基于多维谱效及分子对接的温郁金加工炮制药效物质及机制研究

Traditional Chinese Medicine(TCM) processing is an important part of Chinese herbal pieces production process, and it is also the characteristic property of Chinese clinical medicine. But the processing theory which instructs the reasonable application of Chinese herbal pieces is still lacking modern scientific basis, which impedes the development of TCM Industry. The Curcuma wenyujin is processed to Pianjianghua、Ezhu and CuEzhu by fresh processing, steaming and vinegar processing. All kinds of Curcuma wenyujin pieces have the effect of promoting blood circulation for removing blood stasis and regulating qi-flowing for relieving pain, but being different in clinical application, and until now there is no research on the mechanism of Chinese herbal pieces effect changes. Based on the problems mentioned above and the prior research, the project innovatively introduces multidimensional spectrum-effect, molecular docking into TCM processing. Pharmacodynamic material and target protein as the breakthrough point, through the multidimensional spectrum-effect relationship to identify effective material group, molecular docking technology to screening target protein, combining with metabolomics to evaluate the relevance between the effective material group、target protein and metabolic components. The research is to clarify the connotation of scientific hypothesis ---curcuma wenyujin processing is “multivariate regulation of target protein, plasmin, inflammatory cytokin, endogenous metabolite and metabolic pathway through the effective material group consisting of pieces of the prototype and metabolite, which causes changes in the effect of promoting blood circulation for removing blood stasis and regulating qi-flowing for relieving pain”. The project intends to systematically interprete TCM processing theory from multi-ingredients, multi-targets and multi-way, providing scientific basis for the clinical application of Chinese herbal pieces and a demonstration for the modern research of Chinese herbal pieces.

中药加工与炮制是饮片生产过程的重要环节，为中医临床用药特色，但指导饮片合理应用的加工炮制理论尚缺乏现代科学论据，成为制约中药产业发展的瓶颈。温郁金根茎经趁鲜加工、蒸制及醋制，分别制得片姜黄、生莪术及醋莪术饮片，各饮片均具行气破血止痛功效，但临床应用异同，其效应变化机制未见研究。针对上述问题及前期研究，项目创新性地将多维谱效、分子对接等前沿技术首次引入中药加工炮制研究领域。以药效物质及靶蛋白等为切入点，通过多维谱效关系辨识有效物质群，分子对接筛选出相应靶蛋白，结合代谢组学评价有效群、靶蛋白与代谢组分的关联性。研究阐明温郁金加工炮制是“通过各饮片原型物及代谢产物组成的有效物质群，多元调控靶蛋白、纤溶酶、炎症因子、内源性代谢物及代谢通路等，从而导致行气破血止痛效应的差异变化”假说的科学内涵。从多成分、多靶点、多途径系统诠释中药炮制理论，为饮片临床应用提供科学依据，也为中药饮片现代研究提供示范。

温郁金为临床常用中药，其根茎经趁鲜加工、蒸制及醋制后分别制得片姜黄、生莪术及醋莪术饮片，三者均具活血止痛功效，但临床应用各有侧重。项目通过现代前沿技术辨识效应物质，分子对接筛选出相应靶蛋白，结合网络药理学和代谢组学评价有效群、靶蛋白与代谢组分的关联性，研究阐明温郁金各饮片效应物质及作用机制。.采用LC/MS等进行有效物质群研究，结果显示莪术二酮等在片姜黄、生莪术、醋莪术中依次降低，而莪术醇等先升高后降低；Curcumenolactones A等在生莪术中的含量低于片姜黄，但在醋莪术中含量升高；表明加工炮制对药效物质产生不同程度的影响。活血化瘀及止痛药效评价结果显示，片姜黄对FIB、PAI-1无调节作用；生莪术低剂量组对PAI-1、IL-6、TNF-α无调节作用，醋莪术低剂量组对TT、APTT、PAI-1、IL-6无改善作用，各高剂量组均对血液流变学等相关检测指标具有显著改善作用，且醋莪术高剂量组效果最佳；温郁金三种饮片对原发性痛经均有显著改善作用，醋莪术疼痛相关因子的调控作用更显著。谱-效相关性研究表明，片姜黄中吉马酮等5个成分、生莪术中吉马酮等3个成分以及醋莪术中莪术二酮等5个成分与凝血四项等药效指标相关性较高，表明三种饮片药效的异同与其效应物质变化相关。代谢组学研究表明，温郁金三种饮片对气滞血瘀症的改善作用主要与调控脂质代谢相关，通过抑制并去除氧自由基、抑制COX-2的表达、减少5-HT合成进而抑制血小板聚集从而缓解痛经，其中醋莪术调控作用最显著。网络药理学研究表明，钙信号通路、MAPK信号通路等均与抗血小板聚集相关；分子对接研究显示，CCR5及MAPK1为治疗原发性痛经关键靶点，且均为抗血小板聚集相关通路的重要靶点。温郁金各饮片均具有较好的体外抗血小板活性，且醋莪术抗血小板聚集作用最强，该结果验证了网络药理学和分子对接的预测结果。综上表明温郁金不同饮片可通过调控脂质代谢、抗血小板聚集途径发挥化瘀止痛功效。.项目研究结果表明，温郁金加工炮制是通过对各饮片莪术二酮等原型物及其代谢组分等效应物质的影响，多元调控5-HT等靶蛋白、PAI-1等炎症因子、脂质代谢物及MAPK等代谢通路，从而发挥不同活血化瘀止痛作用。从多成分、多靶点、多途径系统诠释中药炮制理论，为温郁金不同饮片临床应用提供科学依据，也为中药饮片现代研究提供示范。