Specific and ultrasensitive reverse‐phase HPLC assays of the choleretic and biliary antispasmodic hymecromone (down to 0.05 ng ml⁻¹) and its glucuronide, using fluorimetric detection, and sulfate metabolities using UV detection, were developed. Sodium salt solutions of 400 mg (over 3 min) and 800 mg (over 5 min) were infused i.v. into 6–8 normal human volunteers. The half‐life of the major rate constant averaged 28 ± 2 min (SE). Subsequently, less than 0.8 per cent of the dose was eliminated with terminal halflives of 70–359 min. The apparent volume of distribution of hymecromone, referenced to the total plasma concentration, averaged 20.8 ± 1.41 (V_c, central compartment volume) and 36.4 ± 2.11 (V_ss steady state volume). Hymecromone's total body clearance averaged 1413 ± 89 ml min⁻¹. The pharmacokinetics of hymecromone were doseindependent. Only 0.3 ± 0.3 per cent unchanged hymecromone was renally excreted. Mostly dose‐independent glucuronidated drug (93 ± 4 per cent of the dose) was excreted in the urine; a smaller amount was renally excreted as the sulfate (1.4 ± 0.3 per cent of the dose). The oral bioavailability estimated from the relative areas under the hymecromone plasma concentration—time curves following oral and i.v. administration of hymecromone to six volunteer subjects showed no dose‐dependence and was 1.8 ± 0.6 per cent. However, an anomalous c. 200 per cent of the glucuronide produced by i.v. hymecromone was produced from orally administered hymecromone as determined from the ratio of the AUC values of glucuronide obtained after peroral and i.v. administration of the same dose of hymecromone to demonstrate a high first‐pass effect and implicate renal glucuronidation.