Bromobenzene 3,4-oxide alkylates histidine and lysine side chains of rat liver proteins in vivo.

The hepatotoxic effects of bromobenzene (BB) are correlated with and generally ascribed to the covalent modification of cellular proteins by chemically reactive metabolites, particularly BB-3,4-oxide. Previous studies revealed that quinone as well as epoxide metabolites of BB form adducts to protein sulfur nucleophiles, that the quinone-derived adducts are more abundant by a factor of ca. 7, and that collectively these sulfur adducts account for only about 10% of the total protein covalent binding [Slaughter, D. E., and Hanzlik, R. P. (1991) Chem. Res. Toxicol. 4, 349-359]. To examine the possibility that metabolically-formed BB-3,4-oxide alkylates nitrogen nucleophiles on proteins under toxicologically relevant conditions in vivo, we synthesized standards of N tau-(p-bromophenyl)histidine (7) and N epsilon-(p-bromophenyl)lysine (8) as anticipated adduct structures and used them to guide a chromatographic search for their presence in hydrolysates of liver protein from BB-treated rats. While radio-LC chromatography and GC/MS provide unequivocal evidence for their presence, the amounts of 7 and 8 observed are very low ( < 1% of total covalent binding). The apparently small net contribution of epoxide metabolites to covalent binding of BB in vivo suggests the majority of binding may arise via quinone metabolites, but this should not be construed to imply that quinone adducts are necessarily more important toxicologically than epoxide adducts; in this context the identity of the protein targets is probably at least as important as the type of electrophilic metabolite involved.