罗米司亭中聚山梨酯80含量高效液相色谱-电喷雾检测器检测方法的建立、验证及初步应用  

Development,validation and preliminary application of highperformance liquid chromatography coupled with charged aerosol detector method for determination of polysorbate 80 in romiplostim

作　　者：郭莹[1] 李响[1] 毕华[1] 裴德宁[1] 李学普 秦玺[1] 陶磊[1] 史新昌[1] 杨靖清[1] 周勇[1] GUO Ying;LI Xiang;BI Hua;PEI Dening;LI Xuepu;QIN Xi;TAO Lei;SHI Xinchang;YANG Jingqing;ZHOU Yong(National Institutes for Food and Drug Control,Key Laboratory of the Ministry of Health for Research on Quality and Standardization of Biotech Products,Beijing 100050,China;不详)

机构地区：[1]中国食品药品检定研究院卫生部生物技术产品检定方法及其标准化重点实验室,北京100050 [2]赛默飞世尔科技(中国)有限公司,上海201203

出　　处：《中国生物制品学杂志》2024年第11期1361-1366,共6页Chinese Journal of Biologicals

基　　金：国家质量基础设施资助(2021YFF0600804)。

摘　　要：目的建立重组蛋白药物罗米司亭中聚山梨酯80含量高效液相色谱(high-performance liquid chromatography,HPLC)-电雾式检测器(charged aerosol detector,CAD)检测方法,并对方法进行验证及初步应用,以期为重组蛋白药物制剂的质量控制提供参考。方法通过对流动相的筛选以及方法洗脱梯度和CAD参数的优化建立聚山梨酯80含量的HPLC-CAD方法,并对方法的专属性、重复性、准确性进行验证,确定方法的检测限、定量限和线性范围。采用建立的方法检测2批次罗米司亭样品中的聚山梨酯80含量。结果方法优化后的色谱条件如下:色谱柱为Acclaim Surfactant Plus(150 mm×4.6 mm,3μm);流动相A为含0.1%甲酸的水溶液,流动相B为含0.1%甲酸的异丙醇;洗脱梯度为0 min 80%A,1.8 min 80%A,2.0 min 66.5%A,3.0 min 66.5%A,3.5 min 0%A,8.5 min 0%A,9.0 min 80%A,15.0 min 80%A;流速为0.6 mL/min;进样量为10μL;柱温为25℃;CAD雾化温度为50℃,幂律函数值(power function value,PFV)为1.25。该方法样品基质和空白溶液中无其他色谱峰干扰;检测低(10 mg/L)、中(50 mg/L)、高(80 mg/L)浓度聚山梨酯80峰面积的RSD分别为4.32%、1.97%和3.46%,平均加标回收率分别为95.7%、99.0%和98.9%;聚山梨酯80在浓度5.0~100.0 mg/L范围内具有良好的线性关系(R^(2)=1.0000),方法的检测限和定量限分别为1.0和2.0 mg/L。采用建立的方法检测2批次罗米司亭样品中聚山梨酯80含量分别为33.20和32.28 mg/L。结论建立的罗米司亭中聚山梨酯80含量HPLC-CAD检测方法灵敏度高,专属性、重复性、准确性良好,可用于重组蛋白药物质量控制过程中聚山梨酯80含量的检测。Objective To develop,validate and preliminarily apply a high-performance liquid chromatography(HPLC)-charged aerosol detector(CAD)method for the quantitative analysis of polysorbate 80 in recombinant protein drug romiplostim,in order to provide a reference for the quality control of recombinant protein drug preparations.Methods The HPLCCAD determination method for polysorbate 80 content was developed by screening the mobile phase and optimizing the elution gradient and CAD parameters.The specificity,repeatability and accuracy of the method were validated,and the limit of detection(LOD),limit of quantitation(LOQ)and linear range were determined.The content of polysorbate 80 in two batches of romiplostim samples was determined by using the developed method.Results HPLC-CAD conditions after method optimization were as follows:Acclaim Surfactant Plus column(150 mm×4.6 mm,3μm)was adopted,and the column temperature was 25℃.The 0.1%formic acid in water and 0.1%formic acid in isopropanol were used as mobile phase A and B respectively,with a flow rate of 0.6 mL/min and a gradient elution(0 min 80%A,1.8 min 80%A,2.0 min66.5%A,3.0 min 66.5%A,3.5 min 0%A,8.5 min 0%A,9.0 min 80%A,15.0 min 80%A).The injection volume was10μL.The evaporative temperature for CAD was 50℃and the power function value(PFV)was 1.25.There was no interference of other chromatographic peaks in the sample matrix and blank solution in this method.The RSDs of peak area for repeatability at low(10 mg/L),medium(50 mg/L)and high(80 mg/L)concentrations were 4.32%,1.97%and 3.46%,and the mean spike recoveries were 95.7%,99.0%and 98.9%,respectively.Polysorbate 80 had a good linear relationship in the concentration range of 5.0-100.0 mg/L(R^(2)=1.0000),and the LOD and LOQ in the method were 1.0 and 2.0 mg/L respectively.The content of polysorbate 80 detected by the developed method was 33.20 and 32.28 mg/L in two batches of romiplostim samples respectively.Conclusion The developed HPLC-CAD method for the determination of polysorbate 80in romiplostim has high

关 键 词：罗米司亭 重组蛋白药物 高效液相色谱法 电雾式检测器 聚山梨酯80 

分 类 号：R917[医药卫生—药物分析学]