Document M43n0wppo3o3wrOknKaDDYeJa

Scientific Communications Polychlorinated Biphenyls Storage, Distribution, Excretion, and Recovery: Liver Morphology After Prolonged Dietary Ingestion Virlyn W. Burse; Renata D. Kimbrough, MD; Ellen C. Villanueva, MS; Ralph W. Jennings; Ralph E. Linder; G. Wayne Sovocool, PhD Rais were fed 100 ppm of Aroclor 1242 been restricted to closed system. A (0.0 to 3.1 mg/kg/day) or 100 ppm Aro- new PCB, Aroclor 1016, which is a clor 1011 (6.0 lo 3.5 mg/kg/day). Plasma, kldnayt, urine, brain, liver, and adipose (Issue were analysed for polychlorinated biphenyl (PCS) residues at 0.5, one, two, lour, six. eight, and ten months of expo* sure. In additional groups fed the experl* mental dial for six months, PCS tissue levels were determined two, lour, five, end six months slier the exposure to PCBe good dielectric and heat transfer fluid has been proposed. It contains a lower percentage of the higher PCBs and should prove to be leas persistent in the environment. The higher PCBs appear to be more persistent, as evi denced by mass spectral residue anal ysis of environmental samples.4 * It is the purpose of this paper to re port results of feeding studies con approached in two and roaehed in 4 ducted in rats with Aroclor 1016 and months. Ultle PCS-derived materiel was excreted In urine. After discontinuing PCS exposure, Aredor 1010 wee eliminated more rapidly Irom the organs than Aroclor 1242. Mas* suraMe resIdus levels were still present after live (Aroclor 1016) and tlx (Aroclor 1242) months of recovery. Microscopic Aroclor 1242. Aroclor 1242 in its gen eral make-up is very similar to Aro clor 1016, it only lacks the higher PCBs (Papageorge, written commu nication, June 1973). The Aroclors dif fer in the percentages of Axed chlo rine, 43% 05% (1242) vs 41.3% examination ol Me liver showed enlarged 0.6% (1016). Aroclor 1242 contains 9% liver cells with vacuolated eytoplaem and biphenyls with five or more chlorines inclusions. and Aroclor 1016 contains 1% bi phenyls with five or more chlorines. The use and replaceability of poly chlorinated biphenyl (PCBs) has recently been reviewed.' Because of MATERIALS AND METHOD their extensive use in the past and Aroclors 1242 and 1016 were of electrical their slow biodegradability. PCBs are gr-de, lot No. KB45-41S and KB-06-756. served (Table 1): one control and four rata fed Aroclor 1016 or 1242 were killed two weeks after onset of the experiment. Four additional rats of each experimental group were killed one and two months after onset of the experiment The further killing of four experimental rals for each PCB mix ture aa 'veil as four controls that served for both experimental groups were made four, six, eight, and ten months after onset of the experiment. After six months feeding, the experimental diet* were discontinued on 18 randomly selected rata for each PCB mixture. These rata were fed plain chow and four for each PCB mixture were killed together with four control rata at two, four, and five or six months (Table 2) of re covery. Twenty-four hours prior to each killing the rats were housed in individual metabolism cages and urine was collected. During the 24-hour pe -iod prior to death, none of the rats in the study was fed a fortiffed diet Oxalated bio id was obtained by cardiae puncture from rats anesthetized with ether. The animals were killed by cxsanguination and liver, fat. kidney, and brain tissues were collected. The liver was weighed and the tissues were froten for chemical analysis. Liver tissue from the animals killed at four months and at sub sequent killings were also Axed in buffered 4% formaldehyde for miemeropie examina now widely distributed in the envi rtf *ectively. The PCBs were fed in parallel tion, and stained with humatoxyiin-eosin. ronment''-1 Recently the use of these material in the United States has exj .irimenU to male Sherman strain rats. 61t > 73 days old. A total of 158 rata was di vide d into two groups of 27 and two