Studies on the metabolic activation of β-keto nitrosamines: Mechanisms of DNA methylation by N-(2-oxopropyl)- N-nitrosourea and N-nitroso- N-acetoxymethyl- N-2-oxopropylamine

Abstract

At pH 7.35, N-(2-oxopropyl)- N-nitrosourea (OPNU) reacted with calf thymus DNA to yield O 6-methylguanine, 7-methylguanine and 3-methyladenine. Kinetic measurements of the base catalyzed decomposition of OPNU and the extent of methylation of DNA by OPNU suggested that methylnitrosourea is not formed as an intermediate product. Diazoacetone, acetic acid and methanol were identified as products of decomposition of OPNU at pH 7.35. Reaction of OPNU with N-methylmaleimide yielded the product resulting from 1,3-dipolar cycloaddition of diazomethane. Hydrolysis of N-nitroso- N-acetoxymethyl- N-2-oxopropylamine (NAMOPA) in the presence of hog liver esterase also produced diazoacetone, acetic acid and methanol. Enzymatic hydrolysis of NAMOPA in the presence of DNA produced O 6-methylguanine, 7-methylguanine and 3-methyladenine. These results suggest that OPNU undergoes base-catalyzed decomposition and NAMOPA undergoes enzymatic hydrolysis to yield the same intermediate, 2-oxopropyldiazotate. This diazotate then reacts either by protonation followed by loss of water to form diazoacetone, or by internal nucleophilic attack by the diazotate oxygen on the carbonyl carbon to form an oxadiazoline intermediate which then collapses to form acetate and the methylating agent diazomethane. These reaction schemes are used to suggest the mechanism by which N-nitroso-2-oxopropylpropylamine methylates hepatic DNA in vivo.