基于体内过程"层间递进"的中药注射剂药效物质筛选新方法

For the traditional Chinese medicine injection is so complicated that it is hard to identify the pharmacodynamic material basis. Our group has built the research model of chemical materials - metabolites - pharmacokinetic makers - in vitro activity evaluation - pharmacodynamic evaluation in animal models. In our research, Xuebijing injection (XBJ) has been used as a model drug. In order to find out major metabolic pathways and metabolic products of XBJ, the biological sample in plasma, bile, urine and feces will be analysised and identified by mass defect filtering (MDF) of UPLC-Q-TOF/MS after intravenous administration of XBJ to rats. Then, the component of high exposure levels in plasma (prototype drug and metabolites) will be determined by LC-MS/MS, after evaluated pharmacokinetics study, the pharmacokinetics makers of XBJ were found. To clarify the material basis for anti-inflammatory effects of XBJ, the pharmacokinetic makers extracted from the liver microsomal incubations sample in vitro by online fishing and knockout technology, will been evaluated anti-inflammatory activity. Finally, the research ideas of fast screening pharmacodynamic material basis from Chinese medicines by the "whole - part" mode could provide a reference for other Chinese medicines material basic research.

为快速研究中药注射剂药效物质基础，本课题提出了以体内过程分析为切入点，构建化学物质组-代谢物质组-药代标示物-体外活性评价-整体动物药效评价，"层间递进"的中药药效物质筛选模式。以临床疗效确切方剂-血必净注射液为模型药物，在对血必净化学物质及入血原型成分相对清楚的基础上，采用UPLC-Q-TOF/MS 定性分析结合质量亏损过滤(MDF)技术，对整体动物注射血必净后的血浆、胆汁、尿液、粪便进行代谢产物的分析和鉴定，明确其主要成分的代谢产物及代谢途径。 通过HPLC-MS/MS定量分析，对血浆中有较高暴露水平的成分（药物原型、代谢产物）进行药动学评价，确定药代标示物（PK-M）。最后，对药代标示物进行体内、外抗炎活性评价，从而阐明血必净抗炎作用物质基础。本项目体现了从众多中药成分中由"整体"到"部分"快速寻找、筛选药效物质的研究思路，为中药注射剂药效物质基础研究提供了可借鉴的模式。

如何确定中药药效物质，一直是中药现代化研究中的难点和热点。本研究以体内过程分析为切入点，建立了化学物质组-代谢物质组-药代标示物-体外活性评价-整体动物药效评价，“层间递进”的中药药效物质筛选模式。利用UPLC-Q-TOF/MS技术对血必净注射液中的主要化学成分进行定性分析，并在此基础上，利用UPLC-MS/MS法对其15种化学成分开展了定量分析；随后利用质量亏损过滤技术，对大鼠注射血必净后的血浆、胆汁、尿液、粪便进行代谢产物的分析和鉴定，明确其主要成分的代谢产物；通过HPLC-MS/MS 定量分析，对血浆中具有较高暴露水平的9种成分进行药动学评价，确定了药代标示物；最后针对药代标示物开展了对LPS诱导的RAW264.7细胞分泌炎症因子及对脓毒血症大鼠炎性因子TNF-α、IL-1、IL-6、IL-8和IL-10表达的影响的研究，表明血必净药代标示物可抑制脓毒血症大鼠血浆中TNF-α、IL-1、IL-8的表达。本课题以体内过程分析为切入点，层层筛选，体现了从众多中药活性成分中根据其药代属性优劣由“整体”到“部分”快速确定药效物质的研究思路，为中药注射剂药效物质基础研究提供了可借鉴的模式。现已发表学术论文5篇；培养研究生3名。