Document aDvyy4kMpOvqZLa7Zw3DB9r8N

Beilrage zur Okologischen Cheinie LXXXIV+ Metabolism of Lower Polychlorinated Biphenyls-I4C in the Rhesus Monkey by W. Greb, W. Klein, F. Coulston*,L. Golberc*, and F. Korte Intlilui fur dkologische Chentie der Gesellschaft fur Strahlen und Umweltfortchung mbH, M unchcn DS20S St. Augustin 1, W. Germany *lnititute of Experimental Pathology and Toxicology Albany Medical College, Albany, IV.Y. 12208 INTRODUCTION Metabolism studies on PCB are important for estimating the toxicity of. metabolites after knowing their identi ty and biological pathway. Some pure PCB-isomers were shown to be converted to hydroxylated products by rat liver microsomes (GREB et al. 1974), rabbits (BLOCK and CORNISH 1959), rats (HUTZINGER et al. 1972, YOSHIMURA and YAMAMOTO 1973) and pigeons (YOSHIMURA and YAMAMOTO 1973) . The purpose of this study was to identify all major metabolites after PCB-admlnistration to Rhesus monkeys and to measure the amounts of diffe rent excreted metabolites. Correlation of conversions and elimination pattern of PCBs with different chlorine content might lead to conclusions about the behavior of other PCBs. The excretion rates of 2,4'-dichlorobiphenyl and 2,5,21-trichlorobiphenyl have been described elsewhere (GREB et al. 1973)> we now wish to report the identifi cation of their metabolites. EXPERIMENTAL The PCBs investigated were 2,41-dichlorobiphenyl and 2,5,2'-trichlorobiphenyl, both major components^of the lower AROCLOR series. They were labelled with qC. Fema le Rhesus monkeys were housed in metabolic cages and the PCB was injected once in the left forearm vein. Three animals received 2,4'-dichlorobiphenyl in doses of 16.8, 77.6 and 566y(fcg/kg, while 82.6/*g/kg of the 2,5,2'-trichlorobiphenyl were administered to one mon key. Urine and feces were collected daily. Conjugates were hydrolyzed by refluxing urine with an equal volume of 8 N H_SO. for one hour and extracted with ether. Fe ces were mixed with anhydrous sodium-sulfate and extrac ted for two days in a Soxhlet apparatus with CHjCl, and then with methanol. The combined extracts were conceri- +LXXXIII. Communication: GREB, W., W. Klein, F.COULSTON L.GOLBERG,and F.KORTE: Bull .Environ.Contain. Toxicol., in press. Bulletin of Environmental Contamination A Teslcolojy, V1, 13, No. 4 1?75 by Sprlnfrr-Yerlig New York Inc. 471 STLCOPCB4009903 TABLE 1 TLC GLC-- and MS-Data of 2,4'-Dichloroblphenyl Metabolites In Monkeys Data OH-Metabo- OH-Metabo- CH-Metabo- (CH)--Meta (OH),-Me- lite Sx 1ite S2 lite S3 bolite S4. bol. S5 R> before methyiation 1 (ch2ci2) 0.66 0.50 0.25 0.13 < 0.1 Rf after methyiation r (ch2ci2) 5751 0.70 0.66 5759 0.64 |0H)j--Me-- :abof. Sg < 0.1 0.52 GLC after methyiation Rt (min.) MS-fragment a|ter methyiation M 10.2/179C 12.1/185C 11.4/185C 14 .8/200C 13.1/200C 11.6/200C for 9 min. for 9 min. for 8 min., for 8 min. for 8 min., then temp, then temp, then temp, then temp, then temp, raised 5C raised 5C raised 5C/ raised 5Q raised 5C/ min. min. min. min. min. 252 s 252 s 252 s 282 s 282 s 282 s " M+-CH3 " metastable " M+-C1 " M+-HC1 " M+-CH3-CO 217 1 209 m 237 1 .- 217 m - 209 s 237 m 223* m '- 209 m 267 1 - 247 m 246 m 239 m 267 m * 247 m 246 m 239 1 267 m " - 239 1 " M+-CH3-C1 - - 202 1 - -- " M+-CH3-C0-HC1 173 1 - 173 m - -- " M+-CH3-C0-C12 - - 139 m 204 1 - - Peak-Intensities: s*strong, mTnedium, l*low TABLE 2 TLC-, GLC - and MS-Data of 2,5,2'-Trichlorobiphenyl Metabolites in Monkeys Data OH-Metabo- OH-Metabo- OH-Metabo- (OH)--Meta- .