groups Liver tissue at four months of exposure to Aroclor 1016 and at aix months of exposure to Aroclor 1242 together with an equal of 5:1 rats each according to a table of ran number of controls for electron micro " Zi Submitted lor publication Moy 10, 1974; ac cepted July 17. Pram the Environment*! Protection Agency, Chembleo Toxicology Laboratory. Chambloe, Go (Mr. Hurra, Dr, Kimbrough. Mo. Villanueva, Mr. Jennings, and Mr. Under), and the NERC. EPA, Reraareh Triangle Park, NC (Dr. Sovooooi). Mr. Burra is currently with the US Consumer Prodett Safely Commiraion, Atlanta; Dr. Kimbrough U with the Center for Disease Control. Toxivtogy Branch, Atlanta; Ms. Villanueva is with Ihs Coca-Cola Export Corporation, Atlanta; Mr Jennlnp Is with the Environmental Protection Agency, Region IV, AUanta; and Mr. Linder ie with the Pesticides and Toxic Substance* Effects laboratory, NERC, EPA, Research Triangle Park. NC. Reprint request* to the Toxicology Branch, Center for Dinara Control, 1600 Clifton Rd, NE, Atlanta, GA 30333 (Dr. Kimbrough). dom numbers. The rats were group caged and weighed weekly. Their food consump tion was measured, one week prior to each scheduled killing. The groups of 52 rats were fed the experimental diets that con tainer 100 ppm of Aroclor 1016 or Aroclor 1242. '-lie control rata were fed ground laborator- chow ad libitum. The PCBa were incorporated into the diets by dissolving them in ether and pour ing this solution onto 100 gm of corn starch. The ether was evaporated and the PCB cornstarch mixture diluted with in creasing amounts of ground chow in an electric mixer. The following killing schedule was ob scopic examination was Axed in chilled, buffered 5% glutaraldehyde for two hours, postffxed in 1% osmium tetraoxide, dehy drated through changes of alcohol and propylene oxide, and embedded in resin. Sections were cut with a glass knife, stained with lead citrate and uranyl ace tate, and examined with an electron micro scope. The analytical methods used for the de termination of PCB-derived materials have been described previously." BrieAy, 100 to 200 mg of fat, brain, kidney, and liver or 1 to 2 ml of plasma were extracted with hexane. Brain was extracted with acetone, the solution evaporated to dryness Correction Arch Environ Health/Vol 29. Dec 1974 Polychlorinated Slphenyls/Burse et al 301 MCNS 062723 ti c as <Qr> d Fig 1 -Chromatograms ot Aroclor 1242: 3.66 ng, an extract of urine: 4.S mg, and adipose tissue: 0.027 mg from a male rat led a diet containing 100 ppm Aroclor 1242 for eight months. 3* Table 1.--Mean Concentration and Standard Error of PC8*0erived Materials in Tissues ol Four Rats Per Killing Fed 100 ppm Aroclor 1242or 1016 Months on Diet 0.5 10 Ptaeme Are 1241 0.081 0.000 0.180 0.030 0.310 0.057 0 219 0.028 0.528 0.150 0.240 0.052 0.172 0.014 Are 1018 0.182 0.017 0.t75 0.028 0.331 0.010 0.372 0.040 0384 0.024 0 394 0.014 0.334 0.040 Kidneys Are 1242 .Are tetf 0.66 0.63 0.15 0.08 0.51 1.58 0.13 0.11 1.44 2.74 0.18 0.19 1.64 4.78 0.27 0.24 1.89 3.21 0.47 048 2.14 2.07 0.41 0.06 3.13 1.64 0.56 0.38 Urine* Are 1242 <0.02 <0.01 <0.02 0.030 0.02 0.028 0.000 0.168 0.04 <0.01 Aro 1016 0.075 0.042 0.008 0.000 0.162 0.036 0.259 0.063 0.275 0.112 0.057 0.017 0.021 0.000 Brain Aro 1242 <0.25 <0.25 <0.25 0.56 0.08 1.69 0.25 1.63 0.24 2.05 0.08 Are 1016 1 52 0.75 3.05 0.13 3.40 0.13 3.18 0.23 2.96 0.44 2.64 0.36 2.70 0 36 Liver Are 1242 0.55 0.08 t .46 0.10 3.24 0.30 3 94 0.72 421 0.57 4.56 0.81 6.39 0.68 Aro 1016 1.46 0.24 3.05 0.26 3.66 0.38 740 0.79 786 0.75 5.97 0.94 6.70 0.34 Adipose Tissue Aro 1242 35 2.4 65 10 92 1.5 115 it 110 13 143 8 133 15 Aro 1016 69 4 103 8 182 12 214 18 238 9 221 10 168 9 * Confirmed qualitatively by Couison eonduotemetry. Table 2.