Metabolism of (R)-(+)-Pulegone in F344 rats

(R)-(+)-Pulegone, a monoterpene ketone, is a major component of pennyroyal oil. Ingestion of high doses of pennyroyal oil has caused severe toxicity a nd occasionally death. Studies have shown that metabolites of pulegone were responsible for the toxicity. Previous metabolism studies have used high, near lethal doses and isolation and analysis techniques that may cause degr adation of some metabolites. To clarify these issues and further explore th e metabolic pathways, a study of C-14-labeled pulegone in F344 rats at dose s from 0.8 to 80 mg/kg has been conducted. High-pressure liquid chromatogra phy (HPLC) analysis of the collected urine showed the metabolism of pulegon e to be extensive and complex. Fourteen metabolites were isolated by HPLC a nd characterized by NMR, UV, and mass spectroscopy. The results demonstrate d that pulegone was metabolized by three major pathways: 1) hydroxylation t o give monohydroxylated pulegones, followed by glucuronidation or further m etabolism; 2) reduction of the carbon-carbon double bond to give diastereom eric menthone/isomenthone, followed by hydroxylation and glucuronidation; a nd 3) Michael addition of glutathione to pulegone, followed by further meta bolism to give diastereomeric 8-(N-acetylcystein-S-yl)menthone/isomenthone. This 1,4-addition not only took place in vivo but also in vitro under cata lysis of glutathione S-transferase or mild base. Several hydroxylated produ cts of the two mercapturic acids were also observed. Contrary to the previo us study, all but one of the major metabolites characterized in the present study are phase II metabolites, and most of the metabolites in free forms are structurally different from those previously identified phase I metabol ites.