论文部分内容阅读
目的从广西圆斑蝰蛇毒中分离纯化凝血因子X激活剂(factor X activator of Russell’sviper venom,RVV-X)并研究其止血作用。方法应用阴离子交换色谱和凝胶过滤从广西圆斑蝰蛇毒中分离纯化RVV-X,以复钙化时间进行活性鉴定,以SDS-PAGE测定纯度和分子量。观察RVV-X对家兔肝脏创伤出血模型出血时间及出血量,家兔和肝素化小鼠凝血时间的影响。结果广西圆斑蝰粗毒经阴离子交换色谱获得六个蛋白峰,将RVV-X所在第V峰用凝胶过滤获得的组分A为RVV-X纯品,其凝血活性具有钙依赖性。SDS-PAGE非还原性条件下呈单一蛋白条带,分子量约为97.8kDa,还原性条件下呈二蛋白条带,为一条重链(约76.3kDa)和一条轻链(约21.5kDa)。RVV-X可显著减少家兔肝脏切口的失血量,缩短切口出血时间;能显著缩短家兔及肝素化小鼠凝血时间。结论用层析方法从广西圆斑蝰蛇毒中分离得到的凝血因子X激活剂具有较强的止血活性。
OBJECTIVE: To isolate and purify factor X activator of Russell’s ventricular ventricular vasculature (RVV-X) from Guangxi spotted viper venom and study its hemostatic effect. Methods RVV-X was isolated and purified from Guangxi spotted venom by anion exchange chromatography and gel filtration. The activity of RVV-X was determined by time of re-calcification. The purity and molecular weight were determined by SDS-PAGE. To observe the effect of RVV-X on the bleeding time and blood loss of rabbit liver trauma hemorrhage model and the clotting time of rabbits and heparinized mice. Results Six crude protein spots were detected by anion exchange chromatography. The fraction A of RVV-X was gel-filtered to obtain RVV-X purified product. The coagulation activity was calcium-dependent. SDS-PAGE non-reducing conditions, a single protein band, molecular weight of about 97.8kDa, two protein bands under reducing conditions, a heavy chain (about 76.3kDa) and a light chain (about 21.5kDa). RVV-X can significantly reduce the amount of blood loss in rabbit liver incision, shorten the incision bleeding time; can significantly shorten the clotting time in rabbits and heparinized mice. Conclusion The chromatographic method was used to isolate the coagulation factor X activator from Guangxi spotted snake venom and has strong hemostatic activity.