液质联用法测定培唑帕尼中两种基因毒性杂质的含量

Determination of the contents of two genotoxic impurities in pazopanib by high performance liquid chromatography-tandem mass spectrometry method

目的建立高效液相色谱-串联质谱法(HPLC-MS/MS法)测定培唑帕尼中两种基因毒性杂质2,3-二甲基-2H-吲唑-6-胺盐酸盐(杂质A)和2,3-二甲基-6-硝基吲唑(杂质C)的含量。方法采用HPLC-MS/MS方法,使用Poroshell 120 EC-C18色谱柱(100 mm×4.6 mm,2.7μm),以体积分数0.1%甲酸水溶液-乙腈为流动相梯度洗脱,流速0.5 mL·min-1,柱温40℃,进样量10μL,正离子模式采集分析,杂质A母离子和子离子的质核比分别是162.0和147.1,杂质C母离子和子离子的质核比分别是192.0和146.1。结果4批培唑帕尼中杂质A含量为0.06×10^(-6)、0.06×10^(-6)、0.12×10^(-6)和0.05×10^(-6),杂质C含量为0.08×10^(-6)、0.07×10^(-6)、0.05×10^(-6)和0.04×10^(-6)。杂质A和杂质C的线性方程分别为y=5.13×10^(3)x+1.67×10^(2)(R=0.9991),y=8.40×10^(3)x+3.65×10^(2)(R=0.9988),平均回收率范围为93.8%~100.4%(RSD<2.0%)。系统适用性、专属性、重复性试验的RSD均小于10.0%。结论所建立的方法快捷、准确且重复性好,可用于培唑帕尼中基因毒性杂质的控制和培唑帕尼的放行标准。

Objective To establish an high performance liquid chromatography-tandem mass spectrometry method(HPLC-MS/MS method) for determination the contents of two genotoxic impurities namely 2,3-dimethyl-2 H-indazole-6-amine hydrochloride(impurity A) and 2,3-dimethyl-6-nitroindazole(impurity C) in pazopanib.Methods HPLC-MS/MS method was adopted using Poroshell 120 EC-C18column(100×4.6 mm, 2.7 μm),and the mobile phase was 0.1% formic acid aqueous solution-acetonitrile with gradient elution at 0.5 mL·min-1.The column temperature was set at 40 ℃,the injection volume was 10 μL,and the positive ion mode was collected and analyzed.The mass-to-charge ratio of the parent and daughter ions of impurity A was 162.0 and 147.1,respectively.And the mass-to-charge ratio of the parent and daughter ions of impurity C was 192.0 and 146.1,respectively.Results The contents of impurity A in the 4 batches of pazopanib were 0.06×10^(-6),0.06×10^(-6),0.12×10^(-6) and 0.05×10^(-6),and the contents of impurity C were 0.08×10^(-6),0.07×10^(-6),0.05×10^(-6) and 0.04×10^(-6).The regression equations of impurity A and impurity C were y=5.13×10^(3)x+1.67×10^(2)(R=0.999 1) and y=8.40×10^(3)x+3.65×10^(2)(R=0.998 8),respectively, and the average recovery range was within 93.8%-100.4%(RSD<2.0%).The RSD of the system suitability, specificity, and repeatability tests were all less than 10.0%.Conclusion The established method is fast, accurate and reproducible, which can be used for the control of genotoxic impurities in pazopanib and the release standard of pazopanib.