation of some metabolites and these compounds are potentially electro philic and they may form covalently-bound substrate macromolecular adducts. Congener binding to DNA and RNA and other macromolecules has been observed (51-61). Various congeners and arene oxides were shown to be capable of causing strand breaks in DNA (62, 63). Structure activity considerations must be examined (64). . METABOLIC EFFECTS OF PCBs PCBs can affect a variety of normal biochemical pathways. Rats given a single dose of PCBs (190 mg/kg) showed alterations in carbohydrate metabolism two days after the dosing (65). Plasma glucose was decreased while blood lactate, acetoacetate and /3 -hydroxybutyrate were increased. Multiple dosing at this level (onde/week for 5 weeks) showed the above effects and increased serum cholesterol. Other authors have shown that PCBs interfere with carbohydrate and lipid metabolism (66-69). In a recent study, it was found that PCBs increase conversion of acetate or glucose to cholesterol (70). Both in vivo and in vitro studies showed the same trend. In a study on a population accidently exposed to PCBs in sewage sludge (71) it was found that PCBs increased plasma trigly ceride levels. In vitro studies of rabbit muscle lactic dehydrogenase showed inhibition by a group of congeners tested. The authors conclude EX P-2696 Page 15 of 57 PCB-ARCH-EXT0371230 that inhibition is proportionate to the total number of ring-chlorines (72). Plasma and liver cholesterol as well as urinary ascorbic acid were increased in rats by PCBs(73,74). The significance of these re sults is yet to be determined. The role of high and low density lipoproteins and cholesterol on cardi ovascular disease is still a matter of controversy. It has been found (75) that PCBs cause increased serum cholesterol and high density lipoprotein in rats. In addition the ratio of low density lipoprotein and very low density lipoprotein to high density lipoprotein has been decreased. Whether PCB can in fact affect the incidence of cardiovascular disease . is still an open question. But data such as that presented here emphasizes the need for more research in this area. PCBs have also been shown to affect other systems. Phospholipid meta bolism can be drastically changed (76, 77). PCBs have been found to - decrease the activity of choline phosphotransferase and increase the activity of choline kinase (78). The potential effects of these actions has not yet been suggested. In another human study on the effects of PCBs in the Yusho incident, it was found (79) that porphyrin metabolism can be influenced. An increased urinary excretion of delta-aminolevulinic acio and uroprophyrin was seen. A recent study (80) details the difficulties inherent in the understand ing of the effect of PCBs on metabolic systems. Broiler chicks were fed either fat from PCB-treated swine or untreated PCB in lard. The'/PCB " .from swine appeared to be a stronger microsomal enzyme inducer although . the untreated PCB was a better inducer of certain specific enzymes. . While specific effects of PCBs on given metabolic pathways remain to be elucidated, the fact that PCBs can affect nucleic acids and carbohydrate, lipid, phospholipid, sterol and porphyrin metabolism suggests that the compounds may exert severe toxicological effects by interference with normal metabolic pathways. This mandates taking a conservative approach to standard setting for PCBs. (Table 9 summarizes known biochemical effects of PCBs.) ' EX P-2696 Page 16 of 57 PCB-ARCH-FXTn.^719^1 PCBs have been shown to be potent inducers of aryl hydrocarbon hydro xylase (AHH), an enzyme involved in carcinogenesis by polycyclic aromatic hydrocarbons (81). PCBs administered in the ppm range to the sheepshead (Archosareus probatoce phalus) caused significant induction of AHH as well as other enzymes (82). PCBs can induce the enzyme in the skin of the neonatal rat (83). They can induce AHH in renal or hepatic but not testicular tissues in the rat (84). A class of enzymes known as microsomal mixed function oxidases (MFO) are known to play a complex role in the processes of carcinogenicity and mutagenicity. In addition, a complex biochemical mechanism known as the microsomal P-450 linked mono-oxygenase system plays an important role in the conversion of chlorinated hydrocarbons to electrophilic agents which may bind with DNA and proteins. This is suggested as a mechanism of carcinogenic action. The complexity of the problem can be seen by examining a study (85) where the major PCBs identified in human breast milk were prepared as a re-constituted breast milk mixture. This mixture was compared with the commercial PCB mixture, Kanechlor 500. The former was much more efficient in the induction of AHH. The authors claim that the result can be explained by preferential biocon centration of the most toxic congeners. Even if this assumption is accepted, there is still no known explanation for the body preferentially concentrating the most toxic congneers. It is important to note that PCBs can induce mixed function oxidases, cytochrome P-450 and a variety of enzymes such as demethylases, deethylases, and transferases (see 80,S2-98). The mixture can occur in such diverse species as rat, mouse, certain fish, swine and drosophila.' The fact that PCBs can induce such a variety of enzymes of which we have limited knowledge with respect to their cellular function and interactions emphasizes the need for a. cautious approach in assessing the possible effects of these compounds on man. ` TOXICITY From the previous discussion on metabolism and enzyme induction by PCBs, it would be expected that PCBs could induce a variety of pathological EX P-2696 Page 17 of 57 PCB-ARCH-EXT0371232 'manifestations. This is, in fact, the case. While many studies have been performed in animals and toxicities vary with species, it appears that monkeys are more sensitive than rats and man is suggested as being the _ most sensitive species (99). Symptomology is often a valid indicator of the capacity~~crF an "agent to produce disease and the Yusho incident provides data on the wide variety of symptoms produced from FCB exposure. ) In Japan (1968), rice oil and dark oil were contaminated with PCBs that leaked from a heat exchanger. 400,000 chickens were killed and 2 million were found ill (100). A variety of symptoms were observed in humans (see Table 10 taken from 101). It is clear that PCBs have widely varied systemic effects. The minimum toxic dose was calculated to be 70 mg/kg/day for three months (102). Another study_put the figure at an average 0.5 g/120 days..