An investigation was conducted into the stereochemistry of the equine urinary metabolites of 17alpha-methyltestosterone observed after oral administration. Standards of the complete range of C3/C5/C16 stereoisomeric 17alpha-methylandrostane-3,17beta-diols, 17alpha-methylandrostane-3,16,17beta-triols and 17alpha-hydroxymethylandrostane-3,17beta-diols were purchased or synthesised, and were used to unequivocally identify the absolute structures of the metabolites. Phase I metabolism was found to involve combinations of Delta(4)-3-ketone reduction with both 5alpha,3beta- and 5beta,3alpha-stereochemistry, hydroxylation at C16 with both 16alpha- and 16beta-stereochemistry and hydroxylation of the 17alpha-methyl substituent. Phase II metabolism involved mainly sulfation with a lesser degree of beta-glucuronidation.