酒石酸伐尼克兰中6个类苯胺基因毒性杂质的LC-MS法测定

目的：建立液相色谱-质谱法同时测定酒石酸伐尼克兰中6个类苯胺基因毒性杂质：2，3，4，5-四氢-1，5-甲桥-1H-3-苯并氮杂-6，8-二胺、2，3，4，5-四氢-1，5-甲桥-1H-3-苯并氮杂-7，8-二胺、2，3，4，5-四氢-1，5-甲桥-1H-3-苯并氮杂-7-胺、2，3，4，5-四氢-3-（三氟乙酰基）-1，5-甲桥-1H-3-苯并氮杂-6，8-二胺、2，3，4，5-四氢-3-（三氟乙酰基）-1，5-甲桥-1H-苯并氮杂-7，8-二胺和2，3，4，5-四氢-3-（三氟乙酰基）-1，5-甲桥-1H-苯并氮杂-7-胺。方法：采用Inertsil ODS-SP（250 mm×4.6 mm，5 μm）色谱柱，以[0.1%醋酸铵-0.005%甲酸水溶液（pH 6.0）]-乙腈（95：5）为流动相A，乙腈为流动相B进行线性梯度洗脱。以含5 mmol·L^-1二巯基苏糖醇的醋酸铵缓冲液（pH 7.0）-乙腈（95：5）为溶剂制备稳定的供试品溶液，在质谱电喷雾正离子化多反应监测模式下以外标法对6个类苯胺基因毒性杂质同时进行定量测定。结果：6个基因毒性杂质质量浓度在20~200 ng·mL^-1范围内线性关系良好；平均回收率为94.4%~106.0%，RSD均不超过7.4%。最低检测限为15 ng·mL^-1，最低定量限为45 ng·mL^-1，对照溶液和供试品溶液于15℃放置15 h内稳定。结论：建立的LC-MS测定法适用于酒石酸伐尼克兰中6个微量类苯胺基因毒性杂质的同时检测。

Objective:To establish an LC-MS method for the determination of six aniline-like genotoxic impurities(2,3,4,5-tetrahydro-1,5-methano-1H-3-benzazepine-6,8-diamine,2,3,4,5-tetrahydro-1,5-methano-1H-3-benzazepine-7,8-diamine,2,3,4,5-tetrahydro-1,5-methano-1H-3-benzazepine-7-amine,2,3,4,5-tetrahydro-3-(trifluoroacetyl)-1,5-methano-1H-3-benzazepine-6,8-diamine,2,3,4,5-tetrahydro-3-(trifluoroacetyl)-1,5-methano-1H-3-benzazepine-7,8-diamine,and 2,3,4,5-tetrahydro-3-(trifluoroacetyl)-1,5-methano-1H-3-benzazepine-7-amine) in varenicline tartrate.Methods:The separation was established on an Inertsil ODS-SP(250 mm×4.6 mm,5 μm)column by linear gradient elution using[0.1% ammonium acetate-0.005% formic acid buffer solution(pH 6.0)]-acetonitrile(95:5)as mobile phase A and acetonitrile as mobile phase B.Ammonium acetate buffer solution(pH 7.0)containing 5 mmol·L^-1 dithiothreitol and acetonitrile mixture(95:5)was used as solvent for the preparation of stable test solutions.The MS determination was carried out by multiple reactions monitoring mode with positive electrospray ionization.All the genotoxic impurities were quantitatively determined by an external reference method.Results:The calibration curve was linear for each compound in the range of 20-200 ng·mL^-1.The average recoveries were 94.4%-106.0%,with RSD not more than 7.4%.And the limit of detection and limit of quantitation were 15 ng·mL^-1 and 45 ng·mL^-1,respectively.The reference and test solutions were stable at 15℃ for 15 hours.Conclusion:The established LC-MS method is suitable for the simultaneous trace analysis of six aniline-like genotoxic impurities in varenicline tartrate.