(100). Some newborns showed chloracne and skin darkening at birth. It is noted that significant quantities of polychlorinated ' dibenzofurans were found in the rice oil and hence the effects cannot be attributed to PCBs alone. About 1200 persons were affected (102). While the Yusho incident must be kept in perspective because of the contaminants present, it does suggest the need for careful control of PCBs. . A variety of hepatic effects, including lipid accumulation, enlargement, fatty degeneration and necrosis have been reported from PCB exposure . (McConnell in (2) (103-110). In a recent study (111) ifecaca fascicularis monkeys fed 5 mg Kanechlor 400 daily for 20 weeks developed enlarged livers and symptoms resembling Yusho patients. In another study (112) rats fed PCBs (75 ppm) were observed to have focal liver necrosis, atypical centro- lobular regeneration, accumulation of .iron-containing pigment in hepato- . - cytes and kupffer cells and numerous ultrastructural changes observed by .electron microscopy. These changes were similar to those observed in rat livers injured by other heptotoxic agents. The study points out the severe' and varied hepatotoxic effects that can be produced by PCBs. Hepatotoxic agents present cause for grave concern because of the potential for insid ious effects. Keplinger (113) found that Aroclors 1242 and 1254 in con centrations of 100 mg/kg in diet caused, in 18 months, increased liver weight EX P-2696 Page 18 of 57 PCB-ARCH-EXT0371233 V V. -15- down to 1.4 mg/kg body weight. These studies point out the gravity of . hepatotoxic effects of PCBs. The fact that PCBs have shown hepatotoxicity mandates taking a conservative position in PCB regulation. PCBs have been shown to induce goiter and hypothyroidism (115). Coho salmon from the Great Lakes show increased incidence of goiter (116-118). Studies on rats suggest that the toxic effects in thyroid is due to a PCB-induced decrease in serum thyroxin (119). PCBs have also been im plicated in thyroid enlargement in rats. PCBs also have a negative effect on the immune systems of the body (Vos', J.G. et al in (2). Reported effects include small spleens in chickens (120), atrophy of lymphoid tissues in chickens (121), thymic and splenic atrophy (122-126), decreased resistance in ducks to hepatitis, lympho penia (127), decreased numbers of circulating leukocytes and lymphocytes, decrease in antibody titre to tetanus toxoid in guinea pigs (128) and decrease in antibody titre in rabbits (129). Other numerous studies (see Vos, J.G. et al (128),show that PCBs have strong effects on the immune system. In another recent review, a detailed analysis is given . on PCB toxicity to lymphocyte function (130). In addition a recent work where a PCS-exposed population in Taiwan was studied (131) showed multiple defects in the immune system caused by PCBs. The fact that there is prima facie evidence of the potential of PCBs to cause varied immune defects (parameters which are difficult to measure in the human ' population) mandates a conservative approach in standard setting. There is ample evidence that PCBs may also cause histological alterations in many cells. In a recent study (132) it was shown that PCBs were toxic in mouse spleen lymphocytes. The compounds affected the plasma membrane by inhibiting its 5'nucleotidase and ATPase. The authors suggest a general, rather than a specific mechanism based upon lipid solubility. In another study (133) the interaction of PCBs with cellular phospholipid bilayer membranes was studied by utilization of spin labels and electron paramagnetic resonance (EPR). The compounds were shown to affect the fluidity of the membranes. PCBs in mice can affect splenic EX P-2696 Page 19 of 57 PCB-ARCH-EXTT)371934 `lymphocytes , CB cells, T cells, Null cells, and can cause transient splenic lymphocyte depletion (134). The effects of PCBs on lymphocyte function is reviewed (135). PCBs have also been shown to cause anemia suggested to be related to toxic effects in erythropoesis (135, 136). PCBs can also produce skin lesions including erythema, hyperkeratosis, alopecia, acne, blisters and desquamation (137-139). Porphyrins are important to a variety of biochemical systems, one of the most important being the cytochromes. Porphyria is a disease where por phyrin metabolism is adversely affected.' In chronic hepatic porphyria, excess porphyrins are produced and excreted. PCBs are an etiologic agent for this disease. While there is question whether porphyria can be induced in man, it is common in many species (Strik, J.M. et al in (2) (140). CARCINOGENESIS - PCBs have been reported to be sus>ect human carcinogens ( 3) and possess tumor promoting activity (141). The compounds have been found to . cause various liver cancers (142-146). The International Agency for Research on Cancer in evaluating the data on possible carcinogenesis of PCBs concluded as follows: 4. Summary of Data Reported and Evaluation^ 4.1 ExDerimental Data - - Five polychlorinated biphenyl mixtures have been tested in mice and/or rats only by oral administration. Kanechlor 500 and Arocl'or: 1254 are carcinogenic in' mice, and Aroclor 1260 is carcinogenic in rats'; all Subsequent to the finalization of this monograph by the Working Group in October 1977, the Secretariat became aware of a study carried out under the NCI Bioassay Programme (NCI, 1978). Groups of 24 male and 24 female Fischer 344 rats were given Archlor 1254 at concentrations of 25, 50 or 100 mg/kg of diet for 104-105 weeks, when surviving animals were killed. No statistically significant differences between tumour incidences in experimental and control animals were seen. However, a few carcinomas and adenocarcinomas of the gastrointestinal tract were observed in treated animals; no such tumours occurred in controls. Hepatocellular hyperplastic nodules were observed in 11/48, 17/46, 29/48 treated animals, compared with none in controls. EX P-2696 Page 20 of 57 PCR-ARnH.FYTn'?71 r -17- induced benign and malignant liver-cell tumours. In an experiment in rats of only one year's duration, Kanechlor 500, 400, and 300 induced liver lesions described in multiple hyperplastic nodules. 4.2 Human data Human exposure to small amounts of polychlorinated biphenyls is widespread as a result of environmental contamination and the high stability of these compounds. They are commonly found in human tissues. Unusually high levels of exposure to polychlorinated biphenyls have occurred among workers manufacturing or using them and in Japanese who consumed rice oil accidently contaminated with Kanechlor 400. The latter showed acute and chronic toxic effects. , An apparent excess of malignant melanoma has been reported in workers exposed to Aroclor 1254. No melanomas were reported in 9 persons who died from cancer among the 1200 Japanese heavily exposed to Kanechlor 400, but these deaths all occurred within 5^ years of first exposure. Neither the workers exposed occupationally nor the Japanese were exposed solely to polychlorinated biphenyls. 4.3 Evaluation There is experimental evidence of a carcinogenic effect of some polychlorinated biphenyls in rodents. The epidemiological data provide suggestive evidence or a relationship between exposure to polychlorinated biphenyls and the development of malignant- me!anoma. Efforts should be made to obtain both confirmatory experimental and epidemiological evidence; in particular, continuing fn'.lowup of survivors of the Yusho episode is necessary. In the meantime, for practical purposes, polychlorinated biphenyls should be regarded as if they were carcinogenic to humans. Almost without exception, polychlorinated biphenyls contain various levels of polychlorinated dibenzofurans as contaminants, and the poly chlorinated biphenyls responsible for the Yusho episode in Japan were found to contain an unusually high level of polychlorinated dibenzofurans. It is not known if and to what extent polychlorinated dibenzofurans play a role in the observed carcinogenic effects of polychlorinated biphenyls. 