乌苏里蝮蛇血凝酶的分离纯化和质量研究  

Purification and quality of Wusuli haemocoagulase

作　　者：张雪玲[1] 崔艳[2] 崔爱湜 王超众 宋运娜[5] 贾首时 于晓秋[1] 贾翠英[1] ZHANG Xueling CUI Yan CUI Aishi WANG Chaozhong SONG Yunna JIA Shoushi YU Xiaoqiu JIA Cuiying(Department of Pharmacy, the Second Affiliated Hospital of Qiqihar Medical University, Heilongjiang Provlnce, Qiqihar 161006, China Department of Pharmacy, the First Hospital of Harbin City, Heilongjiang Province, Harbin 100010, China Department of Pharmacy, the Third Affiliated Hospital of Qiqihar Medical University, Heilongjiang Province, Qiqihar 161006, China Qiqihar Institute for Food and Drug Control, Heilongjiang Province, Qiqihar 161005, China Basic College of Qiqihar Medical University, Heilongjiang Province, Qiqihar 161005, China)

机构地区：[1]齐齐哈尔医学院附属第二医院药剂科,黑龙江齐齐哈尔161006 [2]哈尔滨市第一医院药剂科,黑龙江哈尔滨100010 [3]齐齐哈尔医学院附属第三医院药剂科,黑龙江齐齐哈尔161005 [4]齐齐哈尔市食品药品检验检测中心,黑龙江齐齐哈尔161005 [5]齐齐哈尔医学院基础学院,黑龙江齐齐哈尔161005

出　　处：《中国医药导报》2017年第1期20-23,47,共5页China Medical Herald

基　　金：黑龙江省齐齐哈尔市科学技术计划项目(SFGG-201538)

摘　　要：目的建立一种分离纯化乌苏里蝮蛇血凝酶(WH)的方法并为其制定质量标准。方法先后采用分子筛层析法、离子交换色谱法、亲和层析法从乌苏里蝮蛇毒中分离纯化得到一种具有凝血活性的酶成分,并采用SDS-聚丙烯酰胺凝胶电泳法、RP-HPLC法测定其纯度,HPSEC法测定其分子量,IEF法测定其等电点,Lowry法测定其蛋白浓度,并用标准人血浆法测定该酶的比活力。结果从乌苏里蝮蛇蛇毒中分离纯化了一种凝血酶成分,SDS-Page显示为一条带、测得分子量约为34 k D,RP-HPLC得到单一的色谱峰,HPSEC法测得该酶分子量为34.7 k D,等电点为5.25,此酶具有体外凝血活性,比活力为5.40μg/U。结论该方法可用于WH的分离纯化,制定的质量标准可用于控制该血凝酶的质量。Objective To establish a method of purification and quality standards for Wusuli haemocoagulase. Methods A component of enzyme with the clotting activity was isolated from Wusuli viper by gel filtration and anion-exchange chromatography and heparin-Sepharose affinity chromatography. SDS-Page and RP-HPLC were used to determine its purity. HPSEC was used to determine the molecular weight. IEF method was used to measure the isoelectric point while its protein concentration was determined by the Lowry method. Standard human plasma method was used to determine the specific activity of the enzyme. Results A kind of thrombin was purified from Wusuli viper. One band was displaied on SDS-Page and the molecular weight was about 34 k D. RP-HPLC got one single chromatographic peak. The molecular weight of this enzyme was measured by HPSEC method was 34.7 k D and the isoelectric point was 5.25. This enzyme possesses the characteristion of extractor clotting activity and its specific activity was 5.40 μg/U. Conclusion This method can be used for Wusuli haemocoagulase separation and purification. And this quality standards can be used to control the quality of this haemocoagulase.

关 键 词：乌苏里蝮蛇 血凝酶 分离纯化 质量标准 

分 类 号：R282.740.2[医药卫生—中药学]