To identify 4 unknown metabolites of benproperine (BPP, 1) in human urine after a po dose, and to investigate the antitussive effect of monohydroxylate metabolites.
The putative metabolite references were prepared using chemical synthesis. Their structures were identified using 1H and 13C nuclear magnetic resonance, and mass spectrometry. The metabolites in human urine were separated and assayed using liquid chromatography-ion trap mass spectrometry (LC/MS/MS), and further confirmed by comparison of their mass spectra and chromatographic retention times with those of synthesized reference substances. The antitussive effects of metabolites were evaluated on coughs induced by 7.5% citric acid in conscious guinea pigs.
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. Four urinary metabolites in human urine showed peaks with the same chromatographic retention times and mass spectra in LC/MS/MS as synthetic 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 min test using citric acid, but did not reduce the number of coughs during the 5 min immediately after the test in conscious guinea pigs.
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.
Sweetman, SC, editor. Martindale, the Extra Pharmacopoeia 34th Edition. London: Pharmaceutical Press; 2005. p 1115.
Kamei J, Ogawa M, Kasuya Y . Monoamines and the mechanisms of action of antitussive drugs in rats. Arch Int Pharmacodyn Ther 1987; 290: 117–27.
Du ZM, Huang HH, Chen XY, Zhong DF . Study on hydroxylated metabolites of benproperine in human urine. Acta Pharm Sin 2000; 35: 916–20.
Clarke NJ, Rindgen D, Korfmacher WA, Cox KA . Systematic LC/MS metabolite identification in drug discovery. Anal Chem 2001; 73: 430A–439A.
Desai RB, Schwartz MS, Matuszewski BK . The identification of three human metabolites of a peptide-doxorubicin conjugate using HPLC-MS-MS in positive and negative ionization modes. J Chromatogr Sci 2004; 42: 317–22.
Tevell A, Bondesson U, Torneke K, Hedeland M . Identification of some new clemastine metabolites in dog, horse, and human urine with liquid chromatography/tandem mass spectrometry. Rapid Commun Mass Spectrom 2004; 18: 2267–72.
Fei X, Lin H . Preparation of benproperine. Hei Long Jiang Yi Yao 1999; 112: 194–5.
Li Y, Chen S, Zhong D, Gan C, Min L, Sun Y, et al. Synthesis of R(+)- and S(−)-benproperine phosphate and their antitussive activity. Chin J Med Chem 2004; 14: 19–22.
Soliman SE, Bassily RW, El-Sokkary RI, Nashed MA . Acetylated methyl glucopyranuronate trichloroacetimidate as a glycosyl donor for efficient synthesis of disaccharides. Carbohydr Res 2003; 338: 2337–40.
Karlsson JA, Ianner AS, Persson CG . Airway opioid receptors mediate inhibition of cough and reflex bronchoconstriction in guinea pig. J Pharmacol Exp Ther 1990; 252: 863–8.
Chen S, Min L, Li Y, Li W, Zhong D, Kong W . Anti-tussive activity of benproperine enantiomers on citric-acid-induced cough in conscious guinea-pigs. J Pharm Pharmacol 2004; 56: 277–80.
Stachulski AV . Glucuronidation of alcohols using the bromosugar–iodonium reagent method. Tetrahedron Lett 2001; 42: 661–13.
Ferguson JR, Harding JR, Lumbard KW, Scheinmann F, Stachulski AV . Glucuronide and sulfate conjugates of ICI 182,780, a pure anti-estrogenic steroid. Order of addition, catalysis and substitution effects in glucuronidation. Tetrahedron Lett 2000; 41: 389–91.
Braga PC, Bossi R, Piatti G, Dal Sasso, M . Antitussive effect of oxatomide on citric acid-induced cough in conscious guinea pig. Arzneim Forsch 1993; 43: 550–3.
Forsberg K, Karlsson JA, Theodorsson E, Lundberg JM, Persson CG . Cough and bronchoconstriction mediated by capsaicin-sensitive sensory neurons in the guinea-pig. Pulm Pharmacol 1988; 1: 33–9.
Ricciardolo FL . Mechanisms of citric acid-induced bronchoconstriction. Am J Med 2001; 111: 18S–24S.
Wong CH, Matai R, Morice AH . Cough induced by low pH. Respir Med 1999; 93: 58–61.
Hay DW, Giardina GA, Griswold DE, Underwood DC, Kotzer CJ, Bush B, et al. Nonpeptide tachykinin receptor antagonists. III. SB 235375, a low central nervous system-penetrant, potent and selective neurokinin-3 receptor antagonist, inhibits citric acidinduced cough and airways hyper-reactivity in guinea pigs. J Pharmacol Exp Ther 2002; 300: 314–23.
Project supported by Grant 2003AA2Z347C of the 863 Program of China.
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- high performance liquid chromatography
- mass spectrometry
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