Structural characterization of mono- and dihydroxylated metabolites of paclitaxel in rat bile using liquid chromatography/ion spray tandem mass spectrometry.

The capability of high performance liquid chromatography/ion spray mass spectrometry (HPLC/ISP-MS) and HPLC/ISP-tandem mass spectrometry (HPLC/ISP-MS/MS) were investigated to achieve mass separation as well as structural characterization of taxol metabolites directly in rat bile, without their previous isolation. HPLC/ISP-MS yielded information on the molecular weights of several hydroxylated derivatives while HPLC/ISP-MS/MS allowed the on-line structural characterization of all metabolites present in different ratios in rat bile. The approach led to the extraction of nine metabolites and their distinction from the other endogenous contaminants. These metabolites were recognized as three dihydroxytaxols, four monohydroxytaxols, one deacetyltaxol and one containing the taxane ring. Among the derivatives, we were able to identify four new metabolites of paclitaxel belonging to the dihydroxy and monohydroxy series, never previously detected. HPLC/ISP-MS/MS enabled the classification of all di- and monohydroxy isomers. These results demonstrate that the high sensitivity of this method, based on the combined use of tandem mass spectrometry with chromatographic separation, can be considered as offering a valid approach to the detection of new taxol derivatives directly in biological fluids.