A specific HPLC method with UV detection was used to investigate the disposition of morphine and its metabolites in the in-situ rat isolated perfused liver preparation. Livers of male Sprague-Dawley rats (n = 4) were perfused under single pass conditions with protein- and erythrocyte-free perfusate, containing 2.66 microM morphine, for up to 90 min. The concentration of morphine, normorphine and morphine-3-glucuronide (M3G) in outflow perfusate, and the biliary excretion of M3G and normorphine glucuronide, all reached steady-state levels within 15-20 min after commencing perfusion. At steady-state, the mean (+/- s.d.) extraction ratio of morphine was 0.87 +/- 0.06 and clearance (26.0 +/- 1.7 mL min-1) approached perfusate flow rate (30 mL min-1). Although M3G was the main metabolite, accounting for 72.8 +/- 12.7% of eliminated morphine, a significant proportion (21.6 +/- 13.5%) was N-demethylated to normorphine and was recovered as unchanged normorphine in outflow perfusate and normorphine glucuronide in bile. The biliary extraction ratio of hepatically-formed M3G was 0.61 +/- 0.31. Results from an additional six experiments, in which livers were perfused with 1.33 and 2.66 microM of morphine for 30 min each in a balanced cross-over manner, indicated that the disposition of morphine and its metabolites was approximately linear within this concentration range.