Metabolism of the food derived mutagen and carcinogen 2-amino-1-methyl-6-phnylimidazo(4,5-b)pyridine (PhIP) by human liver microsomes

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Abstract
Animal studies have shown that 2-amino-1-methyl-6-phenyl-imidazo(4,5-b)pyridine (PhIP) undergoes both activation to a genotoxic metabolite and detoxication, catalysed by CYP enzymes. In this study, using direct chemical analysis, we have examined PhIP metabolism by the microsomal fraction of hunian liver for comparison to that occurring in animals. PhIP was incubated with human liver microsomes in the presence of an NADPH regenerating system and the reaction mixture then analyzed by HPLC. Only one metabolite, identified as N-hydroxy PhIP, was produced. The N-hydroxylation of PhIP by human liver microsomal fraction obeyed Mlchaelis—Menten kinetics, with a Km of 55 µM and a Vmax of 666 pmol/min/mg protein. Furafylline, a potent and specific inhibitor of CYP1A2 in man, inhibited this reaction by >95%, with an IC50 of 0.6 µM. PhIP inhibited high affinity phenacetin O-deethylase activity of human liver microsomes, an activity catalysed specifically by CYP1A2, with an IC of about 80 µM. These data indicate that, in human liver microsomes, N-hydroxylation is the only route of oxidative metabolism of PhIP, yielding a genotoxic species, and that this reaction is catalysed almost exdusively by CYP1A2. Furthermore, the exdusive oxidative activation of PhIP by human liver is in direct contrast to PhIP metabolism in rodents and non-human primates where oxidative detoxica tion products predominate.

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