新型体外胆碱酯酶抑制剂筛选体系的构建  

Establishment of a Screening System for Cholinesterase Inhibitors

作　　者：李利民[1] 胡胜全 马玉翠 金宇[1] 陈可冀[3] 吴正治[1,2] LI Li-min;HU Sheng-quan;MA Yu-cui;JIN Yu;CHEN Ke-ji;WU Zheng-zhi(The First Affiliated Hospital of Shenzhen University,Guangdong Shenzhen 518035;Shenzhen Institute of Gerontology,Guangdong Shenzhen,518035;Xiyuan Hospital of China Academy of Chinese Medical Sciences,Beijing 100091)

机构地区：[1]深圳大学第一附属医院,广东深圳518035 [2]深圳市老年医学研究所,广东深圳518035 [3]中国中医科学院西苑医院,北京100091

出　　处：《深圳中西医结合杂志》2021年第12期1-5,共5页Shenzhen Journal of Integrated Traditional Chinese and Western Medicine

基　　金：国家自然科学基金项目资助课题(81574038);广东省基础与应用基础研究基金资助课题(2020A1515011427);深圳市基础研究重点(学科布局)项目资助课题(JCYJ20170413161352000);深圳市医疗卫生三名工程项目资助课题(SZSM201612049)。

摘　　要：目的:构建新型简便可靠的体外胆碱酯酶抑制剂筛选体系。方法:从SD大鼠大脑和血清中提取乙酰胆碱酯酶(AChE)和丁酰胆碱酯酶(BuChE),应用Ellman比色法构建体外AChE和BuChE抑制剂筛选实验体系。结果:选择10倍稀释的雄性大鼠脑匀浆液〔蛋白浓度(1.69±0.10)mg·mL^(-1)〕和5倍稀释的血清作为AChE/BuChE酶源,使用AChE/BuChE抑制剂他克林作为参考。他克林对AChE/BuChE的半数抑制浓度(IC50)分别为262 nmol·L^(-1)和168 nmol·L^(-1),体外胆碱酯酶抑制剂筛选体系构建成功。在等量蛋白浓度情况下,雌性大鼠脑匀浆液中AChE酶活性为雄性大鼠的1.84倍。结论:本研究从大鼠大脑和血清里分别提取AChE和BuChE粗酶,构建了一种新型的简单易行且稳定可靠的AChE/BuChE抑制剂筛选平台。Objective To establish a screening system for cholinesterase inhibitors.Methods Acetylcholinesterase(AChE)and butyrylcholinesterase(BuChE)were obtained from the brain and serum of SD rats.Ellman colorimetry assay was used to establish an in vitro experimental system for the screening of AChE and BuChE inhibitors.Results The 10–fold dilution of male rat brain homogenate(protein concentration,1.69±0.10 mg/mL)and 5–fold dilution of serum were selected as AChE/BUCHE enzyme sources,and the AChE/BuChE inhibitor,tacrine,was used as the reference.It was found that tacrine inhibited AChE and BuChE with an IC50 of 262 nM and 168 nM,respectively,an observation consistent with the literature reports,suggesting that the screening system for cholinesterase inhibitors in vitro has been successfully established.In addition,the activity of AChE enzyme activity in the brain homogenate of female rats was 1.84 times than that of male rats under the same amount of protein.Conclusions AChE and BuChE crude enzymes were extracted from rat brain and serum respectively to construct a sensitive and stable AChE/BuChE inhibitor screening platform.

关 键 词：乙酰胆碱酯酶 丁酰胆碱酯酶 胆碱酯酶抑制剂 他克林 动物实验 

分 类 号：R965.1[医药卫生—药理学] R971.91[医药卫生—药学]