Quantitative Determination Of Total And Unbound Paclitaxel In Human Plasma By Liquid Chromatography-tandem Mass Spectrometry

Background Abraxane was created by American Biosciences Inc,and was a tumor targeting drug dosage form by nano technology for the first time.It was the combination of paclitaxel and human plasma albumin with an average diameter of 130 nm particles.Compared with the solvent-based paclitaxel,Abraxane was manufactured without toxic solvents and used the natural unique transport mechanisms of albumin（gp60-cellar proteins-SPARC）to make more distribution of taxol in tumor tissues and achieve higher concentration within the tumor cells,so as to guarantee the safety and efficacy of this drug.Objective An accurate and sensitive liquid chromatography/tandem mass spectrometric（LC-MS/MS）assay was developed and validated for the quantification of both unbound and total paclitaxel in plasma following treatment with two formulations of paclitaxel.The method was applied to pharmacokinetics study of total and free paclitaxel and the bioequivalence study.Method This assay used an electrospray ionization source（ESI source）and multiple reaction monitoring mode（MRM）.Chromatography was carried out with a Agilent C₁₈ column（50×2.1 mm,1.8μm）and the running time was 1.7 min.The mobile phase included acetonitrile and 0.1mmol?L^-1 lithium carbonate solution（50:50）.D₅-paclitaxel was internal standard of the deterministation of total paclitaxel and paclitaxel was extracted from plasma by protein precipitation with acetonitrile.Rapid equilibrium dialysis was used to separate sample and paclitaxel was extracted from buffers with docetaxel as internal standard by liquid-liquid extraction in the deterministation of free paclitaxel.Acquisition of mass spectrometric data was performed with the transitions of m/z 860.3→m/z292.2 for paclitaxel、m/z 865.4→m/z 297.2 for d₅-paclitaxel and m/z814.4→m/z 288.2 for docetaxel.Results The linear range of paclitaxel in human plasma（total concentration）and retentates was both 5.00 to 15000 ng?mL^-1 and the linear range of paclitaxel in the dialysates was 0.100 to 100 ng?mL^-1.For total paclitaxel,the intra-and inter-day precisions were within 10.3%,and accuracy ranged from-0.3%to 14.1%.For free paclitaxel,the intra-and inter-day precisions were within 11.8%,and accuracy ranged from-7.6%to 6.6%.The method was successfully applied to pharmacokinetics study of total and free paclitaxel and the bioequivalence study of Abraxane.Free concentration=dialysate concentration/retentate concentration﹡total concentration.The 90%confidence interval of the test and reference geometric mean ratio of AUC_0-72,AUC_0-∞,C_max for Abraxane were both in the range of 80%to 125%.The two formulations were bioequivalent.Conclusion This method was developed to determine the total paclitaxel and free paclitaxel in human plasma,with a short running time and high sensitivity.The results of method validation showed this method was accurate and reliable.It was successfully applied to pharmacokinetics study of total and free paclitaxel and the bioequivalence study.