RESUMO
A GLP-validated, sensitive and specific LC-MS-MS method for the quantification of paclitaxel and its 6-alpha- and 3'-p-hydroxy metabolites is presented. A 0.400 ml plasma aliquot is spiked with a (13)C(6)-labeled paclitaxel internal standard and extracted with 1 ml methyl-tert.-butyl ether. The ether is evaporated and the residue is reconstituted in 130 microl of 30% aqueous acetonitrile (ACN) containing 0.1% trifluoroacetic acid. Isocratic HPLC analysis is performed by injecting 50 microl of the reconstituted material onto a 50x2.1 mm C(18) column with an ACN-water-acetic acid (50:50:0.1) mobile phase at 200 microl/min flow. Detection is by positive ion electrospray followed by multiple reaction monitoring of the following transitions: paclitaxel (854>509 u), 6-alpha-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 13C(6) 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.
