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期刊名称：药物分析杂志
主管单位：中国科学技术协会
主办单位：中国药学会承办：中国食品药品检定研究院
主编：金少鸿
地址：北京天坛西里2号
邮政编码：100050
电话：010-67012819,67058427 电子邮箱：ywfx@nifdc.org.cn
国际标准刊号：ISSN 0254-1793
国内统一刊号：CN 11-2224/R
邮发代号：2-237
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蛇毒血凝酶纯度分析方法研究
Study on the purity analysis method of hemocoagulase
单位(英文)：1. Institutes of Biochemical and Biological Drugs, School of Pharmaceutical Sciences, Shandong University, Jinan 250012, China;
2. National Institutes for Food and Drug Control, Beijing 100050, China;
3. Qiqihaer Institute for Drug Control, Qiqihaer 161006, China;
4. Shenyang Pharmaceutical University, Shenyang 110016, China
分类号：
出版年·卷·期（页码）：2012,32 (8):0-0
DOI:
10.16155/j.0254-1793.2017.01.01
-----摘要：-------------------------------------------------------------------------------------------
目的: 建立蛇毒血凝酶纯度测定方法。 方法: 分别采用SDS-聚丙烯酰胺凝胶电泳法(SDS-PAGE)和等电聚焦法(IEF),对凝胶考染并扫描;采用体积排阻高效液相色谱法(SEC-HPLC),使用TSK G3000SW(7.8 mm×300 mm)凝胶柱,流动相为55 mmol·L-1 柠檬酸钠-0.01%Tween 80溶液;流速0.5 mL·min-1,检测波长280 nm;采用反相高效液相色谱法(RP-HPLC),使用C8 Vydac(4.6 mm×250 mm,5 μm)色谱柱,流动相为0.1%三氟乙酸溶液(A)-含0.1%三氟乙酸的90%乙腈(B),梯度洗脱,流速0.8 mL·min-1,;柱温为45℃,检测波长:280 nm。 结果: 4种方法测得蛇毒血凝酶的纯度分别为89.3%,88.5%,95.4%,87.4%。 结论: 对于蛇毒血凝酶纯度测定,非还原SDS-PAGE、IEF及RP-HPLC法均可准确地反映其样品纯度。
-----英文摘要：---------------------------------------------------------------------------------------
Objective: To establish a method to determine the purity of Haemocoagulase(HC). Methods: SDS-polyacrylamide gel electrophoresis (SDS-PAGE),isoelectric focusing (IEF),size exclusion column-HPLC (SEC-HPLC)and reversed phase-HPLC (RP-HPLC) were used respectively to determine the purity of HCA.TSK G3000SW (7.8 mm×300 mm) column was applied for SEC-HPLC,using 55 mmol·L-1 citric acid sodium-0.01% Tween 80 solution as the mobile phase the flow rate of 0.5 mL·min-1,the detection wavelength at 280 nm;C8 Vydac (4.6 mm×250 mm,5 μm) column was employed for RP-HPLC,using 0.1% triflouroacetic acid (TFA) as the mobile phase A,and 90% acetonitrile containing 0.1% TFA as the mobile phrase B,at the flow rate of 0.8 mL·min-1,the detection wavelength 280 nm. Results: The results of purity detected by the four methods were 89.3%,88.5%,95.4% and 87.4%,respectively. Conclusion: Except SEC-HPLC,the other three methods were feasible to determine the purity of HCA.
-----参考文献：---------------------------------------------------------------------------------------
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2 Tang SS,Zhang JH,Tang BS, et al.Biochemical and hemostatic mechanism of a novel thrombin-like enzyme.Thromb Res,2009,124(5):631
3 ZHOU Su-fang(周素芳),ZHONG Man-sen(钟满森),YU Qing-sheng(余清声),et al.A study of the enhancing blood coagulation effect of thrombin-like enzyme from Deinagkistrodon acutus venom(尖吻蝮蛇毒类凝血酶促凝作用的研究).J Snake(蛇志),1997,9(4):l
4 ZHENG Ying(郑颖),SHEN Ju-ren(沈居仁),FAN Quan-shui(范泉水).Hemocoagulase comparison(蛇毒血凝酶的比较).J Snake(蛇志),2008,20(3):218
5 FU Hong-yi(付宏义),ZHOU Lei(周磊).Rencent advances of research on hemocoagulase(蛇毒类凝血酶的研究进展).Chin Pharm J(中国药学杂志),2008,43(4):245
6 WANG Jun-zhi(王军志).Research and Development and Quality Control of Biotech Drugs(生物技术药物研究开发和质量控制),Beijing(北京):Science Press(科学出版社),2007.79
7 ChP(中国药典).2010.Vol Ⅱ(二部):Appendix(附录)54
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