- N-Nitrosobenzylmethylamine hydroxylation and coumarin 7-hydroxylation: Catalysis by rat esophageal microsomes and cytochrome P450 2A3 and 2A6 enzymes
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N-Nitrosobenzylmethylamine (NBzMA) is a potent and selective esophageal carcinogen in the rat and may be a causative agent for human esophageal cancer. This nitrosamine, like most, must be metabolically activated to exert its carcinogenic potential. NBzMA may be metabolized by P450-catalyzed methyl or methylene hydroxylation; the latter is believed to be the activation pathway. The sensitivity of the esophagus to NBzMA-induced tumorigenesis is believed to be due, at least in part, to the presence of efficient P450 catalysts in this tissue. However, while it was reported almost 20 years ago that the rat esophagus catalyzes the methylene hydroxylation of NBzMA, the P450 that catalyzes this reaction has yet to be identified. We report here that human P450 2A6 and the closely related extrahepatic rat enzyme P450 2A3 both efficiently catalyze NBzMA methylene hydroxylation, characterized as benzaldehyde formation. The catalytic efficiency of P450 2A3 in this reaction was 3-fold greater than that of P450 2A6, 7.6 (K(m) = 0.63 ± 0.18 μM and the V(max) = 4.8 nmol min-1 nmol of P450-1) versus 2.3 (K(m) = 6.7 ± 2.9 μM and the V(max) = 15.7 nmol min-1 nmol of P450-1), respectively. Both enzymes catalyzed methylene hydroxylation at least 4-fold more efficiently than methyl hydroxylation. In addition, P450 2A6, but not P450 2A3, catalyzed benzyl ring hydroxylation, generating N-(p-hydroxybenzyl)methylamine. The identity of this metabolite was confirmed by synthesis of a standard and LC/MS and LC/MS/MS analysis. P450 2A6 is an efficient coumarin 7-hydroxylase, and we report here that P450 2A3 is an equally good catalyst of this reaction (K(m) = 1.7 ± 0.41 μM and V(max) = 1.7 ± 0.08 nmol min-1 nmol of P450- 1). Rat esophageal microsomes (REM), like P450 2A3, were efficient catalysts of NBzMA methylene hydroxylation. However, in contrast to P450 2A3, the major product of this reaction was the product of benzaldehyde oxidation, benzoic acid. Antibody to the closely related mouse P450, 2A5, did not inhibit REM- catalyzed NBzMA metabolism, and most importantly, REM did not catalyze the 7- hydroxylation of coumarin. Therefore, P450 2A3 does not appear to be the P450 in the rat esophagus responsible for catalyzing the methylene hydroxylation of NBzMA.
- Von Weymarn, Linda B.,Felicia, Nadia D.,Ding, Xinxin,Murphy, Sharon E.
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p. 1254 - 1261
(2007/10/03)
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- Synthesis and Reactions of Phenolic Alkylbenzylnitrosamines
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N-Alkyl-p-hydroxybenzylnitrosamines have been synthesised from p-benzoyloxybenzaldehydes via imines and amines which were nitrosated before removal of the protecting group with n-propylamine.N-Alkyl-o-hydroxybenzylnitrosamines were similarly obtained from o-methoxymethoxybenzaldehydes, the final deprotection being achieved with dilute acid.The nitrosamines decompose in hot aqueous alkali by two mechanisms, one through a quinone methide to yield a benzyl alcohol and the other by an elimination eventually yielding a benzaldehyde.The decomposition of the nitrosamines in the presence of 2-naphthol in alkaline solution results in alkylation of the naphthol and production of compounds from the reaction of 2-naphthol with the intermediate quinone methides.The N-sec-butylnitrosamines alkylate 2-naphthol on carbon but the N-methylnitrosamines alkylate on oxygen and during the methylation 94percent of deuterium exchange with methyl hydrogen occurs in deuterium oxide and there is 6percent intact transfer of the methyl group from the N-methylnitrosamine to the 2-naphthol.
- Bowman, Peter J.,Brown, Ben R.,Chapman, M. Andrew,Doyle, Paul M.
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p. 701 - 770
(2007/10/02)
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