高效液相色谱串联质谱法同时检测人血浆伊马替尼及代谢物

A HPLC-MS/MS method for simultaneous determination of imatinib and its metabolite in human plasma

目的:建立一种同时测定人血浆中伊马替尼及其活性代谢产物N-去甲基伊马替尼的高效液相色谱串联质谱(HPLCMS/MS)法,并应用于检测胃肠间质瘤患者伊马替尼及代谢物的浓度。方法:血浆样品经甲醇沉淀蛋白后,以含0.1%甲酸的水溶液和甲醇溶液为流动相;采用梯度洗脱,Waters ACQUITY UPLC BEH C18(2.1 mm×50 mm,1.7μm)色谱柱进行分离,流速为0.3 m L·min^(-1);柱温为35℃。ESI离子源,正离子模式,多反应监测,用于定量分析的离子对为m/z 494.2→m/z 394.3(伊马替尼)、m/z 480.3→m/z 394.3(N-去甲基伊马替尼)、m/z 502.5→m/z 394.4(内标,伊马替尼-D8)。结果:伊马替尼和N-去甲伊马替尼的线性范围均为50~10 000 ng·mL^(-1),定量下限为50 ng·mL^(-1),伊马替尼及代谢物的准确度分别为97.9%~109.0%,95.5%~103.5%,日内和日间变异系数<10%。结论:本方法简便、准确、灵敏、专属性强,适用于人血浆中伊马替尼及其代谢物浓度的检测。

OBJECTIVE To develop a HPLC-MS/MS method for simultaneous determination of imatinib and N-desmethyl imatinib in human plasma,and further apply to analyzing trough concentrations of imatinib and its metabolite at steady state in Chinese patients with gastrointestinal stromal tumor （GIST）.METHODS Protein precipitation with methanol was used for sample preparation.High performance liquid chromatographic separation was performed on a Waters ACQUITY UPLC BEH C18 （2.1 mm×50 mm,1.7 μm） with 0.1% formic acid in both methanol and water as the mobile phase in a gradient manner.The flow rate was set at 0.3 mL·min-1 with a column temperature at 35℃.Electrospray ionization （ESI） with a positive-ion and multiple reactions monitoring mode was adopted.The precursor ion→product ion transitions were m/z494.2→m/z394.3 for imatinib,m/z480.3→m/z394.3 for metabolite,and m/z502.5→m/z394.4 for internal standard.RESULTS The assay had a lower limit of quantitation （LLOQ） of 50 ng·mL^-1 and was linear in range of 50-10 000 ng·mL^-1 for both imatinib and N-desmethyl imatinib.The recovery was 97.9%-109.0% for imatinib and 95.5%-103.5% for N-desmethyl imatinib.Inter-and intra-day variation coefficients were all less than 10%.CONCLUSION The established method is simple,accurate,sensitive and specific,and has been proved to be suitable for simultaneous determination of imatinib and its metabolite N-desmethyl imatinib in human plasma.