Detection of PCB Adducts by the ³²P-Postlabeling Technique

Abstract

The purpose of this study was to determine whether lower chlorinated biphenyls would be bioactivated to electrophilic metabolites by microsomes alone or in combination with peroxidase. Monochloro- and dichlorobiphenyls were incubated with liver microsomes of rats treated with phenobarbital and beta-naphthoflavone, an NADPH-regenerating system, and deoxyguanosine 3'-monophosphate (dGp). The resultant adducts were analyzed by 32P-postlabeling either following microsomal incubation alone ("preoxidized") or coupled with subsequent oxidation with horseradish peroxidase/H2O ("oxidized"). The incubation of 4-monochlorobiphenyl (4-MCB) resulted in the formation of two minor adducts by microsomal activation alone. However, the oxidized sample showed two additional major adducts. Formation of the latter adducts was almost completely (> 80%) inhibited when the oxidation reaction was performed in the presence of ascorbic acid. The other test mono- and dichlorobiphenyls also formed 1-3 major adducts. Compared with microsomal activation alone, these adducts were enhanced after the oxidation reaction or detected only in the oxidized samples. These data suggest that (1) some adducts of the lower chlorinated biphenyls are derived from arene oxides and (2) many adducts may be formed by metabolism of the parent compounds to catechol and p-hydroquinone species, which are oxidized to semiquinones and/or quinones. The involvement of quinones and/or semiquinones was supported by UV/vis spectroscopic measurements, which showed that metabolites of 4-MCB can be oxidized to products with spectra characteristic of quinones. These data raise the possibility that lower chlorinated biphenyls may be genotoxic and may explain the fact that commercial polychlorinated biphenyl mixtures are complete rodent carcinogens.

Metabolic activation of PCBs to carcinogens in vivo—A review

“…The metabolism of PCB3 by cytochrome P-450 seems to involve arene oxide intermediates (McLean et al, 1996b). These are converted to mono-hydroxy metabolites and in further steps, probably through a secondary arene oxide intermediates, from the mono-to the di-hydroxy forms (Figure 3).…”

Understanding Population and Individual Risk Assessment: The Case of Polychlorinated Biphenyls

“…Overwhelming evidence shows that the PCBs are not mutagenic (9). DNA binding has been reported in some crude studies (10), although a specific nucleotide adduct has not been identified (11)(12)(13)(14)(15).…”

Polychlorinated Biphenyls and Polybrominated Biphenyls

“…Formation of protein and DNA adducts was observed in vitro with PCB-3 and the tetrachlorinated congeners PCB-47, PCB-49, PCB-52, and PCB-77 (Wyndham et al, 1976;Shimada & Sawabe, 1984). DNA-adduct formation was also observed with a series of 15 mono-and dichlorinated PCBs, with but not without activation by microsomes, horseradish peroxidase and hydrogen peroxide (McLean et al, 1996b). This suggested that quinones were the ultimate genotoxic agents.…”

“…Rodent and human liver microsomes produced similar or different adduct patterns depending on the PCB congener used, indicating that species differences exist. Reactive intermediates can bind to cellular macromolecules, including DNA and DNA-maintenance proteins, and such adducts can be detected in multiple organs (Morales & Matthews, 1979;Amaro et al, 1996;McLean et al, 1996b;Oakley et al, 1996aOakley et al, , 1996bLin et al, 2000;Pereg et al, 2001Pereg et al, , 2002Srinivasan et al, 2002;Arif et al, 2003;Zhao et al, 2004;Bender et al, 2006;Bender & Osheroff, 2007).…”