基于斑马鱼模型探究药食同源复方降尿酸作用及成分分析

金灵泰; 张铭; 方双琪; 徐蔷

doi:10.13386/j.issn1002-0306.2022110251

基于斑马鱼模型探究药食同源复方降尿酸作用及成分分析

Study on Lowering Uric Acid Effect and Component Analysis of Drug Food Homologous Compound Based on Zebrafish Model

摘要

摘要: 目的：探究芹菜籽、蒲公英、菊苣和玉米须四种药食同源中药联用下的降尿酸活性和作用机制，分析这四种中药复合提取物中具备降尿酸作用的主要效应成分。方法：随机选取5 dpf野生型AB品系斑马鱼，通过250 μmol/L氧嗪酸钾和10 μmol/L黄嘌呤钠盐诱导建立斑马鱼高尿酸模型，将已建立的斑马鱼模型随机分为对照组、模型组、别嘌醇组（136 µg/mL）和复方提取物低、中、高（250、500及1000 µg/mL）剂量组，每组30尾，连续干预24 h后，测定斑马鱼体内的尿酸荧光值和次黄嘌呤磷酸核糖转移酶1（HPRT1）、葡萄糖转运蛋白9（GLUT9）、有机阴离子转运体（OAT1）基因表达水平；采用超高液相色谱-串联高分辨质谱仪为主要手段，以乙腈-甲酸水为流动相，梯度洗脱，ESI离子源正、负离子扫描模式，结合软件数据库搜索及相关文献进行成分鉴定。结果：与模型组相比，复方提取物低、中、高剂量组均能极显著降低高尿酸斑马鱼模型的血尿酸水平（P<0.01），且能够有效上调高尿酸斑马鱼体内HPRT1、OAT1的相对表达量、下调GLUT9的相对表达（P<0.05）；初步筛选鉴定出黄酮及其苷类、有机酸、香豆素、萜类等22种活性成分，均为复方提取物中具有降尿酸机制的效应物质。结论：复方提取物可能通过上调模型组斑马鱼体内HPRT1、OAT1的表达量、下调GLUT9的表达，发挥多方位降尿酸的作用；所筛选鉴定的化学成分为下一步筛选作用靶点提供参考。

Abstract: Objective: In order to explore the uric acid-lowering activity and mechanism of four traditional Chinese medicines, celery seed, dandelion, chicory and corn silk, and analysis of the main functional components with uric acid-lowering effect in these four kinds of traditional Chinese medicine compound extracts. Methods: Hyperuricemia model in zebrafish was induced by potassium oxazinate and sodium xanthine, then they were randomly divided into control group, model group, allopurinol group (136 µg/mL) and compound extract low, medium, high doses group (250, 500 and 1000 µg/mL), with 30 rats in each group. After continuous intervention for 24 h, the fluorescence level of uric acid and the gene expression levels of hypoxanthine phosphoribosyltransferase1 (HPRT1), glucose transporter9 (GLUT9), and organic anion transporter (OAT1) in zebrafish were measured. Using liquid chromatography-series high resolution mass spectrometer as the main method, and acetonitrile formate water as the mobile phase, gradient elution, ESI ion source positive and negative ion scanning mode, and then combined with software database search and related literature for component identification. Results: Compared with the model group, the low, medium and high dose groups of the compound extract could reduce the blood uric acid level of the hyperuric acid zebrafish model (P<0.01), and could effectively up-regulate the relative expression of HPRT1 and OAT1 and down-regulate the relative expression of GLUT9 in the hyperuric acid zebrafish (P<0.05). At the same time, 22 active ingredients including flavonoids and their glycosides, organic acids, coumarins, and terpenes were initially screened and identified, all of which are effective substances in the compound extract with a uric acid-lowering mechanism. Conclusion: The compound extract may play a multifaceted role in lowering uric acid by up-regulating the gene expression of HPRT1, OAT1, and down-regulating the gene expression of GLUT9 in the model group of zebrafish. The chemical components screened and identified can provide a reference for the next step of screening the target.