Reduction of Amphetamine Hydroxylamine and Other Aliphatic Hydroxylamines by Benzamidoxime Reductase and Human Liver Microsomes

Abstract

For the reduction of N-hydroxylated derivatives of strongly basic functional groups, such as amidines, guanidines, and aminohydrazones, an oxygen-insensitive liver microsomal system, the benzamidoxime reductase, has been described. To reconstitute the complete activity of the benzamidoxime reductase, the system required cytochrome b(5), NADH-cytochrome b(5)-reductase, and the benzamidoxime reductase, a cytochrome P450 enzyme, which has been purified to homogeneity from pig liver. It was not known if this enzyme system was also capable of reducing aliphatic hydroxylamines. The N-hydroxylation of aliphatic amines is a well-known metabolic process. It was of interest to study the possibility of benzamidoxime reductase reducing N-hydroxylated metabolites of aliphatic amines back to the parent compound. Overall, N-hydroxylation and reduction would constitute a futile metabolic cycle. As examples of medicinally relevant compounds, the hydroxylamines of methamphetamine, amphetamine, and N-methylamine as model compounds were investigated. Formation of methamphetamine and amphetamine was analyzed by newly developed HPLC methods. All three hydroxylamines were easily reduced by benzamidoxime reductase to their parent amines with reduction rates of 220.6 nmol min(-1) (mg of protein)(-1) for methamphetamine, 5.25 nmol min(-1) (mg of protein)(-1) for amphetamine, and 153 nmol min(-1) (mg of protein)(-1) for N-methylhydroxylamine. Administration of synthetic hydroxylamines of amphetamine and methamphetamine to primary rat neuronal cultures produced frank cell toxicity. Compared with amphetamine or the oxime of amphetamine, the hydroxylamines were significantly more toxic to primary neuronal cells. The benzamidoxime reductase is therefore involved in the detoxication of these reactive hydroxylamines.

R-Deprenyl: Pharmacological Spectrum of its Activity

“…This finding also raises the possibility of retro-reduction of DNO metabolite to the parent compound in case of R-deprenyl treatment. Previously, the reduction of N-hydroxylamines by benzamidoxime reductase and human liver microsomes to their parent amines has been demonstrated [15]. This reduction mechanism may also apply in case of N-oxide metabolites formed from tertiary amines.…”

CHARACTERIZATION AND PARTIAL PURIFICATION OF THE RAT AND HUMAN ENZYME SYSTEMS ACTIVE IN THE REDUCTION OFN-HYDROXYMELAGATRAN AND BENZAMIDOXIME

“…The reductase system has been shown to be responsible for the N-reduction of other aliphatic hydroxylamines such as N-methylhydroxylamine, as well as the clinically relevant detoxification process of the hydroxylamines of methamphetamine and amphetamine (Clement et al, 2000).…”

Assessment of the N‐oxidation of deprenyl, methamphetamine, and amphetamine enantiomers by chiral capillary electrophoresis: An in vitro metabolism study

“…(The formation of phenylpropanone was not investigated.) In other studies, a further metabolic transformation of hydroxylamines, namely the reduction of amphetamine-hydroxylamine to amphetamine, and methamphetamine-hydroxylamine to methamphetamine by a cytochrome P (CYP)-dependent metabolic pathway, have also been reported [20,21].…”

“…In the latter pathway conversion of methamphetamine-hydroxylamine to amphetamine-hydroxylamine and its further reduction to amphetamine was assumed. Components of a reductase system, termed benzamidoxime reductase, responsible for the reduction of N-hydroxylated derivatives have been identified [20,21]. The system contains benzamidoxime reductase, which is a CYP450 enzyme, cytochrome b 5 , and NADH-cytochrome b 5 -reductase, and is suggested to be involved both in the retroreduction of the N-oxygenated metabolites to the parent compounds, and in demethylation processes [20,21].…”