'It would appear from the data that PCBs have been shown to be carcinogenic in animals. (Table 11 summarizes the work on carcinogenesis of PCBs in animals.) The question as to whether they are carcinogenic in man is still unanswered. Carcinogenesis is thought to proceed by two processes--initiation and promotion. Initiation is the process whereby an agent provides the stimulus to induce the first step leading to change in the genetic EX P-2696 Page 21 of 57 PCB-ARCH-EXT0371236 material. Promotion is the process whereby initiated cellsrare encour aged or stimulated to evolve into cancgr (153). There is significant evidence that PCBs are promotors. It has been found (141) that PCBs administered after the carcinogen 3'-methyl-4 dimethylaminoazobenzene caused a significantly increased incidence of hepatoma. PCBs administered with or before the agent did not induce these tumors. In another study (154) rats treated with the carcinogen diethylnitrosamine were given commercial % aroclor 1254 and f\roclor 1254 in which contaminating dibenzofurans were . t removed. Both PCB mixtures caused increased hepatocellular carcinomas although the former have shown a higher incidence. This study shows that the PCBs alone can act as promoters. Another recent study ((155) showed that polybrominated biphenyls (PBBs) can promote the action of diethylnitrosamine. The similarity in structure between appropriate cogeners of PCBs and PBBs would lend further credence to the evidence that PCBs are promoters. . FETOTOXIC AND TERATOGENIC EFFECTS PCBs are known to pass the placental barrier. Offsprina of pregnant mice treated with PCBs showed an increase in hepatic DMN demethylase. Similar results were obtained for .mono-o :ygenase activities in fetal livers. PCBs were shown to cause ultrastructural lesions in thyroid follicular cells and a reduct.on of serum levels of thyroid hormones in neonatal rats exposed to PCB in utero and by milk. In addition, rats fed PCBs showed decreased liver size (156). Viable hatch of flounder was decreased at concentrations of PCBs of 120 ppb (157). This work showed that PCBs in the environment can.yfeld fetotoxfc effects. Low-dose fertility effects have been observed in monkeys and mink (106) and PCBs have been shown to pass the placenta in humans (158). In cows exposed to Aroclor 1254, the concentration in the fetal kidney was greater than that found in the mother (159). This same compound admin istered to rabbits showed greater concentrations in fetal livers as compared to that of the mother (160). . EX P-2696 Page 22 of 57 PCB-ARCH-EXT0371237  -19- Pregnant rats fed dichloro and tetrachlorobiphenyls passed these compounds to their offspring. Metabolites were also found in the fetuses (161). That PCBs may affect male reproduction is evidenced by a work (162) describing effects from feeding cpd PCBs at 1, 5, 10, 25 and 50 jug/kg daily for 5.5 months. Testicular abnormalities including disorganization of lobules and spermatogenic elements, inhibition of spermatogenesis, fibrosis of lobule walls, fatty necrosis and in one case, total disintegration of the elements in many lobules was observed. In mink, diets of Aroclor 1242 caused total reproductive failure at concentrations as low as 5 ppm of the diet. Ferrets were found to be more resistant with complete reproductive failure occurring at concentrations as low as 20 ppm of the diet (163). It is suggested that the immune response of offspring of rabbits fed PC3s may be impaired but only at high concentrations (164). PCBs caused cleft palate and other congenital anomalies in the offspring of treated mice (165). Other authors (166,167) suggested PCBs could induce behavioral abnormalities in offspring. SYNERGISTIC EFFECTS PC3s may be contaminated with polychlorinated dibenzofurans (PCDF) and/or polychlorinated naphthalenes (2) (168). While synergistic studies are virtually non-existent, practical experience shows (e.g. see section on Yusho) that a combination of chlorinated hydrocarbons can produce devastatina health effects. The toxicity of PCDFs approach those of the dioxins. \ -- For example, 2,3,7,8-tetrachlorodibenzofuran has an LD^q in guinea pigs of 7 ojg/kg (169) and trichlorodibenzofurans in dosages of 0.5 to 1 mg/kg have caused severe and often lethal liver necrosis in rabbits (170). The con- . taminants originally associated with a given amount of PCB might also be found in the same fish as the PCB. However, the presence of these and toxic dioxins may end up in fish alternatively contaminated with PCBs. In addition, fish may be contaminated with other highly toxic polychlorinated hydrocarbons such as polychlorinated dibenzodioxins such as 2,3,7,8 tetra- chlorodibenzo-p-dioxin. In setting a standard, possible additive and synergistic effects must be considered. In an analysis of Great Lakes fish EX P-2696 Page 23 of 57 PCB-ARCH-EXT0371238 performed in 1978 (169), 77% were found to contain PCBs, 100% contained DDT, and chlordane and hexachlorobenzene were found respectively in 38% and 65% of the fish tested. Chlorobenzene, chlorostyrenes, chlorophenols, and chlorinated aliphatics were also identified. As was previously suggested, because PCBs can apparently promote during carcinogenesis, and act as inducers of enzymes involved in metabolism of other carcinogens (e.g. AKH), their presence along with other toxic agents could be expected to have a more severe effect on the population as con trasted to the other agents alone. In a recent study (127), Kanechlor 400 was tested with and without PCDFs for toxic effects on Macaca fascicularis monkeys. PCBs alone produced loss in body weight, symptomology similar to patients involved in the Yusho incident and enlargement of the liver. The combination produced altera tions in the immune system and ultrastructural alterations in the liver, kidney, and skin. PCBs have also been shown (171) to increase the muta genic effect of other agents in Drosophila. Since synergistic effects have not adequately been determined, it is recommended that fish be measured for: a. total organic chlorine and bromine b. total PCBs and P33s c. total chlorinated dibenzofurans c. total chlorinated dibenzodioxins Standards must be set based upon which combination of b, c, and d are present. For example, the detection of quantities of c and d even slightly above the limit of.detection should either render a decision that the fish are inedible or at least cause the acceptable standard for PCBs to be lowered. EX P-2696 Page 24 of 57 PCB-ARCH-EXT0371239 PCBs IN HUMANS PCSs appear to be present in a large percentage of the human population. It has been found (172) that of 637 human fat samples analyzed at surgery or autopsy, PCB concentrations in%68.9% were less than 1 mg/kg. In 25.9%, the concentration was 1-2 mg/kg and in 5.2%, it was greater than 2 mg/kg. It has also been reported (173) that 43% of blood samples from 723 volun teers showed a mean concentration of PCBs of 0.5 mg/100 ml blood. In setting any standard, the factor of tissue concentration from prior exposure must be considered. ' Another recent work has shown PCBs in all samples in the milk of Michigan mothers tested (174). The samples were collected from 68 of the State's 83 counties. The concentrations ranged from trace amounts to 5.1 ppm. Table 12 shows the levels correlated with the percentage of the population tested. The study shows that 73% of the population has PCBs greater . than 1 ppm in the milk. The authors suggest breast milk monitoring for PCBs in nursing mothers who have had potentially high exposure to PCBs (175). By contrast to the situation in Michigan, in milk samples of women analyzed in British Columbia (176), 2% of the samples were >50 ppb, 4% were between 25 and C ppb, 29% were between 50-5 ppb and 65% of the sample showed <5 ppb. The fact that the sample size (49) was considerably smaller than that taken in Michigan (1,043) must be considered. However even taking this into account it appears that the situation is significantly more serious in Michigan. Studies in Colorado (177) (8 of 39 women had PCBs ranging from 0.04 to 0.1 ppm) and Japan (108) (concentrations i . ranged from 0.1 to 0.7 ppm) also suggested a problem of less gravity than in Michigan. In an