LC-MS／MS法测定全血中他克莫司浓度的方法学研究

目的：建立测定人全血中他克莫司液相色谱-串联质谱联用（LC-MS／MS）方法，并与微粒子免疫分析（MEIA）方法测定的结果进行了比较。方法：运用LC-MS／MS方法，血样经乙腈沉淀蛋白，上清液氮气吹干流动相复溶进样。色谱柱：Zorbax Extend—C18柱（150mm×4．6mm，5μm）；流动相：甲醇-乙腈-20mmol·L^-1醋酸胺溶液（80：15：5）；流速：0．4ml·min^-1。采用电喷雾离子化四极杆串联质谱，多反应检测方式测定样品的浓度。检测离子对分别为m／z821．7→m／z 768．4和m／z 821．7→m／z 786．5。结果：他克莫司在0．5～128ng·ml^-1浓度范围内，峰面积与浓度线性关系良好（r=0．9991，／2=5），最低定量限为0．5ng·ml^-1，平均回收率为99．0％，日内精密度、日间精密度的RSD分别为3．38％和4．16％。MEIA分析采用标准IMX测定方法。两种方法比较，其标准差小于3．0，表明相关性好。结论：本方法具有良好的灵敏度、准确度、精确度及专属性，可用于他克莫司血药浓度监测和人体药药动力学研究。

Determination of Tacrolimus in Patient Plasma by Liquid Chromatographic-Tandem Mass Spectrometry

Objective： To develop a liquid chromatography with tandem mass spectrometry （LC-MS/MS） method for the determination of taerolimus in paient plasma. Method ： The results of the concentration of taerolimus were determined by LC-MS/MS and mieroparticle enzyme immunoassay （MEIA） in patient plasma that were compared, protein of plasma was precipitated by aeetonitrile. The supernatant was evaporated to dryness and reconstituted with the mobile phase. A Zorbax Extend-C18 （ 150 mm× 4. 6mm,5 μm） was used,mobile phase conslaed of methanol-acetonitrile -water-formic acid （80：15：5 ） at a flow rate of 0. 4 ml.min^-1. Eleetrospray ionization （ESI） sources was applied and operated in the position ion mode. Multiple reaction monitoring （MRM） mode with the transition of m/z 821.7→ m/z 768.4 and m/z 821.7 →m/z 786. 5 were used to quantify tacrolimus. Result： The method was proved to be linear in the range of 0.5-140 ng.ml^- 1 with a regression coefficients of 0. 999 1 （ n = 5 ）. The lower quantitative limit was 0. 5 ng- ml - 1. The average recovery rate was 99. 0% （n = 5 ）. The RSD of average contents of intra-day and inter-clay was 3.38% and 4. 16% , respective- ly. The results of both these two methods could show the standard deviations were less than 3.0, the good relevance both LC-MS/MS and IMX were obtained. Conclusion： This method is accurate, precise, sensitive and specific to be suitable for eoncetration of therapeutic drug monitoring （TDM） and pharmacokinetics study of tacrolimus.