@article {Murphy1166, author = {Sharon E. Murphy and Vytautas Raulinaitis and Kathryn M. Brown}, title = {NICOTINE 5'-OXIDATION AND METHYL OXIDATION BY P450 2A ENZYMES}, volume = {33}, number = {8}, pages = {1166--1173}, year = {2005}, doi = {10.1124/dmd.105.004549}, publisher = {American Society for Pharmacology and Experimental Therapeutics}, abstract = {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 {\textpm} 15 μM and 1.30 {\textpm} 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}, issn = {0090-9556}, URL = {https://dmd.aspetjournals.org/content/33/8/1166}, eprint = {https://dmd.aspetjournals.org/content/33/8/1166.full.pdf}, journal = {Drug Metabolism and Disposition} }