EPA study (178) under the National Human Monitoring Program (NHMP) it was found that measurable residue levels of PCBs occur in a large per centage of the general population. The authors found that between 1972 and 1976 there was an increase in the population with > 3 ppm in their adipose tissues, (i.e. From 2.58% to 7.34% for whites and from 7.55% to 16.67% for non whites.) In addition, for this time period there was an increase in total EX P-2696 Page 25 of 57 PCB-ARCH-EXTn371?an population with some detectable level of PCBs. The mean ranged from 84.58% in 1972 to 95.54% in 1976. A representative calculation (Table 13) suggests that a mother with 1.5 ppm in breast milk might be expected to transfer up to 1.5 mg PCB daily to a suckling child. With ingestion of 1 fish con taining 5 ppm PCBs, a conservative approach to standard setting is mandated. EPIDEMIOLOGY Several recent studies suggest that PCBs may cause various diseases in humans. In a recent work (182), it was found that in a cohort of electrical workers with potential for exposure to PCBs, excess mortality was noted for rectal and liver cancer. Although neither was statisticajJX-significant, all cancer mortality was lower than expected. In another study (183), electrfcaT^orkers were found to have total PCB blood concentrations of 80-1319 ppb. In another study on this group (184), the authors noted signi ficant increases in skin diseases and hepatic involvement (e.g. hepatomegaly with altered ccncentratons of liver enzymes). It is suggested that the pathological manifestations associated with fiver can be associated with^^H blood trichlorobiphenyl but not pentachlorobiphenyl levels. The authors suggest that 20% of cases of abnormal liver injury would be found in those with 200 ppb PCB in blood. Other works (185, 186) suggest a relationship between human melanoma and PCB exposure. It has also been found (187) that in a population exposed to low levels of PCBs in sludge, alterations in liver metabolism occur at levels where no overt symptoms were induced. An epidemiology study (188) on a normal population with moderate PCB blood levels suggested a positive correlation between PCB exposure and_hypertension, liver function and blood cAoI^teroTjleyels. Other reviews (Kimbo rough, R.. (chap. 9) and Kuratsune, M. in (2) outline other epidemiology studies . involving PCBs. * ' . While it is difficult to reach conclusions on PCB effects based on the epidemiological evidence available to date, the works discussed above sug gest a causal relationship. Until the questions presented are resolved, it is imperative that all measures to limit human exposure be taken. EX P-2696 Page 26 of 57 PCB-ARCH-EXT0371241 ANALYTICAL TECHNIQUES The methodologies available for measuring PCB concentrations are such as to allow accurrate measurements down to the ppt level. For example, a recent study (189) discussed methodologies where PCB levels in v/aters of the Great Lakes were measured at levels as low as 1 ppt (nanogram/1)(See Table Recent development of methodologies for synthesizing standards (e.g. 190) has made it possible to analyze for specific congeners. PCBs can be measured by gas chromatography (GC) or combined gas chromato graphy/mass spectrometry (GC/MS). For total PCB analysis, if only GC is available, perchlorin ation with antimony pentachloride is advi.s a hip (Spp (191) for review). However, more sophisticated analysis should involve the use of GC/MS (192) although new techniques are continually being developed to improve analysis by. both GC and GC/MS (e.g. 193, 194). General metho dologies for the use of GC/MS for water samples i.s well-documented (e.g. Rappe, C. in (2) (195). Specific techniques have been developed for identi fying any given isomer (e.g.(195). Specific methodologies have been developed for measurement of PCBs in air (197, 198). .. The analysis of PCBS from tissues present special problems that require use of'pre-extraction techniques. A recent study (199) d?>c:ribes the techniques available to solve the problem of varying fat content in analysis of PCBs in human milk. Other authors describe cleanup techniques for fatty materials in vegetable samples (200) and in fat samples from steers (201). New techniques involving PCB measurement in both blood and milk is described (.202). The state of tjeiart has progressed to the point where the techniques are available both for the separation of PCBs from other~chTorinated hydrocarbons It is clear that the methodology exists for accurately measuring PCBs and specific congeners and identifying contaminants and metabolites. Thus the techniques to implement a 2 ppm standard are available. EX P-2696 Page 27 of 57 PCB-ARCH-EXT0371242 CONCLUSION The data available on PCBs is quite detailed. Yet still many unanswered questions remain. However, application of the known information mandates a conservative approach in the seating of PCB standards. It is for this reasonTthat the Toxic Substance Control Commission has recommended a 2 ppm standard. The data presented does not support the maintenance of the current 5 ppm standard. EX P-2696 Page 28~of 57 PCB-ARCH-EXT0371243 -25TABLE 1 N.AME GENERAL INFORMATION ON POLYCHLORINATED BIPHENYLS 1 EMPIRICAL FORMULA ' MOLECULAR WEIGHT NUMBER OF -NUMBER OF POSSIBLE. CHLORINES ISOMERS ' PERCENT CHLORIN! Chlorobiphenyl Dichlorobiphenyl Trichlorobiphenyl Tetrachlorobiphenyl Pentachlorobiphenyl Hexachlorobiphenyl Heptachlorobiphenyl Octachlorobiphenyl Nonachlorobiphenyl Decachlorobiphenyl C12H9C1 C12H8C12 C12H7C13 C12H6C14 C12H5C15 C12H4C16 C12H3C1/ C12H2C18 c12hci9 C12C110 . ,188.7 223.1 257.6 292.0 326.4 360.9 * 395.3 329.7 364.1 398.5 1 3 2 12 3 24 . 4 42 5 46 ' 6 42 7 24 ; 8 12 9 3 10 1 Total:209 18.79 31.77 41.30 48.56 54.30 58.93 62.77 65.98 68.73 71.18 EX P-2696 - Page-29 -of 57 ... .. PCB-ARCH-EXT0371244 V nSLt Z S*r ENVIRONMENTAL CONTAMINATION BY PCBs LOCATION Australia Arctic SAMPLE Mussels Fish ' Crustacea Moll usk Marine Worms Porpoise Fat PCB LEVEL N.A. 6.7 Northwest Atlantic Ocean Gulf of Mexico Fish mean - 0.32 ppm highest - 1.8 ppm Eastern Pacific & coasts of California, Hawaii, Japan and Uruguay Cetaceans (e.g. porpoise) Maximum - 2,695 ppm Sweden Fish Various marine organisms N.A. Finland Fish Birds Birds N.A. Maximum - 5 ppm Maximum - 80.1 ppm Italy . Marine Animals N.A. England Seal s N. r. Spain Birds N.A. Japan United States Boston, Massachusetts Columbia, South Carolina .Corpus Christi, Texas Galveston Bay, Texas Utah Northwest, Midwest Lake Michigan Air Tokyo Bay Surface Water . Maximum - 2 ppb Maximum - 320 ppt Air Maximum - 7.1 ppt Air Maximum - 4.4 ppt Birds Maximum - 6.64 ppm Water N.A. Birds N.A. Fish N.A. Air over lake Average - 1 ppt Air over surroundinq urban areas Average - 5 ppt Surficial Sediments Mean - 9.7 ppt N.A. = Information not available to author. REFERENCE (8, 9) OO) .. (13) (14) (11,12) (15) ' (16) (17) (18) (19) (20) (21,22) (23) '.(24) 24) (25) (26) (27) (28) (29) (25) (30) EX P-2696 Page 30 of 57 PCB-ARCH-EXT0371245 TABLE 3* HCBP LEVELS IN RABBIT, RAT AND GUINEA PIG ADIPOSE TISSUE, TROUT AND JAPANESE QUAIL CARCASSES 29 DAYS AFTER ADMINISTRATION OF THE ISOMER MIX ISOMER TISSUE LEVELS (ppm) ( Rabbit3 fat Rat^ fat Guinea Japanese quail d Trout g fat carcass Carcass 2,2',4,4' ,6,6'-HCBP + 0.302 0.056 2,2' ,4,4' ,5'6-HCBP * 0.357 0.077 2,2',4,4' ,5,5'-HCBP + 2.043 0.655 + 2`,3,4,4' ,5,5'-HCBP . 