Ⅳ型胶原酶模型的建立及其在中药活性成分筛选中的应用  

Establishment of Type Ⅳ Collagenase Model and Its Application in TCM

作　　者：薛南南 曹寅秋 房康 陈萌 张欣钰 王辉[1] 吴高荣 贾晓慧[1] 郭文博[1] 王鹏龙[1] 雷海民[1] XUE Nan-nan;CAO Yin-qiu;FANG Kang;CHEN Meng;ZHANG Xin-yu;WANG Hui;WU Gao-rong;JIA Xiao- hui;GUO Wen-bo;WANG Peng-long;LEI Hai-min(School of Chinese Pharmacy,Beijing University of Chinese Medicine,Beijing 102488,China)

机构地区：[1]北京中医药大学中药学院,北京102488

出　　处：《现代中药研究与实践》2019年第1期20-23,共4页Research and Practice on Chinese Medicines

基　　金：国家自然科学基金项目(81603256);北京中医药大学创新团队资助项目(2011-CXTD-15)

摘　　要：目的建立Ⅳ型胶原酶筛选模型,并运用此模型对中药活性成分进行评价。方法利用Ⅳ型胶原酶特异性水解明胶释放的末端氨基与2,4,6-三硝基苯磺酸(TNBS)发生显色,吸光度值可以衡量Ⅳ型胶原酶含量,优化Ⅳ型胶原酶浓度、琥珀酰凝胶浓度、显色剂用量及时间参数。结果确定了Ⅳ型胶原酶筛选模型的条件:5.0×10^(-5) g/L酶溶液、20 g/L琥珀酰凝胶溶液、60μl 0.05%TNBS、显色30 min,阳性药盐酸四环素在1.0 g/L下对该酶的抑制率是19.39%,并呈现一定的量效关系,表明成功建立该筛选模型。应用该模型筛选发现,阿魏酸和川芎嗪具有明显的抑制Ⅳ型胶原酶活性,而水杨酸和小檗碱则具有促进作用。结论Ⅳ型胶原酶的筛选模型成功建立并应用到中药活性成分筛选中,为快速评价中药活性成分及其作用机制提供了有益参考。Objective To establish a type Ⅳ collagenase screening model and use it to evaluate the active ingredients of TCM. Methods As type Ⅳ collagenase hydrolyze gelatin specifically, the released terminal amino group react with 2,4,6-trinitrobenzene sulfonic acid(TNBS) with color;the absorbance can be measured by UV for type Ⅳ collagenase content. The screening model is established by optimizing the concentration of type Ⅳ collagenase, succinyl gel concentration, the amount of color reagent and time. Results The conditions of the type Ⅳ collagenase screening model are finally determined: 0.05 mg/L type Ⅳ collagenase solution, 20 g/L succinyl gel solution, 0.05 % 60 μl TNBS, coloration for 30 min, the inhibitory rate of positive drug tetracycline hydrochloride at 1.0 g/L is 19.39 %, and showed a certain dose-effect relationship, indicating that the screening model of type Ⅳ collagenase is established successfully. Using the model, we find that ferulic acid and ligustrazine show inhibitory activity, while salicylic acid and berberine promote inversely. Conclusion The screening model for type Ⅳ collagenase is successfully established and applied to screen the active ingredients of TCM, which provide a useful reference for rapid evaluation of active ingredients and the mechanism of action.

关 键 词：Ⅳ型胶原酶 筛选 模型 中药活性成分 

分 类 号：R285[医药卫生—中药学]