摘要
目的建立从黄酮化合物库中筛选胰蛋白酶抑制剂的新方法。方法利用毛细管固定化酶反应器技术,通过环氧基的开环反应,将胰蛋白酶与poly(GMA-co-PEGDA)有机聚合物整体材料共价键合制备毛细管固定化胰蛋白酶反应器;对酶反应浓度与反应时间等参数进行了系统研究,利用荧光显微镜、扫描电子显微镜和拉曼光谱对酶反应器进行了表征;采用微径液相色谱法对酶反应器的活性和动力学参数进行了系统评价;将制备的酶反应器用于20种中药黄酮化合物中胰蛋白酶抑制剂的筛选;通过分子对接手段进一步考察胰酶与抑制剂之间的作用机制。结果16种黄酮化合物具有不同程度的胰蛋白酶抑制作用,其中,漆黄素的抑制效果最佳,抑制率为(62.9±1.2)%,分子对接结果进一步说明了漆黄素与胰蛋白酶之间的作用机制。结论建立了一种简单、有效的胰蛋白酶抑制剂的筛选方法,可进一步推广应用。
A new method for screening trypsin inhibitors from flavonoids was successfully developed by immobilized capillary enzyme reactor(ICER).An organic polymer monolithic ICER was developed by covalently bonding trypsin to poly(GMA-co-PEGDA)(poly(glycidyl methacrylate-co-poly(ethylene glycol)diacrylate))monolith via ring opening reaction of epoxy groups.The parameters including enzyme reaction concentration and time were systematically studied.Fluorescence microscopy,scanning electron microscopy and raman spectroscopy were used to characterize monolithic trypsin-ICER.The activity and kinetic parameters of the monolithic trypsin-ICER were also systematically evaluated by micro-LC.The prepared ICER was applied to screen trypsin inhibitors from 20 flavonoids of TCM.Additionally,molecular docking was also applied for the study of the interactions between trypsin and the screened compound.Sixteen flavonoids showed inhibitory effect on trypsin.Among them,fisetin exhibited the highest inhibitory potency with the inhibition rate of(62.9±1.2)%,and the interaction between fisetin and trypsin was explained by molecular docking study.A high throughput screening method for trypsin inhibitors was established in this paper,which will be further applied and popularized.
作者
郭嘉亮
林航
范莹盈
沈焕圻
张丽华
曾煦欣
孙平华
GUO Jia-liang;LIN Hang;FAN Ying-ying;SHEN Huan-qi;ZHANG Li-hua;ZENG Xu-xin;SUN Ping-hua(Department of Pharmacy,Foshan University,Foshan 528000,China;Pharmacy College,Jinan University,Guangzhou 510632,China)
出处
《中国药物化学杂志》
CAS
CSCD
北大核心
2019年第6期440-447,共8页
Chinese Journal of Medicinal Chemistry
基金
国家自然科学基金项目(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(34016006)
佛山科学技术学院博士启动项目(gg040952).
