乙酰胆碱酯酶微反应器的制备及其中药抑制剂筛选方法的建立  

Study on preparation of immobilized acetylcholinesterase microreactor and its application in CMM inhibitors screening

作　　者：郭嘉亮[1,2] 沈焕圻 林航[2] 段倩倩 黄若诗 张婷婷 GUO Jia-liang;SHEN Huan-qi;LIN Hang;DUAN Qian-qian;HUANG Ruo-shi;ZHANG Ting-ting(School of Stomatology and Medicine,Foshan University,Foshan 528000,China;College of Pharmacy,Jinan University,Guangzhou 510632,China)

机构地区：[1]佛山科学技术学院医药工程学院,广东佛山528000 [2]暨南大学药学院,广东广州510632

出　　处：《中草药》2018年第22期5292-5297,共6页Chinese Traditional and Herbal Drugs

基　　金：国家自然科学基金资助项目(81503031;81872832);广东省科技计划项目(2016A020226004);广东省中医药建设专项资金项目(20161068);佛山科学技术学院博士启动项目(gg040952);大学生创新训练项目(XJ2018189)

摘　　要：目的制备固定化乙酰胆碱酯酶微反应器,并建立可用于中药乙酰胆碱酯酶抑制剂的快速筛选方法。方法以戊二醛为交联剂,氨基磁性微球为载体,制备一种新型的固定化乙酰胆碱酯酶微反应器,并对其理化性质和色谱性能进行了表征;应用该酶反应器对千层塔Huperziaserratum各提取部位进行酶抑制剂的筛选。结果酶反应体系的最佳底物浓度为50μmol/L,孵育时间5 min;红外表征、专属性验证、酶动力学考察以及稳定性考察均说明了该酶反应器的有效性;从千层塔提取物中筛选获得了石杉碱甲等乙酰胆碱酯酶抑制剂。结论建立了一种有效的乙酰胆碱酯酶抑制剂高通量筛选方法,可进一步推广与应用。Objective To prepare an immobilized acetylcholinesterase(AChE) microreactor and establish a rapid screening method for Chinese materia medica(CMM) acetylcholinesterase inhibitors(AChEIs). Methods A novel immobilized AchE microreactor was prepared by crosslinking with glutaraldehyde, using aminated magnetic microspheres as carrier. The characterizations were conducted by physicochemical properties and chromatographic performance. The immobilized AChE reactor was used to screen ACh EIs from the Huperzia Serratum extracts. Results In the enzyme reaction system, the optimum substrate concentration was 50 μmol/L, and the incubation time was 5 min, respectively. IR characterization, specificity verification, enzyme kinetics, and stability study results all demonstrated the effectiveness of the enzyme reactor. The CMM AChEI, huperzine A, was obtained from the screening of H. Serratum extracts. Conclusion A high throughput screening method for AChEIs is established in this paper, which will be further applied and popularized.

关 键 词：乙酰胆碱酯酶抑制剂 千层塔 微反应器 磁性微球 氨基磁性微球 石杉碱甲 

分 类 号：R283.6[医药卫生—中药学]