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目的:建立大鼠体内去甲基斑蝥素的 HPLC-MS 分析方法。方法:选用烟尿酸为内标,血浆样品经乙腈沉淀蛋白处理。色谱条件为 Hypersil SAX 阴离子交换色谱柱(4.6mm×250mm,5μm),甲醇-水(25∶75,含2.5%甲酸)为流动相,流速0.5mL·min~(-1),采用 HPLC-MS检测系统,AP-ESI 负离子模式,质谱检测参数如下:AP-ESI 负离子模式;雾化压力276kPa;温度350℃;保护气流量8 L·min~(-1);毛细管电压3500V;裂解电压为90V(NCTD)和120V(内标);SIM 检测,NCTD m/z185(M+H_2O-H)、烟尿酸m/z 179(M-H)。结果:血浆中去甲基斑蝥素检测方法的线性范围为0.2-20μg·mL~(-1),最低检测限可达2ng·mL~(-1)。血浆中去甲基斑蝥素的平均回收率为96.7%-100.3%,日内、日间 RSD 均小于8%。结论:本法灵敏、准确、选择性高,可用于去甲基斑蝥素的药代动力学研究。
Objective: To establish a HPLC-MS method for the determination of cantharidin in rats. Methods: Cholecalciferol was used as internal standard and plasma samples were treated with acetonitrile precipitation. The chromatographic conditions were Hypersil SAX anion exchange column (4.6mm × 250mm, 5μm), methanol-water (25:75, containing 2.5% formic acid) as mobile phase and flow rate of 0.5mL · min -1 AP-ESI negative ion mode and mass spectrometry detection parameters were as follows: AP-ESI negative ion mode; atomization pressure 276kPa; temperature 350 ℃; protective gas flow rate 8 L · min -1; capillary voltage 3500V; (NCTD) and 120 V (internal standard); SIM test, NCTD m / z 185 (M + H 2 O-H), nocodine m / z 179 (MH). Results: The linear range of detection of desmethylcantharidin in plasma was 0.2-20μg · mL -1 with the detection limit of 2ng · mL -1. The average recoveries of demethylcantharidin in plasma ranged from 96.7% to 100.3%. The intra-day and inter-day RSD were less than 8%. Conclusion: This method is sensitive, accurate and selective and can be used to study the pharmacokinetics of norcantharidin.