--Mean Concentration and Standard Error of PCB-Derlved Matsrlal In Tissues of Four Rats Two, Four. Five, or Six Months After They Had Been Fed 100 ppm Aroclor 1242 or 1016 for Six Months Months Seeovary 2 5. Plasms Are 1242 0.436 0 091 0.149 0.000 0 122 0.026 ArelOtl 0.194 0.034 0.193 0.050 0.062 0.011 Kidneys Aro 1242 2.07 0.40 1.04 0.27 0.34 0.06 Are 1016 i.oa 0.27 1.S2 0.44 0.44 0.14 Urine Aro 1242 0.02 0.01 <0.01 0.004 0.000 Are 1016 0.033 0.004 0.006 0.002 0.006 0.002 Brain Are 1242 1.06 0 24 1.22 0.53 0.51 0.06 Aro 1016 1.40 0.07 1.35 0.21 0.36 0.06 Liver Ate 1242 4.32 0.72 3.46 1.04 0.97 0.35 Are 1016 2.25 0 32 3.13 0.78 toe 0.09 Adipose Tissue Are 1242 60 3.7 44 9 24 Are 1616 104 13 36 6 21 1 * Experiment had to be prematurely terminated. 302 Arch Environ Health/Vol 29, Dec 1974 Polychlorinated Biphenyls/Burse et al HONS 062724 M ( Fig 2. taimnn Lenff'h xp*ii 6 ny. and l! 10 to mnli'l' volum. lilirn hoxiiHi Snni|< noumin t hr* Uilmn Arch L 100 90 Aroclor 1016 Standard g 60 Urine Adipose Tissue Time, min Fig 2.--Chromatograms of Aroclor 1016: 2.64 rtg, extract of urine: 5.4 mg, and adipose tissue: 0.008 mg Irom a male rat led a diet con taining 100 ppm Aroclor 1016 for eight months. I * Only rats fad Aroclor 1242 were examined microscopically. and the solute taken up In hexane. Urine, t 10 to 20 mi, was extracted at approxi mately pH 7 with hexane. Each sampla in a volume of 0.6 ml wee eluted from a micro silica gel column using s 1:1 bensene- hexane mixture. t Samples were analysed with a tritium l source electron capture detector operated i in the direct current mode el 12 v. Quanti tation of the standard and of the different kinds of samples was based in esch in stance on total response In peak heights. However, as reported more fully below, there were qualitative differencei in the peaks for the standard, the urine, the tis sues collected during dosing, and the tis sues collected after dosing was discontin ued. Under these circumstances exact quantitation is impossible, but the use of total response in peak flights is consid ered to offer the best estimate of quantita tion. RESULTS The rats did not suffer from any overt clinical symptoms of poisoning. During the course of the experiment five rats that were fed Aroclor lOlfi and one rat that was fed Aroclor 1242 Arch Environ Heallh/Vol 29, Dec 1974 Polychlorinated Biphenyls/ Burse el al 303 MGNS 062725 / i Fig 3.--Chromatograms ot adipose tissue: 0.011 mg from a rat fed a diet containing 100 ppm Aroclor 1242 for six months and six months of a control diet: 0.016 mg from a ret fed diet containing 100 ppm Aroclor 1016 for six months and a control diet for five months. (Relative retention times were computed relative to the most intense electron capturing peak present In Aroclor 1018 or 1242. All gat chromatographic columns were composed of 1.5% OV-17/1.95% QF-1 on 100/120 Oat Chrom O.) died. The cause of death was not es tablished. No difference between the experimental and the control rata was observed in the body weight gain and the food consumption. As the rata aged food consumption decreased relative to their body weight. The amount of Aroclor 1242 consumed by the rata ranged from 6.6 mg/kg body weight/day to 3.89 mg/kg body woight/day, and the amount of Arodor 1016 consumed by the rats ranged from 8.9 mg/kg body weight/day to 3.6 mg/kg body weight/day. At autopsy the organs appeared grossly normal except for one rat fed Aroclor 1016 that had leukemia (a dis ease that occurs occasionally sponta neously in our colony). The liver weights of the experimental animals did not differ from thoae of the con trols. The concentrations of the PCB-de rived materials are given in Table 1. The observed levels in adipose tissue are also shown in Fig 1. The highest concentrations for both compounds were found in adipose tissue, ranging from 36 to 143 ppm for Aroclor 1242 and 69 to 236 ppm for Aroclor 1016. The levels for brain, liver, and kidney were tower. The concentrations in urine and plasma were always leas than 1 ppm. In adipose tissue a steady state of storage of both compounds was approached in about two months and essentially achieved in four months. 