基于体内动态变化成分民族药滇白珠抗风湿药效物质基础及其质量控制研究

This project is the continuation and further study of the finished 81001697 NSFC. Compared with monomer salicylic acid methyl glucoside which has good anti-inflammatory activity, the active parts of Dianbaizhu, form the plant Gaultheria leucocarpa var. yunnanensis (Franch.) T. Z. Hsu & R. C. Fang, process better activity. Not only the salicylic acid methyl glucoside, the active parts also contain some anti-inflammatory and analgesic activity compounds like flavones and organic acid. In view of the effective substances of Chinese/folk medicine should being various prototype group and active metabolites group that process suitable dynamic characteristics in vivo, using the active part of Dianbaizhu as the object, and chemical components in vitro, in normal and pathological body combined pharmacokinetics and pharmacodynamics (PK/PD) as the research method, basing on the technology platform of analytical test, cell experiment and animal experiment, this project carries out the study of gastrointestinal bioconversion and absorption, constituent isolation and identification, the analysis and identification of the constituent in normal and pathological body, and the establishment of their relationship of the time-concentration-efficiency. This project aimed at clarifying anti-inflammatory and analgesic metabolites in vivo and prototype in vitro and their change law, and further speculate the mechanism of action. Otherwise, establishing the multidimensional spectrum of active part fingerprint, dose-effect fingerprint and drug metabolism fingerprint, in order to control quality of Dianbaizhu and guide rational use of drugs.

本项目是已结题国家自然基金项目延续和深入。民族药滇白珠在多个民族中用于治疗风湿疾病，但是其活性成分颇有争议。基于滇白珠大极性部位有显著抗风湿活性，其所含多类成分均有很好的抗炎镇痛活性，申请者提出“滇白珠是多成分整体发挥抗炎镇痛作用”的工作思路。由于药物发挥药效是体内具有适宜动力学特征多种原形成分组及活性代谢物组的整体作用，本项目拟以前期化学成分、药理活性和分析方法为基础，以滇白珠抗风湿活性部位为对象，以体外化学成分、正常和病理状态下体内成分及药代动力学与药效动力学（PK/PD）相结合的方法，开展胃肠道生物转化吸收和成分分离鉴定，正常和病理机体体内成分确认分析及其体内发挥药效时间-浓度-效应三者关系建立的研究，阐明滇白珠抗炎镇痛体内代谢成分和体外原型有效成分及其变化规律，推测其抗风湿作用机制；建立活性部位指纹谱、量效指纹谱以及药物代谢指纹谱的多维抗风湿组分谱从而有效科学的质量控制和指导用药。

民族药滇白珠在多个民族中用于治疗风湿疾病，但其抗风湿药效物质基础却颇有争议。本项目以滇白珠抗风湿活性部位（ARF）为对象，经拓扑分析搭建了“疾病-通路-靶点-药物”相互作用网络，促进从系统生物学角度理解ARF抗风湿物质基础。应用UPLC-ESI-Orbitrap-MSn技术对ARF及其在AIA模型大鼠血浆、人工胃肠液和两个人的肠内菌模型中的原形成分和代谢产物进行表征，建立了ARF的原形成分和代谢产物的化学成分库，验证了ARF在胃肠道内的相对稳定性。16S rRNA菌群测序结果提示ARF能显著改变模型大鼠肠道菌群结构。基于离体外翻肠囊法确定了ARF吸收机制，结果表明：ARF部位中MSTG-B、（+）-catechin、MSTG-A、Gaultherin、chlorogenic acid、quercetin、kaempferol在四个肠段中皆有吸收，各自吸收难易程度不一，总体表现为空肠＞十二指肠≈回肠＞结肠；同一种成分的吸收方式可能是被动扩散或与主动转运同时存在。采用房室模型和统计矩模型分析了ARF在正常和模型大鼠体内的动力学过程，检测并发现MSTG-B、MSTG-A、Gaultherin均在1.81-2.93 h内达峰，均符合一室模型，且可能存在相互影响；结合以TNF-α、IL-1β、IL-6、IL-2为药效指标的PD模型构建了PK-PD模型，说明MSTG-B、MSTG-A、Gaultherin可以通过调节炎症因子的水平来发挥抗风湿作用。本课题建立了ARF药效成分的体内外定量分析方法，考察了其在胃肠道稳定性及预测作用靶点及机制，筛选出了最佳吸收肠段及PK、PD模型，并构建了药-时-效曲线，为阐明ARF药效物质基础、临床开发、剂型设计等提供了数据支撑及基础依据。研究成果已发表论文7篇，SCI收录3篇，正在撰写4篇文章，公开发明专利1项，培养硕士3名，博士1名。