2.103 0.500 3,3' ,4,4' ,5,5'-HCBP + 2.030 0.495 1.253 T 0.460 + 2.003 0.847 3.053 + 0.855 + 1.433 0.681 0.529 + 0.222 0.120 + 0.020 0.074 + 0.004 + 0.933 0.004 + 2.108 0.230 1.500 + 0.010 ND NO ND ND 0.215 - 0.010 + 0.612 0.148 0.830 + 0.174 + 0.559 0.131 0.462 + 0.001 0.553 0.009 3C5, b4, , C2n , d5r and, e3,, speci.es per group. ^ND, non-detectable. ' ' ' NOTE: Numbering System is in error but as reported in original article. Dosage: 5 mg/kg/29 days. *From: Sparling, J. et al (36). EX P-2696 Page 31 of 57 PCB-ARCH-EXT0371246 ANALYSIS OF PC3 CONGENERS IN ADIPOSE TISSUE ANu ..-LASKA AMONG CAPACITOR MANUFACTURE WORKERS *7 rom: Wolfe, M.(40). Concentration in *. . *: . Peak . Structure Assignment adipose (ug/g) Median Range Plasma (ng/ml) Median Range AdiposePlasma Parti tit 1A 4,4'- , 0.08 0.002-0.2 1 2,4,4'-(2,5,4'-) 9 0.5-183 - 2 2,5,2',5'- . 0.5 0.07-3 3 2,4,2',5'- 0.3 0.06-4 4 2,4,2',4`- 0.6 0.09-11 5 . 2,3,2',5'- 0.5 0.1-5 5 A (2,3,4,2'-;3,5,3',5'-) . 0.3 0.04-3 6 2,4,5,4'- 7 . .*0.5-36 7 2,5,3',4'- 8 2,4,3',4'- Q 2,3,6,2',5'- 10 2,3,5,2',5'- 0.4 0.1-2 2 0.2-18 0.2 0.05-2 0.1 0.08-0.3 n 2,4,5,2',5'- . 0.1 0.07-0.5 12 2,4,5,2',4'- 1 0.07-5 13 (2,3,4,4',-; 3,'4,3',5'-) 0.9 0.03-6 14 : 2,3,6,2',3'- 15 ' 2,3,5,2',3'- ' 15 2,4,5,2',3'- p,p'-DDE 2 0.3-8 17 2,3,4,2',5'- 0.2 0.08-0.5 18 2,3,6,3',4'-(3,4,3',4'-) 0.1 0.07-0.3 15 20 21 . 22 2,3,5,3',4'2,4,5,3',4'2,3,5,2',4',5'. 2,4,5,2',4',5'- 0:1 0.09-0.2 2 0.09-5 0.1 C.05-0.7 1 0.09-4 23 2,3,4,3',4'- 1 0.2-4 24 2,3,4,2',3',5'- ' 0.1 0.05-0.3 25A (2,3,4,5,3',5'-) 0.3 0.05-1 25 . 2,3,4,2',4',5'- 1 0.06-4 26 2,3,5,6,2',4',5'- 0.2 0.05-0.7 27 2,3,4,6,2',4`,5'- 0.1 0.06-0.2 28 (2,4,5,3*,4',5'-) 0.1 0.05-0.4 25 . 2,3,4,2',3',4'- 0.07 0.05-0.2 30 2,3,4,5,2',3',6'- Q.09 0.05-0.2 31 -2,3,5,6,2',3',4'-' 0.07 ' ' 0.05-0.3 32 2,3,4,5,3' ,4'- 0.4 0.08-2 33 2,3,4,5,2',4',5'- 0.6 0.05-3 34 2,3,5,6,2',3' ,4' ,5'- 0.2 0.05-0.5 35 2,3,4,5,2',3',4'- 0.2 0.08-1 36 2,3,4,2',3', 4', 6`- 0.1 0.05-2 37 2,3,4,5,2',3',4',5'- 0.1 0.7 27 4 2 3 3 1 26 2 7 1 0.9 0.8 4 3 0.03-2 1-850 0.5-23 0.2-42 0.2-24 0.4-55 0.2-42 0.7-180 0.2-27 . 0.3-113 0.3-9 0.6-2 0.4-6 0.4-12 0.7-29 90 200 80 90 ` 310 80 60 200 70 160 60 50 240 190 7 1 0.5 ' 0.6 8 0.7 5 2 0.4 1 4 0.6 0.3 0.5 0.8 . 0.3 0.4 2 3 1 1 0.7 _ ' , 0.6-35 0.6-2 0.3-3 0.4-1 0.7-41 0.2-2 0.7-12 0.6-20 0.2-2 0.4-4 0.5-17 0.2-2 0.2-0.8 0.2-2 0.7-0.8 0.2-0.3 0.2-0.5 0.2-5 0.6-8 0.2-2 0.5-3 0.2-1 170 180 300 270 130 360 110 370 160 120 - '310 270 260 360 Note: Concentrations were calculated from response factors of standards for peaks 1(1); 11(2-20, 23); peak 22 (21,22,24-36). Peaks 14-15 were negligible in fat and plasma. Parti; was derived.from regression coefficient (slope) of adipose tissue vs plasma levels. Peaks v' no partition values had fewer than four cases with detectable values in both adipose and pi; or the correlation was not statistically significant (peaks 17.18.24). The number of c2s- = with detectable peaks in both adipose and Dlasma were(peak,n): 1A,14; 1,25; 2,16; 3,9; 4. r 5A,15; 6,26; 7,7; 8,20; 9,8^ 10,3; 11,9; 12,23; 13,17; 20,^2; 21,(6; 22,23; 23,19; 25A,l/; 25,23; 26,16; 27,7; 28,10; 32,15; 33,22, 34,4; 35 EX P-2696 Page 32 of 57 PCB-ARCH-EXT0371247 -zy- TABLE 5 CONGENER PHARMACOKINETICS OF PCBs SPECIES HALF-LIFE OR OTHER PARAMETERS (see notes) REFERENC 4,4- Dog Monkey 1 day 21 days (39) 2,3,6,2',3',6'2,4,6,2' ,4' ,6`- 2,4,6,Z' ,4' ,5*- Dog i Monkey Rat Trout Rabbit Guinea Pig Japanese Quail Rat Trout 1 day 9 days 1.253-RR3* 0.612(48.8%)RRa 0.302(24%)RRa 0.120(9.5%)RRa 0.000(0%)RRa 2.003-RR3* 0.838(41.872)RRa (43) (36) (36) Rabbit 0.357(17.8S)-RRa Guinea Pig 0.074(3.7%)RRa Japanese Quail 0.000(0%)RRa . 2,4,5,21,4' ,5'- Rat Rabbit 3.053-RR9* 2.043(66.9%)RRa (36) Guinea Pig 0.933(30.6S)RRa Trout 0.559(18.3S)RRa 3,4,5,2*,4',5'- - Japanese Quail Guinea Pig Rabbit Rat Trout Japanese Quail 0.000(0%)RRa - 2.108-RR3* 2.103(99.8%)RRa 1.433(67.9%)RRa 0.462(21)9%)RRa 0.000(0%)RRa (36) 3,4,5,3*,4*,5*- Rabbit Guinea'Pig Trout 2.030-RR3* 1.500(73.4%)RRa 0.553(27.2)RRa (36) Rat Japanese Quail 0.529(26.1%)RRa - 0.215(10.6%)RRa Decachlorobiphenyl Cow 4- Rat 4,4'- Rat 2,4,5,2' ,5'- Rat 2,4,5,2' ,4',5`- ' Rat OS-Body Retention 48.2-ED0 24.6-EDb 20.5-EDb 1.3-EDb (205 (205 (a) Relative retention-the level in ppm is given for the species with the asterisk (this is th highest value reported). The values reported for the other species are those percentaoes of PC3 retained as compared to the reference animal which had the highest retention level" (mean values are used). . ' ' EX P-2696 -- Page 33 of 57 PCB-ARCH-EXT0371248 STRUCTURE-ACTIVITY RELATIONSHIPS FOR PCBs STRUCTURAL CHARACTERISTIC SPECIES EFFECT OBSERVED REFERENCE Increased chlorination (1 to 6) Availability of unsubstituted adjacent carbon atoms Availability of unsubstituted adjacent carbon atoms Chlorines in 4,4' position Comparison of chlorines in 4,4' position and those in unsubstituted 3,4 positions Chlorines in 2,4 and/or 3,4 positions on both rings Comparison of 2,4 and/or 3,4 positions on both rings and those with unsubstituted 3,4 positions regardless of chlorination. rat rat mouse man man man man increased body retention increased metabolism and excretion increased metabolism and excretion tendency to bioaccumulate latter has less tendency to bioaccumulate tendency to bioaccumulate (206) (207-20S (210,211 (40) i i former has greater tendency to bioaccumulate. Comparison of 2,4 substitution on both rings and 3,4 substitution on either ring 2 or more ortho chlorines man mammals 3,3',4,4',5,5' hexachlorobiphenyl mammals 3,4 or 4,5 positions unsubstituted mammals 2,3 or 5,6 positions unsubstituted 9 all 3,4 or 4,5 vicinal positions mammals ' " blocked No adjacent pairs of unsubstituted positions mammals- Substitution at both para positions, 2 ortho positions, at least 2 meta . positions and containing mammals 2,3,4 substitution pattern 3,3',4,4' tetra and 3,3'*4,4*,5,5' hexa congeners mammals former has greater tendency to bioaccumulate exclusively cytochrome P-450.(not P-448) inducer .exclusively cytochrome P-448 (not P-450) inducer most easily metabolized and eliminated slowly metabolized (91) eliminated most slowly - i mixed microsomal enzyme induction i (SB) mixed microsomal enzyme induction (212,21 EX P-2696 Page 34 of 57 PCB-ARCH-EXT0371249 O UD 1 c TABLE 7** MEAN AMD RANGE OF AGE, TOTAL AMOUNT OF OIL CONSUMED, THE AMOUNT OF OIL CONSUMED PER KG PER DAY, AND THE BLOOD PCB CONCENTRATION OF YUSHO PATIENTS IN 1973-1976 Year 1973 1 1974 CO 1 1975 1976 . No. of Sex Patients M 21 F 27 M+F 40 M 14 F 24 M+ F . 38 M 13 F 19 M&F . 32 M 9 F 10 M+ F 19 Age (yr) ------------------------------- Mean Range 35.7 29.3 32.1 30.9 33.4 35.4 44.0 32.7 37.3 50.4 42.2 46.1 6-74 0-60 6-74 0-60 9-58 8-68 10-76 10-59 10-76 23-77 11-77 Oil Consumption Total (ml) -----------------------------Mean* ' Range Daily(ug/kg/day) --------------------------- Mean* Range Blood PCB(ppb) --------------------------- Mean* Range 700 704 746 740 842 806 028 056 045 623 800 751 230-3813 195-3375 195-3375 220-2013 212-3375 212-3375 216-2813 208-3375 216-3375 208-1934 280-3375 280-3375 207 75-608 244 60-061 227 60-861 160 32-608 224 55-861 198 ` 32-861 207 49-351 236 i 55-861 224 ' 49-861 196 75-600 . 218 55-061 207 55-861 6.9 7.2 7.1 0.6 9.1 0.9 0.5 0.2 8.3 9.9 11.5 10.5 2-29 1-39 1-39 5-19 2-31 2-31 3-19 2-37 . 2-37 6-14 4-32 4-32 * Geometric mean ** From Hayabuchi, M. et al (44). O v. EX P-2696 Page 35 of 57 PCB-ARCH-EXT0371250 TABLE 0** INDIVIDUAL PCB AND PCQ LEVELS IN THE TISSUES OF YUSHO VICTIMS, UNEXPOSED INDIVIDUALS AND IN THE MILK FAT OF A WORKER OCCUPATIONALLY EXPOSED TO PCB Sample no. . Age Date of ------------------1--------------------------------- Severity GC Pattern PCB level PCQ level Sex Death of Yusho* Tissue (or fluid) of PCB+ (ppb) (ppb) 583 15868 651 166.34 8 AN-77-34 AN-77-100 2 3 4 Yusho victims 0 M 14.10.68 25 M 9.7.69 4 Intestine Liver Adipose Tissue Liver C 29.7 83.6 C 50.5 393.0 A 5090.7 2400.0' A 225.9 217.5 48 F 29,12.70 1 . Adipose Tissue Liver C 270.5 2.2 C - 7.4 1.2 46 M 16,5.72 72 . M 30'. 