304 Aroh Environ Haallh/Vol 20, Dee 107* HONS 062726 Gas chromatographic traces for all tissues were similar to thoae obtained from adipose tissue. The traces ob- tained from urine analysis had an en tirely different appearance. The dif ferent profiles obtained from adipose tissue, urine, and the PCB standard* are illustrated in Fig 1 to 3. When the peaks in the PCB standards were compared with the peaks obtained from PCB-derived material stored in the adipose tissue, it was found that some of the early peaks were absent. the intensity of others was enhanced, and peaks also emerged that were observed only as "shoulders'* in the standard. This observation suggest? either preferential absorption from the gastrointestinal tract, or prefer- , < j 1 , ' I . I 1 . t Polychlorinated Biphenyls/Burse st al I ntial metabolism, distribution, and excretion in the rats or both. The urine samples show a very dif ferent pattern. This suggests that certain isomers are excreted by the kidneys, that the products excreted in the urine are polar metabolites, or that nonchlorinated moieties re sponded to the electron capture de tector. However, a positive response was obtained with a conductometric detector (Coulson). Residue levels arising from Arodor 1016 ingestion were generally found to be higher than for Arodor 1242, Fig 4.--Residues resulting from a con tinuous dial of a 100 ppm concentration of Aroclor 1016 (e) or Arodor 1242 () for a period of ten months Residues following a continuous diet for only six months and re covery for five months for Aroefor 1016 (O) and for six months for Aroclor 1242 (o). art Htualreted by the broken llnee. Each point represents the mean and stanCard error (1) of four rete. Fig 6.-Uver of rat fed Aroclor 1016 for four months. Note dieted vesicles of smooth endoplasmic reticulum (SER). Some of the mho* chondrla are degenerated and others show an Increased number of parallel arranged crlstae (lead citrate, uranyt acetate, x 35,000) I* 10 the 1016 Of of 1 L.% for nil ittaimd Itt olf i on rn* he dirtdiponi1 ndanis iien Die were Uirinl ired in id thnt iboi'iii, mn<vil, uro !> in 1 he W1** . from ,reHT' *e el si Arch Environ Health/Vol 29, Dec 1974 MGXS 06272? Polychlorinated Blphenyls/fiurse f el 305 T with thn exception of the residues ol>served in the kidneys. However, this wax not true for all organs at every killing. After a ten-month feeding luriud the concentration of both PCNa in the liver was about the same, while at four months almost twice as much Aroclor 1016 as Aroclor 1242 was found. The levels found in adi pose tissue indicate a very sharp rise during the first month for Aroclor 1242, whilo Aroclor 1016 also main tains a sharp rise through the second month. After two months of exposure no appreciable increase was observed in adipose tissue of rats exposed to ei ther Aroclor 1026 or 1242. Very Utile PCB-derived material was excreted in the urine, and the ex cretion was very erratic. When rats were fed Aroclor 1242 they only ex creted measurable amounts during four to eight months of exposure. Kata fed Aroclor 1016 excreted meaaurablc amounts at each urine collec tion. The variation in urinary excre tion of PCB-derived material haa been observed previously in this labo ratory* and by other workers in the field.' The data resulting from the analy sis of tissues from rata that were fed the respective PCB-contsining diets for six months end were then contin ued on plain chow for various inter vals up to six months are given in Table 2. Tho concentrations found in adipose tissue arc also illustrated in Fig 4. Storage of material as detected by gas chromatography decreased more rapidly following discontinua tion of dosing in rats previously fed Aroclor 1016 than in those previously fed Aroclor 1242. Following a recov ery period of six months the residue Icvols in adiposo tissue were 21.8% of those observed after a six-month exIHNture period to Aroclor 1242. Follow ing a roenvory period of five months the residue levels in adipose tissue of rats fed Aroclor 