双检测器串联测定聚乙二醇化人生长激素平均修饰率方法研究  

The research of a SEC-UV-RI method to determine the modifiction degree of PEG-rhGH

作　　者：李懿 张孝明 胡馨月 孙悦[1] 王绿音 陈度宇 吕萍[1] 梁成罡[1] 李晶[1] LI Yi;ZHANG Xiao-ming;HU Xin-yue;SUN Yue;WANG Lü-yin;CHEN Du-yu;LÜPing;LIANG Cheng-gang;LI Jing(Division of Hormone in National Institutes for Food and Drug Control,State Key Laboratory of Drug Regulatory Science,NMPA Key Laboratory for Quality Research and Evaluation of Biological Products,NHC Key Laboratory of Research on Quality and Standardization of Biotech Products,Beijing 102629,China)

机构地区：[1]中国食品药品检定研究院激素室,药品监管科学全国重点实验室,国家药品监督管理局生物制品质量研究与评价重点实验室,国家卫生健康委生物技术产品检定方法及其标准化重点实验室,北京102629

出　　处：《药学学报》2024年第12期3367-3373,共7页Acta Pharmaceutica Sinica

基　　金：国家药典委员会药品标准制修订研究课题(2023S002);国家重点研发计划(2021YFF0600804);中国食品药品检定研究院化药所重点实验室学科建设基金(2024HYZX24)

摘　　要：按照各监管机构的要求,聚乙二醇(PEG)平均修饰率是PEG化蛋白类药物表征与质量控制中的关键属性。本研究基于“PEG化蛋白的示差信号及紫外信号等于该偶联蛋白中PEG和蛋白部分各自处于未偶联状态单独存在时所产生相应信号之和”的基本假设,通过进样20μL的1 mg·mL^(-1)人生长激素(hGH)对照品、2 mg·mL^(-1) PEG对照品和1 mg·mL^(-1) PEG化hGH原液样品,经分子排阻色谱柱分离,测定各溶液在紫外和示差检测器下产生的信号按照推导的公式计算,成功建立了基于分子排阻色谱紫外示差双检测器串联技术的PEG化hGH (怡培生长激素)平均修饰率测定方法,该法专属性良好,重复性良好(RSD=0.63%,n=9),与同时建立的经典酸水解法结果比较,回收率为100.0%,准确性良好。该法操作简单,可及性好,适用于本品PEG平均修饰率质量控制检测。According to the requirements of the regulatory authorities,degree of modification (DP) should be included in the characterisation of the PEGylated protein drug substance,and is one of the critical quality attributes for quality control.In this study,based on the fundamental assumption that the refractive index (RI) signal and the ultraviolet (UV) signal of PEGylated protein are equal to the sum of the corresponding signal produced by the polyethylene glycol (PEG) and protein parts of the conjugates in their uncoupled state,we developed a method to determine the DP of PEGylated recombinant human growth hormone (inpegsomatropin).In this method,20μL of1 mg·mL^(-1) human growth hormone (hGH) standard,2 mg·mL^(-1) PEG reference substance and 1 mg·mL^(-1) drug substance solution were each injected to size exclusion chromatographic (SEC) column for separation,detected with ultraviolet and refractive index (UV-RI) detectors in series.Finally,the DP was calculated as the formula derived from the fundamental assumption.The developed SEC-UV-RI method showed good specificity,repeatability (RSD=0.63%,n=9) and accuracy,with a recovery of 100.0%,compared with the result obtained from the simultaneously established classical acid hydrolysis method,demonstrating pretty handleability and accessibility,and is appropriate for quality control test of DP for this drug substance.

关 键 词：分子排阻色谱 紫外示差串联检测 聚乙二醇化 人生长激素 平均修饰率 

分 类 号：R917[医药卫生—药物分析学]