液相色谱分离纯化五步蛇毒凝血酶样酶  

Purification of Thrombin-like Enzyme from Agkistrodon Acutus Venom by Liquid Chromatographic

作　　者：苏煌财[1] 许云禄[2] 林玉仙[1] 

机构地区：[1]福州市第一医院药学部,福建福州350009 [2]福建医科大学蛇毒研究所,福建福州350000

出　　处：《药品评价》2014年第24期29-31,共3页Drug Evaluation

基　　金："五步蛇毒凝血酶样酶分离;纯化及生物活性研究"。福州市科技计划项目;编号:2013-S-122-6

摘　　要：目的:改进分离工艺,找到液相色谱分离纯化五步蛇毒中凝血酶样酶的最佳方案。方法:五步蛇毒依次经Superdex G75凝胶过滤、DEAE-Sephrose F.F阴离子交换和Superdex G30凝胶过滤,通过改变洗脱液pH值、流速、Tris浓度、盐浓度及上样量来考察对分离纯化五步蛇毒中凝血酶样酶的影响。结果:Superdex 75凝胶过滤色谱分离纯化受洗脱液流速、Tris浓度、盐浓度、pH值、上样量的影响;DEAE-Sephrose F.F阴离子交换色谱受盐浓度的影响;Superdex G30凝胶过滤色谱受上样量的影响。三步分离纯化后获得一种五步蛇毒凝血酶样酶,分子量约23kD。结论:依次经Superdex G75、DEAE-Sephrose F.F和Superdex G30凝胶,通过改变洗脱液流速、Tris浓度、盐浓度、pH值、上样量等能得到最佳分离条件,最终可获得五步蛇毒凝血酶样酶的纯品。Objctive： This study was designed to separate and purify the component of thrombin-like enzyme from Agkistrodon acutus venom by the new technology. Methods： Proteins were separated and purified by Superdex 75, DEAE-Sepharose Fast Flow and Superdex 30, with the changing of the flow rate, pH value, concentrations of riffs and NaC1, and the sample volume. Regults： The flow rate, pH value, concentrations of riffs and NaC1, as well as the sample volume affected Superdex 75＇ s separation effect. The concentration of NaC1 influences DEAE-Sepharose Fast Flow＇ s separation. The sample volume influences Superdex 30＇ s separation effect. The thrombin-like enzyme protein was purified from Agkistrodon acutus venom with a molecular weight around 25 kD after the three-step separation. Conelusioa： By changing the eluent flow rate, the concentrations of Tris and NaCI, pH value, volume of sample, the best separation conditions and purified thrombin-like enzyme from Agkistrodon acutus venom can be obtained.

关 键 词：五步蛇毒 凝血酶样酶 凝胶过滤色谱 阴离子交换色谱 

分 类 号：O657.7[理学—分析化学] O657.72[理学—化学]