Ketonitrosamines as metabolites of methyl-n-amylnitrosamine (MNAN) and its hydroxy derivatives in the rat

In a previous study of the metabolism of methyl-n-amylnitrosamine(MNAN) in the rat, 2- to 5-hydroxy-MNAN (HOMNAN) were provisionallyidentified as metabolites and the identity of 4-HO-MNAN wasconfirmed by mass spectrometry. We now describe syntheses andmass and other spectra for 2- to 5-oxo-MNAN. Two previouslyunidentified MNAN metabolites were shown to be 3- and 4oxo-MNAN.In addition to 4-HO-MNAN, we confirmed 3-HO-, 4oxo- and (lesscertainly) 2-HO-MNAN as urinary MNAN metabolites by GLC-MS ofHPLC fractions. Analysis with and without ß-glucuronidasetreatment showed that the urinary HO-MNANs occurred as theirß-glucuronides. MNAN (25 mg/kg injected i.p.) hada blood half-life of 21 min in adult male rats. The blood alsocontained 4-HO- and 4oxo-MNAN, which showed maximum levels thatwere 13 and 26% respectively of that for MNAN, and were clearedmore slowly than MNAN. On incubation for 3 h with MNAN, ratesophagus produced 3- and 4-oxo-MNAN in yields that were 5%of those for the corresponding HO-MNANs. For MNAN metabolism,the 4-oxo-/4-HO-MNAN ratio of metabolites was 5% for adult ratliver and was 22% for adult hamster liver and 9-day-old ratliver. On incubation with 4-HO-MNAN for 3 h, oxidation to 4oxo-MNANwas 16–25% for adult hamster or 9-dayold rat liver slicesand for adult hamster liver homogenate. Homogenate activitywas concentrated in the microsomal fraction, for which NAD wasa more effective co-factor than NADP. A bacterial alcohol dehydrogenaseoxidized 4-HO- to 4-oxo-MNAN in 38% yield/3 h. None of thesepreparations oxidized 2-HO- to 2-oxo-MNAN. It was concludedthat 3- and 4-oxo-MNAN were metabolites of MNAN, apparently(for 4-oxo-MNAN) via HO-MNAN oxidation by a microsomal NAD-dependentenzyme, that 4-HO- and 4-oxo-MNAN formation was a major routeof MNAN metabolism, and that 4-oxo-MNAN might play a role inMNAN carcinogenesis.