α-淀粉酶功能化MIL-53(Al)-NH_(2)对葛根提取物的配体垂钓研究  

Ligand Fishing Study on Puerariae Lobatae Radix Extract Based onα-Amylase Functionalized MIL-53(Al)-NH_(2)

作　　者：张勇 王静[2] 陈宇萌 冯苏宇 高桂花[1,2] ZHANG Yong;WANG Jing;CHEN Yumeng;FENG Suyu;GAO Guihua(School of Pharmacy,Jining Medical University,Rizhao 276826,China;School of Pharmacy,Shandong University of Traditional Chinese Medicine,Jinan 250355,China)

机构地区：[1]济宁医学院药学院,山东日照276826 [2]山东中医药大学药学院,济南250355

出　　处：《中国现代应用药学》2023年第6期730-735,共6页Chinese Journal of Modern Applied Pharmacy

基　　金：国家自然科学基金项目(82003706);日照市自然科学基金项目(RZ2021ZR18)。

摘　　要：目的采用配体垂钓策略筛选葛根95%乙醇提取物中的具有α-淀粉酶抑制活性的物质。方法通过戊二醛将α-淀粉酶固定在一种水稳定的金属-有机骨架材料(Metal-Organic Frameworks,MOFs)MIL-53(Al)-NH_(2)的表面,得到固定化酶。固定化酶与葛根95%乙醇提取物的水溶液相作用,利用抑制剂与酶活性位点特异性结合的特点,依据配体垂钓策略,从中萃取出与α-淀粉酶发生作用的化学成分。结果α-淀粉酶成功固定在MIL-53(Al)-NH_(2)表面,且具有酶活性;垂钓筛选的化合物经HPLC分离,采用对照品比较法,确定了11种活性成分。通过分子对接技术,进一步确认这11种活性成分与α-淀粉酶的结合情况。结论本研究首次将α-淀粉酶固定于MIL-53(Al)-NH_(2)上,建立了基于MOFs-固定化酶的配体垂钓方法,该方法可用于从复杂的混合物中筛选生物活性成分,这为MOFs更多地用于酶固定化提供了新思路,也为基于酶功能化的MOFs材料的配体垂钓用于中药中的活性组分特异性识别和筛选提供了参考。OBJECTIVE To screen the substances withα-amylase inhibitory activity from 95%ethanol extract of Puerariae Lobatae Radix using ligand fishing strategy.METHODSα-Amylase was immobilized on the surface of MIL-53(Al)-NH_(2),which was a water-stable Metal-Organic Frameworks(MOFs),by glutaraldehyde to obtain the immobilized enzyme.According to the ligand fishing strategy,the immobilized enzyme interacted with the aqueous solution of 95%ethanol extract of Puerariae Lobatae Radix to extract those chemical components that could interact withα-amylase.RESULTSα-Amylase was successfully immobilized on the surface of MIL-53(Al)-NH_(2)and had enzymatic activity.The compounds screened by fishing were separated by HPLC,and 11 active compounds were determined by the comparison with reference substances.Besides,the binding of these 11 active compounds toα-amylase was further confirmed by molecular docking technology.CONCLUTIONα-Amylase is immobilized on MIL-53(Al)-NH_(2)for the first time,and establish a ligand-fishing method based on MOFs-immobilized enzymes,which can be used to screen bioactive components from complex mixtures.In addition,this study provides a new idea for more use of MOFs for enzyme immobilization,and a reference for the specific identification and screening of active components from traditional Chinese medicine based on ligand-fishing of enzyme-functionalized MOFs.

关 键 词：MIL-53(Al)-NH2 Α-淀粉酶 配体垂钓 固定化酶 葛根提取物 分子对接 

分 类 号：R945[医药卫生—微生物与生化药学]