4'. 75 3 # Adipose Tissue . A Liver A Intestine A Liver A 6091.3 68.5 3471.8 114.4 1444.0 143.5 1770.0 51.7 59 M 17,3.77 2 Intestine Liver A 3630.3 1125.0 A 68.4 27.0 69 M 4,9.77 1 Intestine Liver B 1273.4 24.5 C 17.7 1.0 Unexposed Individuals 46 M 19.9.78 * Adipose Tissue Liver 1477.8 2.7' 71.1 0.8 70 M 20,9.78 54 F ''*21,9.78 35 F . 24.9.78 Adipose Tissue Liver Liver Adipose Tissue 530.1 2.7 17.9 0.6 22.0 0.7 248.0 1.3 PCB Worker 37 F ..... Mother's Milk Fat .. .. 6241.1 0.3 *Grade of Severity of skin lesions; grades increase with increasing severity **Kashimoto, T. et al {46) j v' ex P-2696 Hhe PCH GC patterns arc classified.according to Masuda et a'l (1974). A is peculiar to Yusho patients, pai $6W$f t0 A & ------ -- 1 ' <- n r<--... </li n if.'__________________ 81 Pr.R-ARri-l-PYTn'57'IOC'l TABLE 9 BIOCHEMICAL EFFECTS OF PCBs EFFECT REFERENCE Alterations in carbohydrate metabolism Alterations in carbohydrate and lipid metabolism Increased conversion of acetate or glucose to cholesterol Increased plasma triglycerides Inhibition of lactic dehydrogenase (65) (66-69) (70) (71) (72) Increase in plasma and liver cholesterol and urinary ascorbic acid Increase in serum cholesterol and high density 1 ipoprotein;:Decrease ratio of low density and very low density lipoprotein to high nsity lipoprotein (73,74) (75) eration in phospholipid metabolism (76,77) ase activity of choline phosphotransferase and increase ity of choline kinase \ce porphyrin metabolism (increase urinary excretion of deltaulinic acid and uroprophyrin) (78) (79) nzyme induction ' . (80-98) EX P-2696 Page _ 37 of 57 PCB-ARCH-EXT0371252 TABLE 10* PERCENT DISTRIBUTION OF SYMPTOMS OF YUSHO REPORTED BY 189 PATIENTS EXAMINED BEFORE OCTOBER 31, 1963* SYMPTOMS MALES FEMALES ) (N = 89) (N = 10( Dark brown pigmentation of nails 83.1 75.0 Distinctive hair follicles 64.0 56.0 Increased sweating at palms 50.6 55.0 Acnelike skin eruptions . 87.6 82.0 Red plaques on limbs 20.2 16.0 Itching 42.7 52.0^ Pigmentation of skin 75.3 72.0 Swelling of 1imbs 20.2 41.0 Stiffened soles in feet & palms of hands 24.7 29.0 Pigmented mucous membrane 56.2 47.0 Increased eye discharge 88.8 83.0 Hyperemia of conjunctiva 70.8 71.0 Transient visual disturbance 56.2 . 55.0 Jaundice 11.2 11.0 Swelling of upper eyelids 71.9 74.0 Feeling of weakness 58.4 52.0 Numbness in limbs 32.6 39.0 Fever 16.9 19.0 Hearing difficulties 18.0 19.0 Spasm of limbs 7.9 8.0 Headache Vomiting . . 30.3 23.6" 39.0 " `28.0 Diarrea 19.1 17.0 *From: Kuratsune, M. (101) EX P-2696 Page 38 of 57 PCB-ARCH-EXT0371253 TA3LEJ1 ' STUDIES ON CARCINOGENICITY OF PCSs S-ECIES TEST SU3STANCE TOTAL CARCINOGENESIS TESTING cd mice (n) Kanechlor 300,400,500 Selves mice (m) ArocTor 1254 Srerman rats(f,m) Arcolor 1260 Arcolor 1254 Dcnyru rats(f,a) Kanechlor 400 . Vi star rats(m) Kanechlor 300, 400, 500 Sherman rats(t) Arch!or 1260 ROUTES OF ADM!HI STRATTON DOSAGE INCIDENCE REFERENCE oral oral oral oral oral 500 mg/kg/32 weeks 7/12 - liver nodules 5/12 - hepatocellular carcinoma 300 mg/kg diet/11 mos 9/22 - hepatoma 22/22 - adenoflbrosis 0, 20, 100, 500 or 1000 mg/kg diet/8 mos 2/10 (m) adenoflbrosis 1000 mg/kg 1/10 (f) adenofibrosls 100 mg/kg 1/10 (f j adenoflbrosis 500 mg/kg 4/10 (fj adenoflbrosis 1000 mg/kg 0, 20, 100 or 500 mg/kg diet/8 mos 10/10(m) adenoflbrosis 500 mg/kg 9/10 (f) adenoflbrosis 500 mg/kg 1/10 (m) adenoflbrosis 100 mg/kg 7/10 (f) adenoflbrosis 100 mg/kg 047) (148) 049) 049) 38.5 - 616 mg/kg diet/400 days 1000 mg/kg diet N 500 mg/kg diet 100 mg/kg diet m 100 mg/kg diet 21-22 mos 6/10 (f) adenomas 050) 2/15 - cholangioflbrosis-Kanechlor 300 2/10 - cholangiofibrosis-Kanechlor 400 4/13 - cholangiofibrosis-Kanechlor 500 5/13 - hyperplasia-Kanechlor 500 3/10 - hyperplasia-Kanechlor 400 5/10 - hyperplasia-Kanechlor 500 1/22 - hyperplasia-Kanechlor 300 2/16 - hyperplasia-Kanechlor 400 3/25 - hyperplasia-Kanechlor 500 26/184 - hepatocellular carcinoma 144/184 - neoplastic nodules (152) mal es r females SECIES E. PROMOTION TEST SU3STANCE Donryu rats . Kanechlor 400 ROUTES OF ADMINISTRATION oral Sprague Dawley Aroc1or U5. oril INITIATOR DOSAGE INCIDENCE 3!-methyl-4 dimethyl ami noazobenzene (3`HeDAB) 600 mg/kg diet/6 mo. 3`Me0AS, then Q/11 400 mg/kg diet Kanechlor3'" 041) diethylnitrosamine (DEN) 65 jug/kg diet/5 wk DEN, 21/33(63.6-)a then 100 mg/kg diet/ . (154) 18 wk. Aroclor 27/32(84.45) i?C3s previously treated to remove polychlorinated dibenzofurans. 'PC5s untreated; contaminated wnth polychlorinated dibenzofurans. EX P-2696 Page 39 of 57 PCB-ARCH-EXT0371254 TABLE 12* LEVEL PCBs IN MOTHERS MILK IN MICHIGAN PERCENTAGE OF TEST GROUP WITHIN LEVEL > 3 ppm * 6.14 . 2-3 ppm 17.4 1-2 ppm . ' -49.5 < 1 ppm 26.96 *From Wickizer, T.M: et al (174). t ' - * ` '* * - -- * EX P-2696 Page 40 of 57 PCB-ARCH-EXT0371255 TADLE 13 REPRESENTATIVE CALCULATIONS OF POSSIBLE PCB INGESTION FROM FISH The calculations in this table are mere approximations only suggested for use in most general terms. A. Maximum intake at one meal for fish contaminated at 5 ppm PCB ) Fish at 5 ppm =5 mg/1000 gm Fish = 5 mg/2 lb fish = 2.5 mg/1 lb fish (average serving) Maximum dosage = 1.25 mg/meal assuming even distri bution throughout fish and that H lb fish is edible (179). B. Reproductive effects in monkeys (taken from (102). Reproductive effects in monkeys seen at dosages of 0.12 mg/kg/day. 110 lb female = 50 kg,thus 0.1 mg/kg/day = total body intake of approxi- .' mately 5 mg/day. This is 4 times the average intake expected from ingestion of one fish contaminated at the 5 ppm level. C. Yusho* . Sevare effects were seen in individuals receiving 0.5 g over 120 days. Assuming approximately equal exposure per day, the total dosage per day is 4^.16 mg. * -- This is approximately 3.3 times the dosage expected from ingestion of one fish contaminated at the 5 ppm level. D. Potential dosage to children Assume an average Michigan mother has 1-2 ppm (avg. 1.5 ppm) PCB in milk. i Assuming further a six-month baby weighs . 7.5 kg (180) and daily milk intake is 500 ml (180) to 1000 ml (181), the total daily PCB intake based upon the Michigan study would be 0.75 to 1.5 mg/day. With the uncertainty of the burden an infant might receive from the mother who ingested 1.25 mg/day (i.e. eating 1 fish in a given day),.the setting of a standard should be based upon a most conservative approach. * The fact that the PCB's were contaminated by chlorodibenzofurans must be "considered in interpretation of results. EX P-2696 Page 41 of 57 PCB-ARCH-EXT0371256 TABLE 14* POLYCHLORINATED BIPHENYLS IN THE GREAT LAKES WATER (nq/1) Sediments (nq/ml) Superior Michigan Ontario Huron Erie 5.0 31.0 1 -3 5 - 7 27 30 38 9-33 75 - 250 43 - 240 *From National Academy of Sciences (189). i EX P-2696 Page 42 of 57 PCB-ARCH-EXT0371257 FIGURE I STRUCTURES OF RELATED POLYCHLOR1NED AROMATIC HYDROCARBONS Polychlorinated Biphenyls (PCB) 3 2 2' 3' Polychlorinated Dibenzofurans (PCDF) 1 6 t EX P-2696 Page 43 of 57 PCB-ARCH-EXT0371258 - - IN LAKE MICHIGAN SEDIMENTS '*w-- FIGURE 2 Distribution of polychlorinated biphenyls in surficial sediments of Lake Michigan (0-3 cm). Arrows indicate sources and are discussed in text. From Frank, R. et_ aj_. (30) EX P-2696 Page 44 of 57 PCB-ARCH-EXT0371259 o O -41- , FIGURE 3* Typical chromatogram from HRGC analysis of adipose sample from a PCB-exposed worker with current high exposure to Aroclor 1016 and with HPCB (peaks 20 to 33) typical of the general population. 