1016 were only 11.8% of tho concentration found after six months of ingestion of this material. The gas chromatographic traces ob tained from the adipose tissue follow ing recovery from exposure to the PCB mixtures are shown in Fig 3. (Itolative retention times were com puted relative to the most intense 1 electron capturing peak present in Aroclor 1016 or 1242. All gas chro matographic columns were composed of 1.5% OV-17/1.95% QF-1 on 100/120 Gas Chrom Q.) If these results are compared with those obtained earlier from adipose tissue, s slight decrease in the number of peaks as well as a difference in the relative heights of the peaks is noticed. Composite urine and fat samples from rats fed Aroclor 1242 and 1016 continuously for varying time periods were analyzed by gas chromatography-maas spectrometry. Spectra re sulting from the composite adipose tissue analysis showed primarily triand Utrmchlorobiphenyla, for both PCB mixtures. Trace amounts of penta- and hexachlorobiphenyla were found in the Aroclor 1016 composites, while significant amounts of pentachlorobiphenyls and hexachloro biphenyla were found in the com posites obtained from rats that were fed Aroclor 1242. Spectra from urine composites of rats fed Aroclor 1016 revealed the presence of the same molecular ions observed in the adipose tissue compos ites. with the exception of hexachlor- obiphenyl. Dichlorohydroxybiphenyls (DCHB) and trichlorohydroxybiphenyls (TCHB) were also observed Di* chlorohydroxybiphenyl was more abundant. Spectra from urine composites of rats fed Aroclor 1242 revealed the presence of the same molecular ions observed in adipose tissue, with the noted absence of hexachlorobiphenyl. In contrast to Aroclor 1016, hydroxylated chlorobiphenyls were not observed in any of the spectra result ing from Aroclor 1242 urine compos ite analysis. The incidence of the various micro scopic changes observed in the liver is given in Table 3. Light microscopic examination of the liver of rats fed the two PCBs showed enlarged hepatocytes predominantly in the center of the lobules. Occasional hepatocytes had coarsely vacuolated cytoplasm or contained inclusions within the cyto plasm. The cytoplasm of some of the hepatocytes waa finely vacuolated and a brown pigment was noticed in the Kupffer cells and macrophages of some of the livers of the experimental rats fed Aroclor 1242 but not in rats fed Aroclor 1016. Vacuolation of the cytoplasm of the liver ceils was more consistently ob served in the livers of rats fed Aroclor 1242. Generally not much difference waa seen in the incidence of the vari ous changes between the two experi mental groups. All of the observed changes have to be considered as mild alterations that have been observed with a number of chlorinated cyclic chemicals Four and six months after exposure to the PCBs was discontinued the hepatocytes were still larger than those of the controls, but only one out of three rats showed vacuolated cyto plasm or inclusions within the cyto plasm. Since the number of animals was small, these results have to be in terpreted with reservation; however, it appears that the morphological changes encountered following expo sure to Aroclor 1242 and 1016 are re versible and disappear gradually af ter doting it stopped. Except for the hemorrhage and necrosis observed in one animal in each experimental group no evidence of serious liver damage was encountered. The livers of rats from the controls were normal when studied with the light microscope. Sections of four liv ers of rats fed Aroclor 1242 for six months and of four rats fed Aroclor 1016 for four months were also stud ied under the electron microscope, to gether with the same number of con trols. The livers of the control animals showed occasional lipid vacuoles but no abnormal changes. The livers of the experimental rats fed both PCB compounds showed an increase in smooth endoplasmic reticulum, an in crease in the number of lipid vacuoles, and atypical mitochondria Stacking of membranes within the mito chondria was also observed (Fig 5). Except for one animal that showed lymphocytic leukemia, no other ab normal findings were made on gross inspection of the organs at autopsy. It must be emphasized, however, that the only organ examined microscopic ally was the liver, since it was this or gan that was primarily affected by Aroclor 1254 and Aroclor 1260 as pre- viously tory.