UPLC-MS/MS法检测雷迪帕韦原料药中潜在基因毒性杂质残留量

Determination of Potential Genetoxic Impurity in Ledipasvir by UPLC-MS/MS

目的:建立超高效液相色谱-质谱联用法(UPLC-MS/MS)检测雷迪帕韦原料药中潜在基因毒性杂质1-(3-二甲氨基丙基)-3-乙基碳二亚胺(EDC)的残留量。方法:色谱条件:采用Agilent ZORBAX SB-C18色谱柱(75 mm×4.6 mm,3.5μm),流动相为甲醇-0.1%甲酸溶液(50∶50),流速为0.3 ml·min^(-1),柱温为30℃,进样量为5μl。质谱条件:采用电喷雾离子源(ESI),多级反应监测(MRM),正离子扫描模式,离子喷雾电压为2 500 V,离子源温度为500℃,雾化气为379.0 k Pa,辅助气为275.6 k Pa,气帘气为137.8 k Pa,碰撞气为41.3 k Pa,离子碰撞能为15 V,扫描时间为100 ms,用于定量分析的MRM离子对为m/z→156.2/86.1。结果:EDC的线性范围为0.03~2.25μg·ml^(-1)(r=0.999 0);检测限为0.03 ng,定量限为0.08 ng;平均回收率为98.3%(RSD=5.7%,n=9)。结论:本方法操作简便、快捷、结果准确,可用于雷迪帕韦原料药中潜在基因毒性杂质EDC的质量控制。

Objective： To establish a UPLC-MS/MS method for the determination of the residual quantity of potential genetoxic im- purity 1-（3-dimethylaminopropyl）-3-ethylcarbodiimide （EDC） in ledipasvir. Methods： The chromatographic conditions were as fol- lows： an Agilent ZORBAX SB-C1s（75 mm x4.6 mm, 3.5 μm） column was used with methanol-0.1% formic acid （50： 50） as the mobile phase. The flow rate was 0.3 ml·min-1. The column temperature was 30 ℃ and the injection volume was 5 ：1. The mass spectrometry conditions were as follows ： electrospray ionization source （ ESI）, multistage reaction monitoring＇ （ MRM）, positive ion scanning mode, ion spray voltage of 2 500 V, ion source temperature of 500 ℃, atomization gas of 379.0 kPa, auxiliary gas of 275.6 kPa, curtain gas of 137.8 kPa, collision gas of 41.3 kPa, ion collision energy of 15 V and scan time of 100 ms. The MRM ion pair for quantitative analysis was m/z=156. 2/86.1. Results： The linear range of EDC was 0.03-2.25 ml·min-1 （ r = 0.999 0）. The limit of detection was 0.03 ng and the limit of quantitation was 0.08 ng. The average recovery was 98.3% （RSD = 5.7% , n = 9 ）. Conclusion： The method is simple, rapid and accurate, and can be used for the cmalitv control of EDC in ledinasvir.