10 20 30 40 50 MINUTES * From Wolfe, M. et al (40). EX P-2696 Page 45 of 57 PCB-ARCH-EXT0371260 FIGURE 4* Typical chromatogram From HRGC analysis of standard PCB solution (Aroclor 1016). Peak numbers refer to structure designation in Table 4. MINUTES *From Wolfe, M. et al (40). EX P-2696 Page 46 of 57 PCB-ARCH-EXT0371261 I I 00 FIGURE 5. . Correlation between severity of Yusho and PCB^Concentration in Liver-extrapolation from Kashimoto, T. et al `(45). V.. P c B o 3 L E V E L rvoIir j | (ppb) J f ) o 2 3 SEVERITY OF YUSHO ) i EX P-2696 Page 47 of 57 PCB-ARCH-EXT0371262 FIGURE 6 A summary of the in vivo PCB metabolites n--r OH H !QH)aclx-i dihydrodiols phenol dechlorination acetates and and methyl ethers rearrangement products sulfur- microbial containing --"'enradation products products EX P-2696 Page 48 of 57 PCB-ARCH-EXT0371263 c -45- 1. Environ. Health Perspect., 24_, 131 (1973). 2. Kimbrough, R.D.; Halogenated Biphenyls, Terphenyls, Napthalenes, Dibenzodioxins and Related Products; Elsevier, New York (1980). > ` 3. International Agency for Research on Cancer (IARC); Polychlorinated and Polybrominated Biphenyls, IARC Monographs on the Evaluation of. the Carcinogenic Risk of Chemicals to Humans; #18, Lyon (1978). 47 Vos, J.6. et al; Food Cosmet. Toxicol.; 625 (1970). 5. Bowes, G.W. et al; Nature, 256, 305 (1975). 6. Nagayama, J. et al; Bull. Environ. Contam. Toxicol.; 15_, 9 (1976). 7. Miyata, H. et al; J. Food Hyg. Soc; Japan, 1J7.> 434 (1976). 8. Richardson,. B.J. et al; Biogeochem. Ancient Mod. Environ.; Proc. Int. Symp., 4th, 423 (1980). . * 9. Shaw, 6.R. et al; Chemosphere, 9(12), 731 (1980). 10. Clauson, J. et al; Bull. Environ. Contam. Tox.; 12(5), 529 (1974). . IT. Edgren, M. et al; Chemosphere; 10(5), 447 (1981). 12. Jehsen, S. et al; Nature; 224, 247 (1969). 13. Stout, V.F.; Fish Bull.; 78(1), 51 (1980). 14. O'Shea, T.J. et al; Pestic. Monit. J.; 14(2), 35 (1980). 15. Pyysalo, H. et al; Chemosphere; 10(8), 865 (1981). 16. ' Paasivirta, J. et al; Chemosphere; 10(7), 787 (1981). . 17. Lemmetyinen, R. et al; Ann. Zool. Fenn; 17(3), 141 (1980). 18. Camoni, I. et al; Ann First Super Sanita; 16(4), 645 (1980). 19. . Donkin, P. et al; Sci. Total Environ.; 19(2), 121 (1981). 20. Baluga, G. et al; Bull. Env. Contam. Tox.; 1_7, 603 (1977). 21. Tatsukawa, R.; In:PCB Poisoning and Pollution, Chicuchi, I. p. 147, Tokyo, Kodansha Ltd. (1976). 22. Tatsukawa, K. et al; Shoko No Kagaku; 55 (1972). 23. Fukushima, M.; M.S. Thesis, College of Agriculture, Ehime Univ., Japan (1974). EX P-2696 Page 49 of 57 PCB-ARCH-EXT0371264 Bidleman, T.S.; Atmos Env.; 15(4), 619 (1981). White, D.H. et al; Pestic. Monit. J.; 14(2), 58 (1980). Hurray, H.E. et al; Bull. Environ. Contain. Toxicol.; 26_, 769 (1981). Lindvall, M.L. et al; Pestic. Monit. J.; 14(3), 108 (1980). Schmitt, C.J. et al; Pestic. Monit. J.; 14(4)> 136 (1981). Doskey, P.V. et al; J. Great Lakes Res.; 7(1), 15 (1931). Frank, R. et al; J. Great Lakes Res.; 7(1), 42 (1981). . Kerkhoff, M: et al; Sci. Total. Environ.; 19(1), 41 (1981). Lawler, G.C. et al; Amoco Cadiz: Consequences Pollut. Accid. Hydrocarbures, Acetes Colloq. Int.; p. 573 (1981). Ross, P.F. Set al;.Bull. Environ. Contam. Guiney, P.D. et al; Arch. Env. Contam. Toxicol.; 9(6), 667 (1980). Abdel-Hamid, F.H. et al; J. Toxicol. Environ. Health; 7(2), 181 (1981). Sparling, J. et al; Toxicol. Lett.; 7_, 23 (1980). Tanabe, S. et al; Agric. Biol. Chem.; 45(3), 717 (1981). r Kuroki, H. et al; Food Cosmet. Toxicol; 18_, 387 (1980). Sipes, I.G.; Toxicol. Appl. Pharmacol.;554 (1980). Wolff, M.S. et al; Toxicol. Appl. Pharmacol.; 62_, 294 (1981). Poland, A. et al; Ann. N.Y. Acad. Sci.; 320, 214 (1979). . . ..Yoshihara,. S. et al; Chemosphere; 8, 531 (1979) . \ . Sipes, I.C.;'Toxicol'. Appl. Pharmacol.; 62, 317 (1982). . Hayabuchi, H. et al; Food Cosmet. Toxicol.; 1_9, 53 (1981). Kashimoto, T. et al; Food Cosmet. Toxicol.; 1_9, 335 (1981). Sipes, I.G. et al; Society of Toxicology; Eighth Annual Meeting, New Orleans, March (1979). ' Sipes, I.G. et al; Toxicol.. Appl. Pharmacol.; 55_, 554 (1980). Muhlebach, S. et al; Xenobiot.; T21_, 249 (1981 ).. EX P-2696 Page 50 of 57 PCB-ARCH-EXT0371265 f i . r t -47- 49. Matthews, H.B. et al; Toxicol. Appl. Pharmacol.; 53, 377 (1980). 50. Matthews, H.B. et al; Drug Metabol. Dis'p.; 3_, 211 (1975). 51. Morales, N.M. et al; Toxicol. App. Pharmacol.; 48^, 397 (1979). ) 52. Hsu, I.C. et al; Proc. Soc. Exp. Biol. Med.; 150, 185 (1975). 53. Seymour, J.L. et al; Proc. Soc. Exp. Biol. Med.; 152, 621 (1976). 54*. Hesse, S. et al; Chem. Biol. Interact.; 20_, 355 (1978). 55. Wyndham, C. et al; Biochem.; 1_7, 208 (1978). 55. Wyndham, C. et al; Res. Commun. Chem. Pathol. Pharmacol.; 1_5, 563 (1976). 57. Hesse, S. et al; Biochem. Pharmacol.; 26_, 2043 (1977). 58. Shimada,T.; Bull. Env. Contam. Toxicol.; 16., 25 (1976). . 59. Ghiasudden, S.M. et al; Toxicol. Appl. Pharmacol.; 36., 187 (1976). 60. Shimada, T. et al; Biochem. Pharmacol.; 27_, 585 (1978). 61. Hargraves, W.A. et al; Res. Commun. Chem. Pathol. Pharmacol.; 25^, ,33 (1979). 62. Stadnicki, S. et al; Res. Comm. Chem. Pathol. Pharmacol.; 24, 313 (1979). 63. Wong, A. et al; Res. Commun. Chem. Pathol. Pharmacol.; 24_, 543 (1979). 64. Kennedy, M.W. et al; Biochem. Pharmacol.; 22.> "577 (1981). 65. Garthoff, L.H.; Toxicol. Appl. Pharmacol.; 60., 33 (1981). 66. Mehlman, M.A. et al; Toxicol. Appl. Pharmacol.; 27., 300 (1979). 67. Messner, B. et al; Nature; 263, 599 (1976). 68. Bernardier, C.D. et al; J..Toxicol. Environ. Health; T_, 91 (1975). . 69. Garthoff, L.H. et al; J. Toxicol. Environ. Health; 3., 769 (1977). 70. Yagi, N. et al; Env. Res.; 22, 139 (1980). 71. Baker, E.L. et al; Amer. J. Epid.; 112(4), 553 (1980).. 72. Nishizawa, M. et al; Bull. Environ. Contam. Toxicol.; 25(4), 630 (1980). 73. Kato, N. et al; J. Nutr.; 3(1), 123 (1981). . 74. Tang, T. et al; J. Nutr. Sci. Vitaminol.; 26(6), 629 (1980). EX P-2696 Page 51 of 57 PCB-ARCH-EXT0371266 -40- r l. < 75. Kato, N. et al; Nutr. Rep. Int.; 23(5), 825 (1981). 76. Ishidate, K. et al; Biochem. Phannacol.; 25_, 1255 (1976). 77. Ishidate, K. et al; Biochem. Pharmacol.; 27_, 2595 (1978). 78. Ishidate, K. et al; Biochem. Biophys. Acta.; 620, 49 (1980). 79. Chang, K. et al; Res. Commun. Chem. Pathol. Pharmacol.; 30(3), 547 (1980). 80. Hansen, L.G. et al; Toxicology; 21 (3), 203 (1981 ). . 81. Gelboin, H.U. et al; Proc. Nat. Acad. Sci. U.S.A.; 64, 1188 (1969). 82. James, M.O. et al; Chem. Biol. Interact.; 36(2), 229 (1981). Mukthar, H. et al; Drug Metabol. Dispos.; 9(4), 311 (1981). Kluwe, W.M. et al; Toxicology; 20, 259 (1981). Parkinson, A. .et al; Biochem. Biophys. Res. Comm.; 96(2), 882 (1980).' "85. Addison, R.F. et al; Environ. Pol. Series A; 25(3), 211 (1981). 87. Inoue, K. et al; Toxicol. Appl. Pharmacol.; 59(3), 540 (1981). 88. Parkinson, A. et al; Chem. Biol. Interactions; 35_, 1 (1981). 89. McKinney, J.D. et al; Chem. Biol. Interactions; 33_, 271 (1981). 90. Haellstrom, I. et al; Chem. Biol. Interactions; 34_, 145 (1981). 91. Albro, P.W. et al; Chem. Biol. Interactions; 34_, 373 (1981). 92. Hara, E. et al; Cancer Res.; 41(1), 253 (1981). 93. 94.' 95.' Carlson, G.P.; Life Sci.; 27(17), 1571 (1980). Narbonne, J.F.; Toxicol. Appl. Pharmacol.; 56(1), 1 (1980)..' Shimada, T. et al; Toxicol. Appl. Pharmacol.; 55(3), 490 (1980). 96. Ueng, T.H. et al; 0. Biol. Chem.; 256(14), 7536 (1981). 97. Ahotupa, M.; Biochem. Pharmacol.; 30(13), 1866 (1981 ). 98. Jannetti, R.A. et al; J. Nat. Cancer Inst.; 67(2), 461 (1981 ). 99. Polychlorinated Biphenyls and Terphyenyls; World Health Organization, Geneva (1976). EX P-2696 Page 52 of 57 PCB-ARCH-EXT0371267 L -49- V../ r 100. Kohanana, K. et al; Nat. Inst. Animal Health Quarterly; 9_, 213 (1979). 