The conversion of pyrrolizidine alkaloids to N-oxides and to dihydropyrrolizine derivatives by rat-liver microsomes in vitro

Abstract

The enzymatic conversion of pyrrolizidine alkaloids to dihydropyrrolizine derivatives (‘pyrrole derivatives’) and to N-oxides has been investigated. In rat liver both reactions were catalysed by an enzyme present in the microsomal fraction, typical of the mixed-function oxidases. Thus the conversion of alkaloids to pyrrole derivatives and to N-oxides required oxygen and reduced NADP and was activated by Mg 2+, Mn 2+ or Ca 2+ at concentrations of 5 · 10 −3 M. The liver microsomal system was less active in the female rat than in the male and was inhibited by carbon monoxide and by SKF 525A. The conversion of retrorsine to both pyrrolic metabolites and to N-oxides was significantly induced by pretreatment with phenobarbitone or DDT, but the induction of pyrrole derivatives was greater than that of N-oxide. The conversion of retrorsine to pyrrole derivatives was lower in liver microsomal fractions from rats which were starved or fed on protein-free diets. Differences between the rates of formation of pyrrole derivatives and N-oxides followed the inclusion of nicotinamide in the incubation mixtures containing microsomes from untreated rats, or substitution of a carbon monoxide-oxygen mixture for air during the incubation period. The K m values towards the formation of pyrrolic derivatives and N-oxides from retrorsine from normal rat-liver microsomes were similar. No conclusive evidence was found to establish whether N-oxides were intermediate in the formation of pyrrole derivatives. However, pyrrole derivatives could not be detected when N-oxides were added to the hepatic microsomal system in vitro. Rat-liver microsomal enzymes converted a number of alkaloids having either 1,2-dehydropyrrolizidine or saturated pyrrolizidine ring structures to pyrrole derivatives. The pyrrole derivatives from these two groups of alkaloids had different chemical properties. Only alkaloids in the first group were hepatotoxic, and in general the more toxic alkaloids in this group were those from which the greatest amounts of pyrrole derivatives were produced in vitro, although there were exceptions. The enzymatic conversion of retrorsine or monocrotaline to pyrrolic derivatives could not be demonstrated in lung microsomal fractions.