C1酯酶抑制剂分离纯化工艺研究  

Study on separation and purification process of C1 esterase inhibitor

作　　者：徐佳彬 李雅文 龚佳莉 罗二华 张猛 XU Jiabin;LI Yawen;GONG Jiali;LUO Erhua;ZHANG Meng(Boya Bio-pharmaceutical Group Co.Ltd.Fuzhou 344000,China)

机构地区：[1]华润博雅生物制药集团股份有限公司,江西抚州344000

出　　处：《中国输血杂志》2022年第11期1121-1124,共4页Chinese Journal of Blood Transfusion

摘　　要：目的 研究以人凝血酶原复合物(PCC)制备过程废弃洗涤液为原料分离纯化C1酯酶抑制剂(C1-esterase inhibitor, C1-INH)的工艺。方法 通过聚乙二醇(Polyethylene glycol, PEG)4000沉淀及阳离子层析两步法分离纯化C1-INH。调整原料pH及PEG4000浓度考察PEG4000沉淀法最佳条件;PEG沉淀后离心,上清液处理后作为阳离子交换层析的上样液,使用Fractogel EMD SE HiCap(M)凝胶及CM Sepharose FF凝胶进行离子交换层析,通过比较C1-INH活性收率及比活性选择最适合的凝胶及分离条件。结果 在pH6.1的条件下,PEG4000质量分数14%时,搅拌10min后离心,C1酯酶抑制剂回收率≈70%,铜蓝蛋白(ceruloplasmin, CER)去除率>95%;使用Fractogel EMD SE HiCap(M)凝胶进行阳离子交换层析,洗脱液盐浓度为0.25M氯化钠时,C1酯酶抑制剂活性收率>80%,比活性>5IU/mg。结论 以PCC制备过程中废弃洗涤液为原料,经过PEG沉淀和Fractogel EMD SE HiCap(M)离子交换层析纯化制备能够得到高比活性的C1酯酶抑制剂。Objective To study the technology of separating and purifying C1 esterase inhibitor(C1-INH) by using the waste washing liquid as raw materia during the preparation of human prothrombin complex(PCC) l. Methods C1-INH was isolated and purified by a two-step method of polyethylene glycol(PEG) 4000 precipitation and cation chromatography. The pH of raw materials and the concentration of PEG4000 were adjusted to investigate the optimal conditions of PEG4000 precipitation method. After PEG was precipitated and centrifuged, the supernatant is treated as the loading solution for cation exchange chromatography, using Fractogel EMD SE HiCap(M) gel and CM Sepharose FF gel for ion exchange chromatography. The most suitable gel and separation conditions were selected by comparing the C1-INH antigen yield, activity yield and specific activity. Results Under the condition of pH 6.1, when the mass fraction of PEG4000 was 14%, the recovery rate of C1 esterase inhibitor was close to 70%, and the removal rate of ceruloplasmin was more than 95% after stirring for 10 minutes. As fractogel EMD SE HiCap(M) gel was used for cation exchange chromatography, when the eluent salt concentration was 0.25 M sodium chloride, the activity yield of C1 esterase inhibitor was greater than 80%, and the specific activity was greater than 5 IU/mg. Conclusions Using the waste washing liquid as the raw material during the preparation of PCC, the C1 esterase inhibitor with high specific activity can be prepared through PEG precipitation and purification by Fractogel EMD SE HiCap(M) ion exchange chromatography.

关 键 词：C1酯酶抑制剂 遗传性血管性水肿(HAE) 聚乙二醇4000 Fractogel EMD SE HiCap(M)凝胶 人凝血酶原复合物(PCC) 

分 类 号：R457.1[医药卫生—治疗学] R554[医药卫生—临床医学]