阿齐沙坦有关物质的LC-MS/MS测定与鉴定

目的：建立阿齐沙坦原料中有关物质的测定方法，鉴定主要降解产物并推测降解机理。方法：采用HPLC方法分离分析阿齐沙坦的有关物质，使用Inertsil ODS-SP色谱柱（150 mm×4.6 mm，5 μm），流动相A为乙腈-水-冰乙酸（20∶80∶0.1），流动相B为乙腈-水-冰乙酸（80∶20∶0.1），梯度洗脱，流速1.0 mL·min^-1，检测波长250 nm，柱温40℃；采用相同色谱条件，ESI正离子化方式LC-MS/MS进行杂质鉴定。结果：阿齐沙坦与有关物质均能有效分离；阿齐沙坦在酸中易水解，LC-MS/MS鉴定主要降解产物为杂质B（2-羰基-1-［［2'-（4，5-二氢-5-氧代-1，2，4-二唑-3-基）联苯-4-基］甲基］苯并咪唑-7-羧酸）。阿齐沙坦、杂质A（2-乙氧基-1-［［2'-（酰胺基）［1，1'-联苯基］-4-基］甲基］-1H-苯并咪唑-7-羧酸甲酯）和杂质B的检测下限及定量下限分别为8.4、8.1、8.2 ng·mL^-1和20.1、19.4、19.8 ng·mL^-1，且在各自的线性范围内线性关系良好（r>0.999，n=5）；校正因子分别为1.00、1.09、1.08。经5批样品检验，均仅检出杂质B，均小于0.05%。结论：建立的方法可对阿齐沙坦的有关物质进行定量测定。

Objective:To establish a method for the determination of related substances in azilsartan,and to identify the main degradation product and speculate mechanism of degradation.Methods:The related substances in azilsartan were separated by RP-HPLC on an Inertsil ODS-SP column(150 mm×4.6 mm,5 μm)using acetonitrile-water-acetic acid(20:80:0.1)as mobile phase A and acetonitrile-water-acetic acid(80:20:0.1)as mobile phase B with gradient elution at a flow rate of 1.0 mL·min^-1.The detection wavelength was set at 250 nm and the column temperature was 45℃.The major impurity was identified by using LC-MS/MS in positive mode with the same LC conditions.Results:The related substances in azilsartan could be separated effectively from azilsartan.The major impurity was identified using LC-MS/MS as impurity B(2-carbonyl-1-[[2'-(4,5-dihydro-5-oxo-1,2,4-oxadiazol-3-yl)biphenyl-4-yl]methyl]benzimidazole-7-carboxylate).The LOD and LOQ of azilsartan,impurity A(1-[[(2-ethoxy-(2'-amido))[1,1'-xenyl]-4-yl] methyl]-1H-benzimidazolyl-7-carboxylic methyl ether)and impurity B were 8.4,8.1,8.2 ng·mL^-1 and 20.1,19.4,19.8 ng·mL^-1,respectively.The calibration curves of azilsartan and the impurities were linear in the concentration range(r>0.999,n=5).The calibration factors were 1.00,1.09 and 1.08,respectively.Impurities B detected in 5 batches of samples were all less than 0.05%.Conclusion:The established method was accurate and effective for qualitative determination of related substances in azilsartan.