Preliminary study of the metabolism of 17α-methyltestosterone in horses utilizing gas chromatography–mass spectrometric techniques

Abstract

Little is known about the metabolism of 17α-alkyl anabolic steroids in horses. In this study, the metabolism of 17α-methyltestosterone is investigated by oral administration of a (1 + 1) mixture of the steroid and its deuteriated analogue. Both compounds were synthesized from dehydroisoandrosterone (DHA), using a Grignard reaction followed by an Oppenauer oxidation. Post-administration urine extracts were analysed by gas chromatography–mass spectrometry (GC–MS) using both electron impact (EI) and chemical ionization (CI). Interpretation of the data was facilitated by observation of the fragment ions present in the mass spectra. Notably, the D-ring fragment ions were indicative of 15- or 16-hydroxylation, where 16-hydroxy metabolites showed ion pairs at m/z 218/221 and at m/z 231/234 while 15-hydroxy compounds gave the 231/234 ion pair alone. Unaltered D-rings showed fragment ions at m/z 143/146. The data showed that the main phase 1 metabolic processes were partial and complete reduction of the 3-oxo-4-ene group, 15-hydroxylation, 16-hydroxylation, 17-epimerization and hydroxylation at at least two other undetermined sites, postulated as the 6 and 11 positions. Phase 2 metabolism, in the form of glucuronide and sulfate formation, was also common. The information provided by this investigation will result in improved effectiveness of confirmatory analytical procedures for 17α-alkyl anabolic steroids.