Aim:To identify 4 unknown metabolites of benproperine(BPP,1)in human urineafter a po dose,and to investigate the antitussive effect of monohydroxylatemetabolites.Methods:The putative metabolite references were prepared usingchemical synthesis.Their structures were identified using ~1H and ~(13)C nuclearmagnetic resonance,and mass spectrometry.The metabolites in human urinewere separated and assayed using liquid chromatography-ion trap mass spec-trometry(LC/MS/MS),and further confirmed by comparison of their mass spectraand chromatographic retention times with those of synthesized referencesubstances.The antitussive effects of metabolites were evaluated on coughsinduced by 7.5% citric acid in conscious guinea pigs.Results:1-[1-Methyl-2-[2-(phenylmethyl)phenoxy]-ethyl]-4-piperidinol(2),1-[1-methyl-2-[2-(phenylmethyl)phenoxy]ethyl]-3-piperidinol(3)and their glucuronides 4 and 5 were obtainedfrom chemical synthesis.Four urinary metabolites in human urine showed peakswith the same chromatographic retention times and mass spectra in LC/MS/MS assynthetic substances 2,3,4 and 5.Phosphates of compounds 2 and 3 prolongedthe latency of cough and reduced the number of coughs during the 3 rain testusing citric acid,but did not reduce the number of coughs during the 5 min imme-diately after the test in conscious guinea pigs.Conclusion:Compounds 2,3,4,and 5 were identified as the metabolites of BPP in human urine.Among them,compounds 2 and 3 are inactive in the antitussive effect. Aim: To identify 4 unknown metabolites of benproperine (BPP, 1) in human urine after a po dose, and to investigate the antitussive effect of monohydroxylate compoundstabolites. Methods: The putative metabolite references were prepared using chemical synthesis. Their structures were identified using ~ 1H and ~ (13) C nuclear magnetic resonance, and mass spectrometry. The metabolites in human urine body separated and assayed using liquid chromatography-ion trap mass spec-trometry (LC / MS / MS), and further confirmed by the comparison of their mass spectra and chromatographic retention times with those of synthesized referencesubstances.The antitussive effects of metabolites were evaluated on coughsinduced by 7.5% citric acid in conscious guinea pigs.Results: 1- [1-Methyl-2- [2- (phenylmethyl) phenoxy] -ethyl] -4-piperidinol (2), 1- [1-methyl-2- [2- (phenylmethyl) phenoxy] ethyl] -3-piperidinol (3) and their glucuronides 4 and 5 were obtained from chemical synthesis. same chromatog raphic retention times and mass spectra in LC / MS / MS assynthetic substances 2, 3, 4 and 5. Phosphates of compounds 2 and 3 prolonged the latency of cough and reduced the number of coughs during the 3 rain test using citric acid, but did not reduce the number of coughs during the 5 min imme-diately after the test in conscious guinea pigs.Conclusion: Compounds 2, 3, 4, and 5 were identified as the metabolites of BPP in human urine. Among them, compounds 2 and 3 are inactive in the antitussive effect.