Redox cycling in the metabolism of the environmental pollutant and suspected human carcinogen o-anisidine by rat and rabbit hepatic microsomes.
We investigated the ability of hepatic microsomes from rat and rabbit to metabolize 2-methoxyaniline (o-anisidine), an industrial and environmental pollutant and a bladder carcinogen for rodents. Using HPLC combined with electrospray tandem mass spectrometry, we determined that o-anisidine is oxidized by microsomes of both species to N-(2-methoxyphenyl)hydroxylamine, o-aminophenol, and one additional metabolite, the exact structure of which has not been identified as yet. N-(2-Methoxyphenyl)hydroxylamine is either further oxidized to 2-methoxynitrosobenzene (o-nitrosoanisole) or reduced to parental o-anisidine, which can be oxidized again to produce o-aminophenol. To define the role of microsomal cytochromes P450 (P450) in o-anisidine metabolism, we investigated the modulation of this metabolism by specific inducers and selective inhibitors of these enzymes. The results of the studies suggest that o-anisidine is a promiscuous substrate of P450s of rat and rabbit liver; because P450s of 1A, 2B, 2E, and 3A subfamilies metabolize o-anisidine in hepatic microsomes of both studied species. Using purified enzymes of rat and rabbit (P450s 1A1, 1A2, 2B2, 2B4, 2E1, 2C3, 3A1, and 3A6), reconstituted with NADPH:P450 reductase, the ability of P450s 1A1, 1A2, 2B2, 2B4, 2E1, and 3A6 to metabolize o-anisidine was confirmed. In the reconstituted P450 system, rabbit P450 2E1 was the most efficient enzyme metabolizing o-anisidine. The data demonstrate the participation of different rat and rabbit P450s in o-anisidine metabolism and indicate that both experimental animal species might serve as suitable models to mimic the fate of o-anisidine in human.
Naiman
Karel
K
Department of Biochemistry, Faculty of Science, Charles University, AlbertoV 2030, 128 40 Prague 2, Czech Republic.
Dracínská
Helena
H
Martínková
Markéta
M
Sulc
Miroslav
M
Dracínský
Martin
M
Kejíková
Lucie
L
Hodek
Petr
P
Hudecek
Jirí
J
Liberda
Jirí
J
Schmeiser
Heinz H
HH
Frei
Eva
E
Stiborová
Marie
M
D000814:Aniline Compounds / Q000378:metabolism*; D000818:Animals; D002274:Carcinogens, Environmental / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D008658:Inactivation, Metabolic; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism*; D010084:Oxidation-Reduction; D011817:Rabbits; D051381:Rats; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry
D000814:Aniline Compounds; D002274:Carcinogens, Environmental; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System; C003898:2-anisidine