N-OXYGENATION OF PRIMARY AMINES AND HYDROXYLAMINES AND RETROREDUCTIONOF HYDROXYLAMINES BY ADULT HUMAN LIVER-MICROSOMES AND ADULT HUMAN FLAVIN-CONTAINING MONOOXYGENASE-3

Adult human liver microsomes catalyze the NADPH-dependent N-oxygenatio n of -N-(n-octylamino)-2-(trifluoromethyl)phenothiazine to the corresp onding oximes through the intermediacy of the hydroxylamine. In the pr esence of adult human liver microsomes, the primary amine is stereosel ectively converted to the cis-oxime, but addition of the alternative c ompetitive substrate hydroxylamine hydrochloride apparently decreases the amount of aliphatic hydroxylamine retroreduction because an increa se in oxime formation was observed. In the presence of hydroxylamine h ydrochloride, however, the oxime product recovered was formed with ver y low stereoselectivity. Studies on the biochemical mechanism of oxime formation suggested that cis-oxime formation in the presence of adult human liver microsomes was largely dependent on the human flavin-cont aining monooxygenase (form 3). This conclusion is based on the effects of incubation conditions on product formation when compared to result s observed in the presence of cDNA-expressed human FMO3. The retroredu ction of the intermediate hydroxylamine was dependent on NADPH but was not catalyzed by human flavin-containing monooxygenase (form 3) or an y one of seven prominent cytochromes P-450 that have been well-charact erized in the human liver microsomes examined. The results suggest tha t aliphatic primary amines are efficiently sequentially N-oxygenated i n the presence of human liver microsomes to hydroxylamines and then to oximes mainly by the human flavin-containing monooxygenase. Retroredu ction of the intermediate hydroxylamine is apparently facilitated by a novel but as yet poorly characterized enzyme system that does not emp loy any of the currently known well-characterized cytochrome P-450 enz ymes present in adult human liver microsomes.