Androstanediol glucuronide (Adiol G) has been reported to be a marker of peripheral androgen metabolism and action. It consists of two isomers, Adiol 3-G and Adiol 17-G. Adiol G is formed from unconjugated precursors by the enzyme glucuronyl transferase. To determine the likely source of Adiol G formation in man, we developed a glucuronyl transferase assay and measured the activity of this enzyme in human liver, abdominal and scalp skin, and prostate. In human liver, glucuronyl transferase activity was linear with respect to time (up to 120 min) and tissue concentration (up to 1 mg/ml). Apparent Michaelis-Menten constant Km (micromolar) and maximum velocity (Vmax) (picomoles per mg/30 min) were 5.6 and 140 for dihydrotestosterone, 8.9 and 1300 for androstanediol, and 3.1 and 46 for androsterone, respectively. Conversion of androstanediol to Adiol G (/0.5 mg tissue.30 min) was 5.8-13.2%. Over 80% of the Adiol G formed in human liver was Adiol 17-G, similar to what has been previously found in human serum. Glucuronyl transferase activity was present at low levels in human prostate (conversion of androstanediol to Adiol G was 0.04-4.6%/50 mg tissue.120 min). Analogous conversion rates (/50 mg tissue.120 min) for human scalp skin were 0.2-0.4% and for human abdominal skin were 0.07-0.14%. Although dihydrotestosterone may be converted to androstanediol in peripheral tissues such as skin and prostate, our results suggest that the principal site of androgen conjugation to glucuronic acid is the liver. The present results cast doubt upon the role of androstanediol glucuronide as a specific marker of cutaneous androgen metabolism.