Stereoselective measurement of E- and Z-doxepin and its N-desmethyl and hydroxylated metabolites by gas chromatography-mass spectrometry

A stereoselective method of analysis of the antidepressant drug doxepin (DO X, an 85:15% mixture of E-Z stereoisomers), its principal metabolites E- an d Z-N-desmethyldoxepin (desDOX) and ring-hydroxylated metabolites in micros omal incubation mixtures is described, DOX and its metabolites were extract ed from alkalinised incubation mixtures by either: 9:1 hexane-propan-2-ol ( method 1) or 1:1 hexane-dichloromethane (method 2), derivatised with triflu oroacetic anhydride and analysed by GC-MS with selected ion monitoring. Bot h methods were suitable for the analysis of individual desDOX isomers as in dicated by correlation coefficients of greater than or equal to 0.999 for c alibration curves constructed between 50 and 2500 nM, and good within-day p recision at 125 nM (C.V. less than or equal to 14%) and 1000 nM(C.V. less t han or equal to 8%). Method 1, however, was unsuitable for the analysis of ring-hydroxylated metabolites of DOX, whereas the hydroxylated metabolites of E-DOX and E-desDOX (generated in situ) were extracted by method 2 with a C.V. of ca. 13%. This is the first assay method that permits the simultane ous measurement of desDOX and hydroxylated metabolites of DOX in microsomal mixtures. (C) 1999 Elsevier Science B.V. All rights reserved.