(OH)--Meta- (OH),-Me- lite Gx lite G2 lite G3 bolite G4 bolice G5 tabol. Gg Rf before methylation r (ch2ci2) 0.49 0.25 0.25 0.11 <0.1 < 0.1 R, after methylation' 1 <ch2ci2) 0.76 0.72 0.72 0.64 0.53 0.32 GLC after methylation Rfc (min.) MS-fragment a^ter methylation M " m+-ch3 13.8/180C 286 s 271 1 15.4/185C 16.2/185C 17.0/20OC 13.7/200C 17.8/200C for 8 min., for 8 min., for 8 min., then temp, then temp, then temp, raised 5C/ raised 5C/ raised 5C/ min. min. min. 286 s 271 1 286 s - 316 s 301 1 316 s 301 1 346 s - " M+-C1 " m+-ch3-co 251 m 243 m 243 1 251 1 243 1 281 1 273 m 273 1 - - " m+-ch3-ci - - - 266 1 - - " M+-C12 186 1 - - 246 1 246 3 276 1 " M+-CH3-CO-Cl2 173 1 173 - 203 1 - - DSW 0 2 5 9 4 9 Peak-Intensitie31 b*> strong, m medium, l*=low After methylation all purified metabolites showed in the mass spectra -OCH_-groups indicating that the original metabolites were phenolic derivatives. In both cases a number of isomers had been formed. Isomers only dif fered in peak-intensities. All Isolated compounds showed the typical isotopic distribution pattern corresponding to the number of chlorine atoms in the parent PCB. For 2,4'-dichlorobiphenyl there was no conversion exceeding the introduction of 2 OH-groups per molecule. For the 2,5,2 *-trichlorobiphenyl we observed a metabolite with three OH-groups. Corresponding to its formation the monohydroxy-metabolite with the lowest concentration should be the precursor of the highest concentrated di hydroxy-compound. Only three monohydroxy-derivatives were formed although six (reap, seven for the trichloro biphenyl) structures are possible. For each PCB a highly polar zone of 1 and 4% resp. of radioactivity could not be identified. Data of the metabolites are shown in Table 1-2. Comparing the excreted amounts (in % of totally excreted radioactivity) of metabolites formed, the dichlorobiphenyl was mainly excreted as monohydroxy-derivatives, while the trichlorobiphenyl was mainly eliminated as di hydroxy-compounds (Table 3). TABLE 3 Comparison of Metabolites formed of PCBs by Rhesus Monkeys Metabolites Dichlorobiphenyl Trichloro biphenyl Monohydroxy-metabolites Dihydroxy-metabolites Trihydroxy-metabolites Polar metabolites, unidentified 66.6% 32.4% - . 1% 7.8% ' 82.2% 6t 4% <7S DSW 025950 STLCOPCB4009906 The following might explain this different behavior. Due to the additional chlorine atom, the trichlorobiphenyl is more lipophilic than the dichlorobiphenyl. The mono hydroxy-metabolites react in the same way: the monohydroxy-dichlorobiphenyl is excreted rapidly, while the water solubility of the monohydroxy-trichlorobiphenyl is still low. After further hydroxylation the trichloro biphenyl -metabolites reach a polarity to be easier eli minated. A dihydroxy-trichlorobiphenyl thus should correspond to a monohydroxy-dichlorobiphenyl as regards speed of elimination. For higher chlorinated BPs, a higher degree of hydroxylation is necessary and will cause a longer retention time of the compound in the bo dy, that is, a Blow excretion rate. This might explain their accumulation. CONCLUSION Metabolism of PCB with high chlorine content will be dif ficult and proceed slowly. Concerning lower chlorinated biphenyls, however, this study reveals that total degra dation to hydroxylated metabolites and the rapid excre tion of the PCBs investigated in primates indicate that accumulation of lower chlorinated biphenyls at low doses will not occur. REFERENCES BLOCK, W.D., and H.H. CORNISH: J. Biol. Chem. 234, 3301 (1959). DE BOER, Th. J., and H.J.BACKER: Org. Syn. Coll. 4, 943 (1963). GREB, W., W. KLEIN, F. COULSTON, L. GOLBERG, and F. KORTE: Chemosphere 2_, 14 3 (1973) . GREB, W., W. KLEIN, F. COULSTON, L. GOLBERG, and F. KORTE: Bull. Environ. Contam. Toxicol., in press (1974). HUTZINGER, O. et al.: Science 178, 312 (1972). VOIGT, K.D. in H.D. BERGMEIER: Methods of Enzymatic Analysis. Acad. Press 1965, p, 462. YOSHIMURA, H., and H. YAMAMOTO: Chem. Pharm. Bull.21 1168 (1973). DSw 025951 476 mawmv STLCOPCB4009907