Validation of a liquid chromatography-mass spectrometry method to assess the metabolism of dextromethorphan in rat everted gut sacs

A rapid, sensitive and selective liquid chromatography-mass spectrometry (LC-MS) method was developed for the simultaneous assay of dextromethorphan and its metabolites in tissue culture medium and its intestinal metabolism studied with the rat everted gut sac model. The method was validated in the concentration range of 0.1-2.5 mu M (27.1 ng/mL-0.677 mu g/mL) for dextromethorphan and 0.005-0.5 mu M for dextrorphan and 3-methoxymorphinan (1.28 ng/mL-0.128 mu g/mL) and 3-hydroxymorphinan (1.22 ng/mL-0.122 mu g/mL). The limits of quantification (LOQ) were 0.0025 mu M (12.5 fmoles, 3.4 pg, 5 mu L injected) for dextromethorphan; 0.0025 mu M for dextrorphan, 3-methoxymorphinan (24.9 fmoles, 6.4 pg injected), and 3-hydroxymorphinan (25.1 fmoles, 6.1 pg injected) with 10 mu L injected. The detection of dextrorphan and 3-methoxymorphinan showed that both the P450 isoforms CYP3A and 2D were active in the intestinal mucosa and metabolised dextromethorphan during its passage across the mucosa. (c) 2005 Elsevier B.V. All rights reserved.