101. Kuratsune, M.; Environ. Health Perspect.; 1_, 119 (1972). 102. 103. Isono, W.; Private Communication. } . Adamson, I.Y.R. et al; Arch. Environ. Health; 27_, 69 (1973). 104. Nishizumi, M.; Toxicol. Appl. Pharmacol.; 45_, 209 (1978). 105. Vos, J.G. et al; Toxicol. Appl. Pharmacol.; 1_7> ^56 (1970). 106. Brucker, J.V. et al; Toxicol. Appl. Pharmacol.; 28_, 189 (1974). - 107. Koller, L.D. et al; Am. J. Pathol.; 70, 363 (1973). . 108. Zinkl, J.G. et al; Vet. Pathol.; 12, 63 (1975). . 109. Kimbrough, R.D. et al; Arch. Environ. Health; 25_, 354 (1972).- 110. Hinton, D.E. et al;Virchows Arch. B. Cell Pathol.; 27, 279 (1978). 111. Hori, Si et al; J. Pharmacobio-Dyn; 4(5), 5-65 (1981). 112. Oonsson, H.T. et al; Arch. Environ. Contam. Toxicol.; 1_0, 171 (1981). 113. Keplinger, M.L. et al; Toxicol. Appl. Pharmacol.; 1_9_, 402 (1971). 114. Kimbrough, R.D. et al; J. Nat. Cancer Inst.; 55_, 1453 (1979). 115. Barsano, C.P.; Environ. Health Perspect.; 38., 71 (1981). 116. Black, J.J. et al; J. Nat. Cancer Inst.; 53_, 725 (1974). 117. ' Sonstegard, R. et al; Cancer Res.; 36., 4467 (1976). 118. Moccia, R.D. et al; Science; 198, 425 (1977). . 119. Bastomsky, C.H. et al; Endocrinology; 98, 1309 (1976). 120. Flick, D.F. et al; Poultry Sci.; 44_, 1460 (1965). '121. Vos, J.G. et al; Food Cosmet. Toxicol.; 8., 625 (1970). 122. Vos, J.G. et al; Toxicol. Appl. Pharmacol.; 19., 617 {1971). 123. Iatropolous, M.J. et al; Toxicol. Appl. Pharmacol.; 41_, 85 (1977). 124. Harris, S.J. et al; Poultry Sci,; 55., 1933 (1976). 125. Hansen, L.E. et al; Am. J. Vet. Res.; 36(1), 23 (1975). EX P-2696 --Page 53 -of-57- PCB-ARCH-EXT0371268 I i 1 126. Street, J.L. et al; Toxicol. Appl. Pharmacol.; 32(3), 587 (1975). 127. Friend, M. et al; Science; 170., 1314 (1970). 128. Vos, J.G. et al; Sci. Total Environ.; 1_, 289 (1972). 129. Roller, L.D. et al; An*. J. Vet. Res.; 34, 1605 (1973). 130. Silkworth, J.B. et al; Environ. Health Perspect.; 3, 105 (1981). 131. Chang, K.J. et al; Immunologic Evaluation of Patients with Polychlorinated Biphenyl Poisoning; Society of Toxicology Meeting; Boston; Feb (1932). 132. 133. Sharom, F.J. et al; Biochem. Pharmacol.; 29(24), 3311 (1980). Reich, T. et al; Environ. Sci. Health, Part A; Al6(1), 65 (1981). 134. Carter, J.W. et al; Immunopharmacology; 2(4), 341 (1980). 135. Silkworth, J.B. et al; Environ. Health Perspect.; 39^ 105 (1980). 136. McConnell, E.E. et al; Ann. N.Y. Acad. Sci.; 320, 138 (1979). 137. Meies, J.W. et al; J. Ainer. Med. Assn.; 154, 1417 (1954). 138. Summary of Health Effects of PCBs, Chemical Manufacturing Assn., Washington (1981). ' 139. Miller, J.W.; U.S. Public Health Dept.; 59, 1085 (1974). 140. Zitko, V. et al; Environ. Health Perspect.; 47 (1972). 141. Kimura, N.T. et al; Z. Krebsforsch; 87_, 257 (1976). 142. Ito, N. et al; Gann; 65_, 545 (1974). 143. Kimborough, R.D. et al; J. Nat. Cancer Inst.; 55_, 1453 (1975). 144. . Nagasaki, H. et al; Gann; 63, 805 (1972). . . . 145. Kimura, N.T. et al; Gann; 64_, 105 (1973). ' .' " * 146. Kimborough, R.D. et al; J. Nat. Cancer Inst.; 53_, 547 (1974). 147. Ito, N. et al; J. Nat. Cancer Inst.; 51_, 1637 (1973). 148. Kimborough, R. et al ; J. Nat. Cancer Inst.; 53_, 547 (1974). 149. Kimborough, R. et al; Arch. Ind. Health; 25_, 354 (1972). 150. Kimura, N.T. et al; Gann; 64_, 105 (1973). EX P-2696 Page 54 of 57 PCB-ARCH-EXT0371269 151. 152. 153. 154. 155. 156. 157. 158. 159. 160. 176. I to, N. et al; Gann; 65_, 545 (1974). Kimborough, R.-et al; J. Nat. Cancer Inst.; 5^_, 1453 (1975). Farber, E.; Biochem. Biophys. Acta.; 605, 149 (1980). Preston, B.D. et al; J. Nat. Cancer Inst.; 66(3), 509 (1981). , Sleight, S.D.; Society of Toxicology Meeting; Boston; Feb. (1981). Collins, W.T. Jr. et al; Am. 0. Pathol.; 99(1 ), 125 (1980). Von Westernhagen, H. et al; Aquatic Toxicol.; 1(2), 85 (1980). Sukamoto, H. et al; Fukuoka Acta. Med.; 60, 496 (1969). Platonow, N.S. et al; Vet. Rec.; 88,, 109 (1971). Grant, D.L. et al; Environ. Physiol.; X, 61 (1971). Toeroeck, P. et al; Arch. Environ. Contam. Toxicol.; 10(3), 289 (1981 ). Sangalang, G.B. at al; Arch. Environ. Contam. Toxicol.; 10(5), 617 (1981). Bleavins, M.R. et al; Arch. Environ. Contam. Toxicol.; 9(5), 627 (1980). Thomas, P.T. et al; Drug Chem. Toxicol.; 3(2), 173 (1980). Watanabe, M. et al; Toxicology; 19(1), 49 (1981 ). Storm, J.E. et al; Neurobehav. Toxicol. Teratol.; 3(1), 5 (1981). Bowman, R.E. et al; Neurobehav. Toxicol. Teratol.; 3(1), 15 (1981). Veith, G.D. et al; Pestic. Monit. J.; 15(1), 1 (1981). Moore, J.A.; Proc. Nat. Conf. on Polychlorinated Biphenyls; Chicago, Nov. (1975). Bauer, H. et al; Arch. 'Gewerbepathol .-Gewerbehyg;. 1_8, 538 (1961). Haellstroem, I. et al; Chem. Biol. Interact.; 34(2), 129 (1931 ). Yobs, A.R. et al; Env. Health Perspec.; X* 79 (1972). Finklea, J. et al; Am. J. Pub. Health; X> 79 (1972). Wickizer, T.M. et al; Am. J. Pub. Health; 71 (2), 132 (1981 )., Wickizer, T.M. et al; Pediat.; 68(3), 411 (1981 ).. Mes, J. et al; Can. Plains Proc. Workshop Pestic. Residue Anal. Ista. ; 9, 99 (1980). EX P-2696 Page 55 of 57 PCB-ARCH-EXT0371270 177. Savage, F.D. et al; Bull. Environ. Contam. Toxicol.; , 222 (1973)."'. Lucas, R.M. et al; FCB Residue Levels in Human Adipose Tissue. ^ ._ 178. Statistical Evaluation by Racial GroupingrEPA Report 560/13-79-1015 (1980). 179. City Fish Market; Personnel' Communication. 180. Schlantia, R.J. et al; 0. Pediat.; 96(4), 679 (1980). 181. Harger, J.R.E. TSCC Report; PCBs and General Low Level Contaminants in Fish (1980). 182. Brown, D.P. et al; Arch. Env. Health; 36(3), 120 (1981). 183. Maroni, M. et al; Br. J. Med.; 38, 49 (1981). 184. Maroni, M. et al; Br. J. Med.; 38, 55 (1981). 185. Bahn, A.K. et al; New Eng. 0. Med.; 296, 108 (1977). 186. Bahn, A.-K. et al; New Eng. J. Med.; 295, 450 (1976). ' Baker, E.L. et al; Am. J. Epid.; 112(4), 553 (1980)-. 188. Kreiss, K. et al; J. Amer. Med.. Assn.; 245(24), 2505 (1981). 189. Polychlorinated Biphenyls; National Academy of Sciences, Washington (1979). 190. Mull in, M. et al; J. Anal. Toxicol.; 5(3), 138 (1981). 191. Hutzinger, D.; The Chemistry of PCBs,; CRC Press, Cleveland (1974). 192. Duinker, J.C. et al; Bull. Environ. Contam. Toxu'ol.; 25(6), 956 (1980). 193. Nowicki, H.G. et-al; J. Assoc. Off. Anal. Chem.; 64(1), 16 (1981). 194. DeKok, A. et al; Int. Environ. Anal. Chem.; 9(4), 301 (1981). 195. Millar, J.D. et al; Anal^ Chem.; 53(2), 214 (1981). . - 196. Tindall, G.W. et al; J. Chromatog.; 196, 109 (1980). A - ' - J. 197. Andrew, A.W. et al; Atmospheric Chemistry of PCBs and PAHs; 9_; EPA Report 905/4-79-029-1 (1980). 198. Jackson, M.D. et al; ASIM Spec. Tech. Pub.; 721; 1SS Sampling Anal. Toxic Org. Atmos. (1980). 199. Mes, J. et al; Chemosphere; 9_, 763 (1980). 200. Tuinstra, L.G.M. et al; J. Chromatog.; 204, 413 (1981). EX P-2696 Page 56 of 57 PCB-ARCH-EXT0371271 9 201. 202. 203. 204. 205. 206. 207. 208. 209. 210. 211. 212. 213. Ross, P.F. et al; Bull. Environ. Contam. Toxicol.; 26(4), 485 (1981). Mes, J. et al; Int. J. Environ. Anal. Chem.; 8(2), 89 (1980). Krupcik, J. et al; J. Cnrcmatog.; 191, 207 (1980). Albro, P.W. et al; J. Chromatog.; 197, 155 (1980). Platanow, N.S. et al; Can. J. Comp. Med.; 39_, 104(1975). Mehendale, H.M. et al; Toxicol. Appl. Pharmacol.; 36_,369 (1970). Lutz, R.J. et al; Drug Metab. Disp.; 5, 386 (1977). Tuey, D.B. et al; Drug Metab. Disp.; 5_,. 444 (1977). Peterson', R.E. et al; Toxicol. Appl. Pharmacol.; 38_, 609 (1976). Gage, J.C. et al; Toxicol. Appl. Pharmacol.; 436, 555 (1976). Brandt, I. et al; Pharmacol. Toxicol.; 40_, 1 (1977). Tierney, B. et al; Arch. Biochem. Biophys.; 200, 513 (1980). Poland, A. et al; J. Biol. Chem.; 251, 4936 (1976). EX P-2696 Page 57 of 57 PCB-ARCH-EXT0371272