Measurement of paclitaxel and its metabolites in human plasma using liquid chromatography/ion trap mass spectrometry with a sonic spray ionization interface

• A quantitative liquid chromatography/ion trap mass spectrometry method for the simultaneous determination of paclitaxel, 6α-hydroxypaclitaxel and p-3'-hydroxypaclitaxel in human plasma has been developed and validated. 6α-,p-3'-Dihydroxypaclitaxel was also quantified using paclitaxel as a reference and docetaxel as an internal standard. The substances were extracted from 0.500 mL plasma using solid-phase extraction. The elution was performed with acetonitrile and the samples were reconstituted in the mobile phase. Isocratic high-performance liquid chromatography analysis was performed by injecting 50 µL of reconstituted material onto a 100 × 3.00 mm C12 column with a methanol:1% trifluoroacetic acid/ammonium trifluoroacetate in H2O 70:30 mobile phase at 350 µL/min. The [M+H]+ ions generated in the sonic spray ionization interface were isolated and fragmented using two serial mass spectrometric methods: one for paclitaxel (transition 854 → 569 & 551) and the dihydroxymetabolite (transition 886 → 585 & 567) and one for the hydroxy metabolites (transition 870 → 585 & 567; transition 870 → 569 & 551) and docetaxel ([M+Na]+, transition 830 → 550). Calibration curves were created ranging between 0.5 and 7500 ng/mL for paclitaxel, 0.5 and 750 ng/mL for 6α-hydroxypaclitaxel, and 0.5 and 400 ng/mL for p-3'-hydroxypaclitaxel. Adduct ion formation was noted and investigated during method development and controlled by mobile phase optimization. In conclusion, a sensitive method for simultaneous quantification of paclitaxel and its metabolites suitable for analysis in clinical studies was obtained.

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