UHPLC-MS分析人促红素产品中CHO宿主细胞蛋白残留

Analysis of residual CHO host cell protein in human erythropoietin products by UHPLC-MS

目的:利用超高效液相色谱-质谱联用技术(UHPLC-MS)分析人促红素(human erythropoietin,hEPO)产品中CHO宿主细胞蛋白(host cell protein,HCP)残留。方法:将hEPO原液和蛋白标准品一起进行变性、还原烷基化、酶切处理;采用Accucore Vanquish C18色谱柱串联超高效液相色谱和Q ExactiveTM Plus组合型四极杆OrbitrapTM质谱仪对样品进行分离和检测;使用BioPharma Finder 4.1软件中的HCP工作流程进行定性及定量分析。结果:10种HCP在4批hEPO原液样品中均被检测到,且不同样品中HCP总量有较大差异,从25×10^(-6)至约1000×10^(-6);含量>50×10^(-6)的HCP,CV值均<10%。结论:UHPLC-MS法可对CHO细胞表达的hEPO产品中HCP进行定性定量分析,不同来源hEPO样品中HCP种类和含量均有差异。

Objective:To analyze residual CHO host cell protein(HCP)in human erythropoietin(hEPO)prod⁃ucts by UHPLC-MS.Methods:The hEPO bulks and internal protein standard were denatured,reduced,alky⁃lated and digested;Accucore Vanquish C18 column connected with UHPLC and Q Exactive Plus Hybrid Quadru⁃pole-Orbitrap Mass Spectrometer were employed to separate and detect the samples;qualitation and quantitation of HCP were performed using HCP workflow in BioPharma Finder 4.1 software.Results:Ten HCPs were detected in all 4 batches of hEPO bulks,and the HCP amounts varied between different samples,ranging from 25×10^(-6)to approximately 1000×10^(-6);the CV%were below 10%for the HCPs above 50×10^(-6).Conclusions:The meth⁃od of UHPLC-MS could be used for qualitative and quantitative analysis of HCP in hEPO products expressed in CHO system,and the HCP types and amounts in different samples varied considerably.