Biotransformations of R-(+)-pulegone and menthofuran in vitro: chemical basis for toxicity.

Incubation of R-(+)-pulegone(I) with PB-induced rat liver microsomes in the presence of NADPH resulted in the formation of menthofuran (II) and 2-Z-[2'-keto-4'-methylcyclohexylidene] propanol (III, 9-hydroxy pulegone) as the major and minor metabolites, respectively. When isopulegone (IV) was used as the substrate, the major metabolite formed was shown to have identical GC-MS fragmentation pattern to that of synthetic 2-[2'-keto-4'-methylcyclohexyl]prop-2-en-1-ol (V) and the minor metabolite was shown to be menthofuran (II). Transformation of menthofuran (II) by microsomes in the presence of NADPH yielded a metabolite identified as 2-Z-(2'-keto-4'-methyl cyclohexylidene) propanal (VI, pulegone-8-aldehyde). Formation of this alpha, beta -unsaturated aldehyde was further confirmed by trapping it as cinnoline derivative by adding semicarbazide to the assay medium. The toxicity mediated by pulegone is discussed in the light of these observations.