The in vivo tissue distribution and metabolism of tibolone was studied in different animals to further investigate the compound's tissue-specificity. Tibolone's metabolism was studied in vivo in rats and rabbits by administration of [16-3H]-tibolone and the metabolic pattern was determined in urine and faeces after oral administration to female rats and dogs. The main excretory pathway was found to be excretion in the faeces. Important phase-I metabolic routes were the reduction of the 3-keto to the 3a- or 3beta-hydroxy functions with a preference for 3alpha-OH in rats and for 3beta-OH in dogs. To a lesser extent, hydroxylation reactions at C2 and C7, and a shift of the delta5(10)-double bond to a delta4(5)-position also occurred. The main phase-II metabolic route was sulphate conjugation of the hydroxyl groups at C3 and C17. Since the oxidation reactions form only a minor part of the metabolism of tibolone, it is concluded that the cytochrome P450 enzymes do not play an important role in tibolone's metabolism. For both phases, quantitative differences were found between the species. In human similar metabolites are found. Profiling of the target organs in female rats and rabbits showed a tissue-specific distribution of metabolites. The majority of the metabolites existed as sulphate conjugates and no glucuronidated conjugates were observed. The same metabolites were found in both the circulation and the tissues. However, different tissues had quantitatively different metabolic profiles.