土茯苓中黄嘌呤氧化酶抑制剂筛选鉴定研究  

Screening and identification of xanthine oxidase inhibitors from Smilax glabra

作　　者：邹惠亮 陈柯 王新财 成霄 孟令杰 刘百联[3] 陈敏 ZOU Hui-liang;CHEN Ke;WANG Xin-cai;CHENG Xiao;MENG Ling-jie;LIU Bai-lian;CHEN Min(Huzhou Institute for Food and Drug Control,Zhejiang,Huzhou 313000,China;Institute of Life Sciences,Zunyi Medical University,Zunyi 563000,China;Institute of Traditional Chinese Medicine and Natural Products,College of Pharmacy,Jinan University,Guangzhou 510632,China)

机构地区：[1]湖州市食品药品检验研究院,浙江湖州313000 [2]遵义医科大学生命科学研究院,贵州遵义563000 [3]暨南大学药学院中药及天然药物研究所,广东广州510632

出　　处：《中成药》2024年第5期1558-1564,共7页Chinese Traditional Patent Medicine

基　　金：湖州市科技计划项目(2020GZ12)。

摘　　要：目的采用磁珠固定化酶和LC-MS/MS联用技术快速筛选土茯苓中的黄嘌呤氧化酶抑制剂,明确土茯苓中的降尿酸活性成分。方法以羧基磁珠作为载体通过共价偶联制成固定化黄嘌呤氧化酶。以固定化酶的特异性吸附作用对土茯苓中的黄嘌呤氧化酶抑制剂进行亲和筛选,以LC-MS/MS和对照品进行分析对比,并对筛选鉴定出的成分进行抑制活性和抑制类型进行考察。结果扫描电镜和红外光谱表征了固定化黄嘌呤氧化酶的成功合成,酶固载量为70.50μg/mg,相对活力为79.44%。从土茯苓提取物中共筛选出13个活性化合物,并鉴定了11个成分。酶活性试验表明黄杞苷和异黄杞苷的抑制活性最强,与阳性对照别嘌呤醇接近,其IC_(50)值和抑制类型分别为32.25μg/mL、混合型抑制,35.12μg/mL、竞争性抑制。结论本方法直接从土茯苓提取物中筛选鉴定具有黄嘌呤氧化酶抑制活性的成分,简单、快速、准确,适用于复杂中药体系中黄嘌呤氧化酶抑制剂的高通量筛选。AIM To rapidly screen xanthine oxidase(XOD)inhibitors from Smilax glabra Roxb.by enzyme-immobilized magnetic microspheres and LC-MS/MS,and to confirm the anti-uric acid constituents from S.glabra Roxb.METHODS The immobilized xanthine oxidase was prepared by covalent coupling with carboxyl magnetic beads as a carrier.The xanthine oxidase inhibitors in S.glabra were screened by the specific adsorption of immobilized enzyme.LC-MS/MS and standard substances were used for analysis and comparison,and the inhibitory activity and inhibition type of the screened and identified components were investigated.RESULTS The successful synthesis of immobilized xanthine oxidase was characterized by scanning electron microscopy and infrared spectroscopy.The enzyme loading was 70.50μg/mg and the relative activity was 79.44%.Thirteen active compounds were screened from the extract of S.glabra,and eleven compounds were identified.The enzyme activity test showed that the inhibitory activites of engeletin and isoengeletin were the strongest,which was close to the positive control allopurinol.The IC_(50)value and inhibition type were 32.25μg/mL,mixed inhibition,35.12μg/mL,competitive inhibition.CONCLUSION The method is simple,rapid,accurate and suitable for directly screened active ingredients which can inhibit XOD from complex extract of traditional Chinese medicines.

关 键 词：土茯苓 固定化酶 磁珠 抑制剂 黄嘌呤氧化酶 

分 类 号：R284.1[医药卫生—中药学]