Measurement of paclitaxel in biological matrices: high-throughput liquid chromatographic–tandem mass spectrometric quantification of paclitaxel and metabolites in human and dog plasma

A GLP-validated, sensitive and specific LC–MS–MS method for the quantification of paclitaxel and its 6-α- and 3′-p-hydroxy metabolites is presented. A 0.400 ml plasma aliquot is spiked with a 13C6-labeled paclitaxel internal standard and extracted with 1 ml methyl-tert.-butyl ether. The ether is evaporated and the residue is reconstituted in 130 μl of 30% aqueous acetonitrile (ACN) containing 0.1% trifluoroacetic acid. Isocratic HPLC analysis is performed by injecting 50 μl of the reconstituted material onto a 50×2.1 mm C18 column with an ACN–water–acetic acid (50:50:0.1) mobile phase at 200 μl/min flow. Detection is by positive ion electrospray followed by multiple reaction monitoring of the following transitions: paclitaxel (854>509 u), 6-α-hydroxy paclitaxel (870>525 u), 3′-p-hydroxy paclitaxel (870>509 u) and internal standard (860>509 u). Quantification is by peak area ratio against the 13C6 internal standard. The method range is 0.117–117 nM (0.1–100 ng/ml) for paclitaxel and both metabolites using a 0.400 ml human or dog plasma sample. Analysis time per sample is less than 5 min.