富马酸丙酚替诺福韦片中有关物质测定及杂质鉴定  

Determination of Related Substances and Identification of Impurities in Tenofovir Alafenamide Fumarate Tablets

作　　者：章海云 金秉德 王玲兰 宿亮 徐彬滨 朱跃芳 ZHANG Haiyun;JIN Bingde;WANG Linglan;SU Liang;XU Binbin;ZHU Yuefang(Zhuzhou Qianjin Pharmaceutical Co.,Ltd.,Zhuzhou,Hunan,China 412000;Zhuzhou Institute for Food and Drug Control,Zhuzhou,Hunan,China 412000)

机构地区：[1]株洲千金药业股份有限公司,湖南株洲412000 [2]湖南省株洲市食品药品检验所,湖南株洲412000

出　　处：《中国药业》2024年第7期53-58,共6页China Pharmaceuticals

基　　金：2020年度湖南省自然科学基金[2022JJ80122]。

摘　　要：目的建立检测富马酸丙酚替诺福韦片中有关物质含量的高效液相色谱法,以及鉴定制剂中未知湿热降解杂质化学结构的二维液相色谱-三重四极杆串联质谱法。方法有关物质测定中,色谱柱为Waters XTerra MS C_(18)柱(250 mm×4.6 mm,5μm),流动相为0.02 mol/L磷酸盐(1 mol/L氢氧化钾溶液调pH至6.0)-甲醇(95∶5,V/V)-甲醇(梯度洗脱),流速为1.0 mL/min,检测波长为260 nm,柱温为35℃,进样量为20μL。化学结构鉴定中,色谱柱为Agilent Zorbax Eclipse Plus C_(18)柱(50 mm×3.0 mm,1.8μm),流动相为0.1%甲酸水溶液-甲醇(55∶45,V/V),流速为0.4 mL/min,柱温为35℃,采用电喷雾离子源(ESI),正离子模式,质荷比(m/z)扫描范围为100~1000,雾化气体(氮气,N_(2))压力为45 psi,干燥气(N2)流速为6 L/min,温度为350℃,碎裂电压为150 V,二级质谱碰撞裂解能量为30 eV。根据保留时间、分子量及二级碎片识别湿热降解杂质。结果丙酚替诺福韦与5个降解杂质A-E的定量限分别为0.4,0.4,2.0,2.0,0.8,0.8 ng。5个降解杂质A-E的杂质校正因子分别为0.61,0.70,3.16,0.75,0.85;平均加样回收率分别为100.20%,95.23%,102.36%,94.48%,97.26%,RSD分别为0.50%,0.23%,0.89%,1.02%,1.67%(n=9)。未知湿热降解杂质分别为杂质D乳糖加合物及丙酚替诺福韦乳糖加合物。结论该方法专属性强、灵敏度好、准确度高,可用于富马酸丙酚替诺福韦片中有关物质的含量测定,通过特征碎片离子可快速识别丙酚替诺福韦乳糖加合物类杂质。Objective To establish a high-performance liquid chromatography method for the content determination of related substances in Tenofovir Alafenamide Fumarate Tablets,and to establish a two-dimensional high-liquid chromatography-triple quadrupole mass spectrometry(2D-HPLC-MS/MS)method for the identification of the chemical structure of unknown hygrothermal degradation impurities.Methods For the content determination of related substances,the chromatography column was Waters XTerra MS C_(18) column(250 mm×4.6 mm,5μm),the mobile phase was 0.02 mol/L phosphate(adjusted to pH 6.0 with 1 mol/L potassium hydroxide solution)-methanol(95∶5,V/V)-methanol(gradient elution),the flow rate was 1.0 mL/min,the detection wavelength was 260 nm,the column temperature was 35℃,and the injection volume was 20μL.For the chemical structure identification,the chromatographic column was Aglient Zorbax Eclipse Plus C_(18) column(50 mm×3.0 mm,1.8μm),the mobile phase was 0.1% formic acid aqueous solution-methanol(55∶45,V/V),the flow rate was 0.4 mL/min,the column temperature was 35℃,the positive ion mode was adopted with the electric spray ion source(ESI),the mass charge ratio(m/z)scanning range was 100-1000,the pressure of atomized gas(nitrogen,N_(2))was 45 psi,the flow rate of dry gas(N2)was 6 L/min,the temperature was 350℃,the fragmentation voltage was 150 V,and the secondary mass spectrometry collision pyrolysis energy was 30 eV.The hygrothermal degradation impurities were identified based on retention time,molecular weight,and secondary fragments.Results The limits of quantification(LOQ)of tenofovir alafenamide and five degradation impurities A-E were 0.4,0.4,2.0,2.0,0.8,and 0.8 ng,respectively.The relative correction factors of impurities A-E were 0.61,0.70,3.16,0.75,and 0.85,respectively.The average recoveries of impurities A-E were 100.20%,95.23%,102.36%,94.48%,97.26% with RSDs of 0.50%,0.23%,0.89%,1.02%,and 1.67%(n=9),respectively.The unknown hygrothermal degradation impurities were identified as impurity D lactose add

关 键 词：富马酸丙酚替诺福韦片 高效液相色谱法 二维液相色谱-三重四极杆串联质谱法 有关物质 乳糖加合物 化学结构鉴定 

分 类 号：R917[医药卫生—药物分析学] R978.7[医药卫生—药学]