影响胃蛋白酶和胰蛋白酶纤溶活性因素初探及其纤溶活性失活研究  

Effect of factors on pepsin and trypsin fibrinolytic activity and deactivation of fibrinolytic activity

作　　者：王正俊[1,2] 李博[1,2,3,4] 郭立玮[1,2] 吴勉华[3,4] 朱华旭[1,2] 

机构地区：[1]南京中医药大学江苏省中药资源产业化过程协同创新中心,江苏南京210023 [2]南京中医药大学中药复方分离工程重点实验室,江苏南京210029 [3]南京中医药大学第一临床医学院,江苏南京210046 [4]南京中医药大学江苏省中医药防治肿瘤协同创新中心,江苏南京210023

出　　处：《中草药》2016年第1期46-56,共11页Chinese Traditional and Herbal Drugs

基　　金：国家自然科学基金资助项目(81274096;81303230);江苏省科技厅产学研联合创新资金-前瞻性研究项目(BY2012036);江苏省科技支撑计划-社会发展项目(BE-2012763);中国博士后科学基金(2013M541704);江苏省博士后科学基金(1301133C)

摘　　要：目的通过探讨影响胃蛋白酶和胰蛋白酶纤溶活性的因素及其纤溶活性失活方法,排除胃蛋白酶和胰蛋白酶对中药鼠妇Armadillidium vulgare活性粗蛋白纤溶活性检测的干扰,以在鼠妇活性粗蛋白酶解过程,获取相对分子质量更小、效价更高的蛋白质或多肽。方法采用纤溶蛋白平板法分别研究pH值、温度、金属离子、酶抑制剂和表面活性剂对胃蛋白酶和胰蛋白酶纤溶活性的影响,以获取较佳的酶失活工艺,并研究较佳酶失活工艺对尿激酶、蚓激酶、鼠妇纤溶酶纤溶活性的影响。结果 pH值、温度、金属离子、酶抑制剂和表面活性剂均可对胃蛋白酶和胰蛋白酶纤溶活性产生影响,其中胃蛋白酶的最优失活方案为pH 6.0~8.0;胰蛋白酶的最优失活方案为质量浓度25 mg/mL TLCK和浓度1 mmol/L EDTA的混合制剂。结论研究所获取的较佳酶失活工艺,可用于以纤维蛋白平板法检测胃蛋白酶、胰蛋白酶降解产物的纤溶活性,操作简单,重复性好,稳定性好。Objective To investigate the method for study on the effect of factors on pepsin and trypsin fibrinolytic activity and deactivation of fibrinolytic activity and to eliminate the interference of pepsin and trypsin on the detection of crude protein fibrinolytic activity of Armadillidium vulgare（porcellio plasmin） in order to obtain the proteins or peptides which have the smaller molecular weight but higher titer during the pepsin and trypsin degradation. Methods To study the effect of pepsin and trypsin deactivation on pH value, temperature, metal ions, enzyme inhibitor, surfactant, and responsing fibrinolytic by fiber fibrin plate assay. The better enzyme deactivation process was obtained and used for studying the effect on the fibrinolytic activity of urokinase, lumbrokinase, and porcellio plasmin. Results All the pH value, temperature, metal ions, enzyme inhibitor, and surfactant have had an impact on pepsin and trypsin fibrinolytic activity. Among them the optimum deactivation of pepsin was pH 6.0—8.0, while the optimum deactivation of trypsin was mixed preparation with TLCK at the concentration of 25 mg/mL and EDTA at the concentration of 1 mmol/L. Conclusion This study has obtained the better enzyme deactivation process which could be used for the detection of fibrinolytic activity of pepsin and trypsin degradation product by fiber fibrin plate assay, the operation is simple, and the repeatability and stability are good.

关 键 词：胃蛋白酶 胰蛋白酶 纤溶活性 纤维蛋白平板法 酶失活 尿激酶 蚓激酶 鼠妇纤溶酶 

分 类 号：R284.2[医药卫生—中药学]