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

We have developed a rapid, sensitive and selective LC-MS method for the simultaneous assay of bupropion and its metabolite hydroxybupropion during its intestinal absorption, studied with the rat everted gut sac model. The method was validated in the concentration range of 1-15 microM (0.024-3.58 microg/mL) for bupropion and 0.005-1 microM (0.00127-0.25 microg/mL) for hydroxybupropion with 10 microL injected. Bupropion is used as a probe for the activity of the CYP2B6 isoenzyme of the P450 family of enzymes in man. Its major metabolite hydroxybupropion was found in the serosal media of the gut sac showing that the isoenzyme of the 2B group was active in the intestinal mucosa and metabolized bupropion during its passage across the mucosa. The metabolite was also quantified in the mucosal media indicating its ability to cross the apical membrane of the epithelial cells.