RT Journal Article SR Electronic T1 NICOTINE 5′-OXIDATION AND METHYL OXIDATION BY P450 2A ENZYMES JF Drug Metabolism and Disposition JO Drug Metab Dispos FD American Society for Pharmacology and Experimental Therapeutics SP 1166 OP 1173 DO 10.1124/dmd.105.004549 VO 33 IS 8 A1 Sharon E. Murphy A1 Vytautas Raulinaitis A1 Kathryn M. Brown YR 2005 UL http://dmd.aspetjournals.org/content/33/8/1166.abstract AB In smokers, the primary pathway of nicotine metabolism is P450 2A6-catalyzed 5′-oxidation. The nicotine Δ5′(1′)-iminium ion product of this reaction is further metabolized to cotinine by aldehyde oxidase. Previous investigators have reported kinetic parameters for cotinine formation using human liver cytosol as a source of aldehyde oxidase. Using [5-3H]nicotine and radioflow high-performance liquid chromatography analysis, we determined kinetic parameters for nicotine 5′-oxidation by P450 2A6 and the closely related human extrahepatic P450 2A13 as well as the rodent P450s 2A3, 2A4, and 2A5. The formation of both cotinine and nicotine Δ5′(1′)-iminium ion was monitored. The Km and Vmax values for P450 2A6 were 144 ± 15 μM and 1.30 ± 0.05 pmol/min/pmol, respectively. Previously reported Km values for cotinine formation by P450 2A6 in the presence of cytosol were much lower, ranging from 11 to 45 μM. P450 2A13 was a somewhat better catalyst of nicotine Δ5′(1′)-iminium formation, with 2-fold lower Km and 2-fold higher Vmax values than P450 2A6. The rat P450 2A3 and the mouse P450 2A5, which are 85 and 84% identical to P450 2A6, were much more efficient catalysts of nicotine 5′-oxidation. P450 2A4 was not an efficient catalyst of nicotine metabolism. Whereas 5′-oxidation was the major pathway of nicotine metabolism for all five P450 2A enzymes, these enzymes also catalyzed methyl oxidation. Nornicotine, the product of this reaction was detected as 5 to 15% of the total nicotine metabolites. Nornicotine is the amine precursor to the esophageal carcinogen N′-nitrosonornicotine. Therefore, methyl oxidation of nicotine by P450 2A6 or P450 2A13 followed by nitrosation of nornicotine are possible endogenous pathways of N′-nitrosonornicotine formation. The American Society for Pharmacology and Experimental Therapeutics