LC-MS/MS测定酒石酸溴莫尼定中苯胺类基因毒性杂质

Determination of Anililines Genotoxic Impurity in Brimonidine Tartrate by LC-MS/MS

目的建立LC-MS/MS方法测定酒石酸溴莫尼定中苯胺类基因毒性杂质(6-氨基喹喔啉、4-硝基邻苯二胺、6-氨基-5-溴喹喔啉)的含量。方法采用色谱柱ZORBAX Eclipse Plus C_(18)(2.1 mm×50 mm,1.8μm),流动相为0.1%乙酸水溶液(A)-乙腈(B),梯度洗脱,流速0.35 mL·min^(−1),柱温为35℃;采用Shimadzu LC30 UPLC-AB Science 4000+三重四极杆液质联用仪和多反应离子监测模式二级质谱MS/MS(ESI)源检测,正离子模式采集数据。结果6-氨基喹喔啉、4-硝基邻苯二胺、6-氨基-5-溴喹喔啉分别在9.98~199.50,9.94~198.74,10.03~200.55 ng·mL^(−1)内线性关系良好;低、中、高浓度(50,100,150 ng·mL^(−1))回收率为94.8%~110.8%,RSD均<4.2%;溶液在12 h内稳定性良好。结论本方法操作简便,结果可靠,可以用于酒石酸溴莫尼定中苯胺类基因毒性杂质(6-氨基喹喔啉、4-硝基邻苯二胺、6-氨基-5-溴喹喔啉)的控制。

OBJECTIVE To establish LC-MS/MS method for the determination of anililines genotoxic impurities(6-aminoquinoxaline,4-nitro-o-phenylenediamine,and 6-amino-5-bromoquinoxaline)in brimonidine tartrate.METHODS The method was achieved on a ZORBAX Eclipse Plus C_(18) column(2.1 mm×50 mm,1.8μm)utilizing a mobile phase of 0.1%acetic acid(A)-acetonitrile(B)with gradient elution at the flow rate of 0.35 mL·min^(−1).The temperature of column was set at 35℃.The Shimadzu LC30 UPLC-AB Science 4000+LC-MS was used to detect(ESI source)in multiple reaction monitoring positive mode.RESULTS The calibration curves of 6-aminoquinoxaline,4-nitro-o-phenylenediamine,6-amino-5-bromoquinoxaline were linear in 9.98−199.50,9.94−198.74,10.03−200.55 ng·mL^(−1).The average recoveries of the control samples at low,middle and high concentrations(50,100,150 ng·mL^(−1))were 94.8%−110.8%,all RSDs were<4.2%,solution stability was good within 12 h.CONCLUSION The method is convenient to operate and has reliable results for the determination of anililines genotoxic impurities of 6-aminoquinoxaline,4-nitro-o-phenylenediamine,6-amino-5-bromoquinoxaline in bromnidine tartrate.