1. The disposition of nalmefene in rat and dog was studied using in vitro and in vivo methodology. In vitro metabolite profiles were obtained following incubation of nalmefene with liver microsomes and biological fluids were assayed to profile in vivo metabolites. Characterization of metabolites was accomplished using hplc, co-chromatography with synthetic standards, or LC/MS. 2. In rat, tissue distribution and metabolite plasma concentration-time data were obtained following intravenous bolus dosing of nalmefene. 3. The results indicate that the primary phase I metabolite of nalmefene from liver microsome incubations was the N-dealkylated metabolite, nornalmefene. Quantitative metabolite production was rat >> dog. In vivo, nornalmefene glucuronide was the major metabolite in rat urine, whereas nalmefene glucuronide(s) were predominant in dog urine. 4. More than 90% of the radioactive dose was recovered in the rat excreta and tissues 24 h after an intravenous bolus dose of 14C-nalmefene, with no apparent organ-specific retention of radioactivity. 5. Pharmacokinetic analysis of the rat plasma metabolite data indicated that terminal half-lives for nalmefene and nornalmefene were comparable (approximately 1 h). However, Cmax and AUC of nornalmefene were < or = 7% that of corresponding nalmefene values.