Cytochrome P450-mediated metabolism and cytotoxicity of aflatoxin B-1 in bovine hepatocytes

Aflatoxin B-1 (AFB(1)) biotransformation comprises cytochrome P450-mediated reactions resulting in hydroxylated and demethylated metabolites as well a s AFB(1) epoxides. As the latter are highly nucleophilic, the species-speci fic rate of epoxidation and the ability for rapid conjugation to glutathion e by glutathione S-transferase determines the individual susceptibility to AFB(1). Here we show the time- and dose-dependent rate of AFB(1)-metabolism in bovine hepatocytes. Aflatoxin M-1 (AFM(1)) is the most prominent metabo lite formed within the first 2-8 hr of incubation, whereas AFB(1)-dhd is de tectable in medium mainly after a prolonged incubation period, The delayed formation of AFB(1)-dhd corresponds to the cytotoxicity demonstrated by the MTT assay. alpha-Naphthoflavone and ketoconazole, inhibitors of CYP1A and CYP3A, respectively in humans, were used to evaluate the contribution of sp ecific P450 isoenzymes in bovine biotransformation of AFB(1), Initial exper iments confirmed that alpha-naphthoflavone and ketoconazole inhibited ethox yresorufin O-deethylation and testosterone 6 beta-hydroxylation also in bov ine hepatocytes. Both inhibitors reduced AFM(1) and AFB1-dhd formation conc entration dependently, suggesting that both enzyme groups contribute to the formation of these metabolites. However, the formation of AFM(1) was less inhibited by both compounds than the formation of AFB(1)-dhd. (C) 2000 Else vier Science Ltd. All rights reserved.