*" LevtW found in level of 9 182 ppm months' containin crease th sumption both irrou higher rc Aroclurl* the gastr dor 1242. storage v: pounds. 1 Aroelnr H sttributa! is known tion and ' substd'iti* tron -apt compound electron proporir* An alien PCB ri.-ri complr'* phenyl.Whethfactor- differnw at simi.. r ent not 1 the olL for Ar that \u crea.-wil .. exposure ued. Froiv. Aroclor ; ' rearlu'il exposui< more r:>, 1016. Mas? PCBs in pies wa the con-tcristu-. (3) obw These h SOS Arch Environ Health/Vol 29. Dec 1974 Polychlorlnated Biphenyls/Burse et al HONS 062728 Arch Z IF"* mm 3oLM. asm \ tl .\::h Hr . I*' t!. .1 r. dim ivt : tt>l Oh liv* six dor iuri. to* onnaif but i of CB in * in>lcn, inn ito* 5). wed nb ron< l*v I'hni .,>iv pr.- viously reported from our labora tory.*''' COMMENT Levels of PCB-derived components found in adipose tissue reached a level of 92 ppm (Aroclor 1242) and of 182 ppm (Aroclor 1016) after two months' exposure of rats to a diet containing 100 ppm, with little in crease thereafter. Although food con sumption of the rats was the same in both groups, rats on Aroclor 1016 had higher residues. It is possible that Aroclor 1016 is absorbed better from the gastrointestinal tract than Aro dor 1242. Another poeaibility ia that storage variea between the two com pounds The higher residue levels of Aroclor 1016 vs Arodor 1242 could be attributable to PCB quantitation. It ia known that the degree of chlorina tion and the position of the chlorine substitution contribute to the elec tron capturing properties of these compounds. In some instances their electron capture responses are not proportions! to theirconcentration.'n An alternative U> quantitating the PCB derivatives in tissue could be complete chlorination of the bi phenyls to decachlorobiphenyl." Whether a combination of the various factors outlined contributed to the difference in the PCB concentration at similar exposure times is at pres* snt not known. The concentrations in the other organs were often higher for Aroclor 1016, and the residues that were built up in the tissues de creased faster for this compound once exposure to the PCBs was discontin ued. Prom previous experience with Aroclor 1254* equilibrium seems to be reached sooner, and elimination after exposure has been discontinued is more rapid with Arociors 1242 and 1016. Maas spectra) identification of PCBs in composite fat and urine sam ples was based on*. (1) the presence of the correct molecular ion, (2) charac teristic chlorine isotopic dusters, and (3) observed fragmentation patterns. These were all in agreement with previously published data. The presence of DCHB and TCHB in Aroclor 1016 urine composite is in keeping with published data on meta bolic products for these compounds.1* However, one would generally not ex pect the method of extraction em ployed here (hexane at pH 7) to ex tract these metabolic products since they are generally found bound in the urine as glucosiduronides," and are detected following hydrolysis of the urine. It is feasible that small amounts of hydroxylated compounds could be found as free phenols and could then be chromatographed under selected conditions. As could be ex pected, the concentrations of the PCBs were much lower in the liver, kidneys, and brain than in adipose tissue, and even lower in plasma. Plasma concentration probably is not a very good indicator of the amount of PCBs stored in adipoee tissue. The morphological changes ob served In the liver are similar to those observed with other chlorinated hy drocarbons.1* They were generally mild, particularly when compared with findings made earlier with Aroclor 1254 and 1260.