重组人促卵泡激素N末端不均一性分析方法研究

目的：建立重组人促卵泡激素（rhFSH）N端不均一性的分析方法，并对不同企业样品rhFSH原液的N端不均一性进行比对研究。方法：通过对亚基的还原与烷基化处理方法、糖苷酶切条件及RP-HPLC条件的优化，将完整与N端缺失氨基酸的亚基有效分离，建立了2种rhFSH的N端不均一性分析方法，并利用质谱分别对目的蛋白进行定性鉴别。以国内外企业的rhFSH原液为样品进行N末端不均一性比较。结果：建立的2种RP-HPLC方法均可实现完整与缺失β亚基的有效分离，各样品rhFSH的N端不均一性基本一致。结论：rhFSH国内生物类似药与国外参照药的N端不均一性比较有较高的相似性；本实验建立的2种RP-HPLC方法可评价产品质量，为生物类似药的可比性研究提供技术支持。

Objective: To establish a method for the analysis of N-terminal heterogeneity of recombinant human follicle stimulating hormone (rhFSH),and to compare the N terminal heterogeneity of rhFSH sample solutions from in different enterprises. Methods: Integrated subunit and subunit with N-terminal amino acid residues losing were separated by optimizing the reduction and alkylation process,the glycosidase digestion,and the RP-HPLC conditions. Two methods of analyzing N-terminal heterogeneity of rhFSH were established,and target proteins were further indentified by mass spectrometry. The N-terminal heterogeneity was compared with the domestic and foreign enterprises' rhFSH solutions. Results: Integrated β subunit and the loss β subunit could be separated by two RP-HPLC methods,and the N-terminal heterogeneity of rhFSH from different enterprises were basically consistant. Conclusion: The N-terminal heterogeneity of rhFSH in the domestic biosimilar druys was of high similarity than that in the international reference drugs;two established RP-HPLC methods can evaluate the quality of the products and provide technical support for the comparative study on biosimilar.

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