Identification of the cytochrome P450 isoforms involved in the O-demethylation of 4-nitroanisole in human liver microsomes

1. 4-Nitroanisole is O-demethylated to 4-nitrophenol by human liver microsomes. Kinetic studies indicate that this metabolic route is mediated by two cytochrome P450 isoforms, one with a K-m = 2.1 mu M and the other with a K-m = 220 mu M. 2. Chemical inhibition and correlation studies in human liver microsomes indicate that the low K-m enzyme is CYP2A6 and the high K-m enzyme is CYP2E1 suggesting that 4-nitroanisole is not a general cytochrome P450 substrate. 3. Studies using expressed recombinant cytochrome P450s indicated that all the cytochrome P450s investigated metabolized 4-nitroanisole but CYP2A6 and CYP2E1 produced the highest rates. Kinetic studies with these two isoforms produced a K-m for CYP2A6 of 9 mu M and 54 mu M for CYP2E1. 4. The involvement of these two isoforms in the O-demethylation of 4-nitroanisole can be rationalized in terms of a hydrogen bond interaction with the nitro group and the active site of CYP2A6 and a hydrophobic interaction with the active site of CYP2E1.