*-* Neither the pronounced lipid accumulation nor the edenofibrosis was observed in this study. However, some brown pigment was observed in the macrophages of the livers of some rata fed Arodor 1242 but not Aroclor 1016. It was not determined whether the pigment rep resented ceroid or whether part of it was caused by an accumulation of hemosiderin and porphyrin in the liver. After feeding rets Arodor 1242 and 1016 for six months, Goldstein et al" found an increase in liver uro porphyrin for Aroclor 1242 at this die tary level but not Arodor 1016 at the same dietary level. Unfortunately the number of rata was not large enough to determine if the occasional liver cell necrosis and hemorrhage that was seen was related to the exposure to Arodor 1242. From this study it may be concluded that both Arodor 1242 as well as Arodor 1016 are less persistent in the ret and have also less of an effect on the liver than ei ther Arodor 1254 or 1260. Arorion 1242 and 1016 were supplied by Mon santo Chemical Company. Si. Louis l Curnay, R. Moore, J. Couk and J. M. Jack gave technical assistance. References 1. Broadhurel MG: Uaa and reptaraabiHty of polychlorinated biphenyls. Efevirva Health Persped 2*1-102. 1972. 2. Koeman JH, Ten Noever De Brauw MC, at al: Chlorinated biphenyls in Ash, muaaria and birds from the river Rhina and the Netherlands coastal area. Nature 221:1122.1128, I960. 5. Kimbrough AW, Riacho P, Peake!) DB. et al: Polychlorinated biphenyls in the global acoeyatem. Nature 220:109*1102,1981. 4. Bagloy GR, Retehel WL, Cromartie C. Iden tification of polychlorinated biphenyls in two bald eagles by combined gat-liquid ehromatofra* phy-maea spectrometry. J <4sear Of Anof Cheat 63:261-261,197Q. 6. Zitko V, Hutalnger O, Choi PMX: Contentnation of tha Bay of Fundy-Gulf of Main* area with polychlorinated biphenyls, polychlorinated terphenyli. chlorinated dtbenodimina, and dlbansofurens. Environ Health Prrsprrttvr* H760. 1971 6. Curley A, Bunt VW, Grim ME,at ah Poly chlorinated bipbcnylr Distribution and storage in body fluids and tiaausa of Sherman rata. Enwran Ret 4:481-496,1971. 7. Pkntonow NW, Llptrap RM. Goiminfer HD: The distribution and excretion of poly chlorinated biphenyle (Arodor 1254) and their effect on urinary gonadal steroid levela in the boar. Bull Enriron Content Tanrul 7*58-966. 1972. 8. Kimbrough RD, Linder RB, Burn VW, et al: AdenoAbreeia in the rat liver. Arch Environ Health 27*90-995, 1979. 9. Kimbrough RD, Under RE, Gainee TB: Morphological changaa in liven of rats fed poly chlorinated biphenyls. Arch Environ Health 26*54-964, 1972. 10. Zitko V, Hutalnger O, Safe 3: Retention times and electron capture detector responses of some individual chlorobipbenyk Bull favire* Content Tuticot 6:160-163, 1971. 11. Rote JW, Murphy PG: A method for quan titation of polychlorinated biphenyl (PCB) isomere, Butt Environ Content Portent 6:977-384, 1971. 12. Berg OW, Sioeady PL, Reea GA. Column chromatographic separation of polychlorinated biphenyls from chlorinated hydrocarbon pesti cides, and their subsequent gas chromatographic ' quantitation in terms of derivatives. Bull Eastren Content Taatcol 7:998-347, 1972. 19. Hutanger O, Nash DM. Safa S: Poly chlorinated biphenyls: Metabolic behavior of pure isomer* in pigeons, rata, and brook trout Sciaut 178:312-314, 1972. 14. Block WD, Cornish HH: Meteboiiam of biphenyl and 4-chlorobiphenyi in the rabbit J Bid Ckrm 284:3301-3902, 1969. 15. Kimbrough RD, Gaines TB, Under RE; The ullrestracture of livers of rets fed DDT and dieldrin. Areh Environ Health 22:460-467, 1971. 16. Goldstein JA. Hickman P. Bergman H.et al: Induction of hepatic porphyria and drug me tabolism by two polychlorinated biphenyls (PCB) mixture containing 42% chlorine (Arorior 11)18 and 1242), abstract. Fed Prve 33:219, 1974. H sl ] Arch Environ Msslth/Vol 29, Dsc 1974 Polychlorinated Biphenyls/Burss at al 307 K0NS 062 7iS