甘精胰岛素注射液有关物质分析方法研究  

Study on analytical methods of related substances in insulin glargine injection

作　　者：丁晓丽[1] 陈莹 胡馨月 李晶[1] 张慧[1] 梁成罡[1] DING Xiao-li;CHEN Ying;HU Xin-yue;LI Jing;ZHANG Hui;LIANG Cheng-gang(Division of Hormone,National Institutes for Food and Drug Control,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国家卫生健康委员会生物技术产品检定方法及其标准化重点实验室,北102629

出　　处：《药物分析杂志》2022年第1期23-32,共10页Chinese Journal of Pharmaceutical Analysis

基　　金：国家科技重大专项课题资助项目(2018ZX09101001-003-004)。

摘　　要：目的:优化甘精胰岛素注射液有关物质分析方法,论证现行标准规定限度科学性,并对主要有关物质进行鉴别。方法:采用Thermo scientific Bio-Basic-C_(18)色谱柱(250 mm×4.6 mm,5μm),以氯化钠-磷酸盐缓冲液-乙腈(18.4 g∶250 mL∶250 mL,加水至1000 mL)为流动相A,氯化钠-磷酸盐缓冲液-乙腈(3.2 g∶250 mL∶650 mL,加水至1000 mL)为流动相B,梯度洗脱,流速1.0 mL·min^(-1),检测波长214 nm,柱温35℃。利用该方法收集含量最大有关物质组分并利用LC-MS方法进行鉴别。分析色谱柱为ACQUITY UPLC BEH C_(18)Column(100 mm×2.1 mm,1.7μm,300A),以0.1%甲酸水溶液为流动相A,0.1%甲酸乙腈溶液为流动相B,进行梯度洗脱,柱温为50℃。质谱数据采集条件为MS;模式,一级质谱能量40 V。结果:该RP-HPLC有关物质分析方法,方法学验证结果良好,能够有效分离并准确测定样品中的有关物质;经鉴定,甘精胰岛素注射液的主要有关物质为B链第3位天冬酰胺的脱-NH;降解产物,即3^(B)-琥珀酰亚胺-甘精胰岛素。结论:优化后的RP-HPLC方法能够有效用于甘精胰岛素注射液有关物质分析,现行国家标准有关物质限度规定科学合理,能够满足甘精胰岛素注射液货架期内的质控需求。Objective:To optimize the analysis method of related substances of insulin glargine injection,demonstrate the scientific limits for the current standards and identify the main related substances.Methods:The separation was performed on Thermo scientific Bio-Basic-C_(18)column(250 mm×4.6 mm,5μm).Sodiumchloride,phosphate buffer and acetonitrile were mixed in a 18.4 g∶250 mL∶250 mL ratio,and added with water to 1000 mL to be used as mobile phase A;sodium chloride,phosphate buffer,acetonitrile were mixed in a 3.2 g∶250 mL∶650 mL ratio,and added with water to 1000 mL to be used as mobile phase B;gradient elution program was used at the flow rate of 1.0 mL·min^(-1).The detection wavelength was 214 nm and the column temperature was 35℃.The maximum content of related substances was collected and identified by LC-MS.The chromatographic separation was carried out on a column ACQUITY UPLC BEH C_(18)column(100 mm×2.1 mm,1.7μm,300A)with the mobile phase consisting of 0.1%formic acid aqueous solution(A)-0.1%formic acid acetonitrile solution(B)in a gradient mode.The column temperature was set to 50℃.The mass spectra was acquired with MS;mode.The primary mass spectrum energy was 40 V.Results:The RP-HPLC method for the analysis of related substances has good methodological verification results,and could effectively separate and accurately determine the related substances in the samples.It was identified that the main related substance of insulin glargine injection was the 3^(B)-succinimide-insulin glargine,which was deaminated degradation product of asparagine at the third position of chain B.Conclusion:The optimized RP-HPLC method could be effectively used for the analysis of related substances in insulin glargine injection.The current national standards for related substances limits are scientific and reasonable,which could meet the quality control of insulin glargine injection during the shelf life.

关 键 词：甘精胰岛素注射液 药品在用期 有关物质 3^(B)-琥珀酰亚胺-甘精胰岛素 结构鉴定 反相高效液相色谱法 液相质谱联用 

分 类 号：R917[医药卫生—药物分析学]