白术芍药散“炮制增效”多靶点作用机理研究

Baizhu Shaoyao powder is a famous traditional Chinese medicine prescription,consisting of the stir-fried Atractylodes macrocephala, stir-fried white peony root, stir-fried Citrus reticulata and crude Saposhnikovia divaricata, which is mainly used to treat irritable bowel syndrome in clinical. On the basis of our previous research, this project proposes the following hypothesis: the main components of the three-flavor Chinese medicines are changed after processing, and the interaction with certain gastrointestinal microbes may alter hormones and.neurotransmitters secretion, impact TLR4/NF-κB signaling pathway and improve irritable bowel syndrome. This project integrated UPLC-Q-TOF/MS analysis technique, 16S rRNA gene sequencing technology and ELISA, Western Blot and other molecular biology techniques to investigate the chemical basis and metabolites of Baizhu Shaoyao powder, intestinal flora distribution, fecal exogenous metabolites,.gastrointestinal hormones and neurotransmitters (vasoactive peptides, substance P, etc.) levels and TLR4 /NF-κB signaling pathway. Collectively, the enhanced role of multi-target effect of Baizhu Shaoyao powder via processing will be clarified from different levels.

白术芍药散是中医传统名方，由炒白术、炒白芍、炒陈皮和防风 4 味药组成，临床主要用来治疗肠易激综合征。在前期的研究基础上，本项目提出以下假说：白术芍药散中3味中药炮制后主要成分发生改变，通过与肠道微生物菌群发生相互作用，改变某些胃肠激素和神经递质的分泌，调控TLR4/NF-κB信号通路，改善肠易激综合征的各种病理状态，发挥炮制增效的作用。项目选择白术芍药散为研究对象，应用UPLC-Q-TOF/MS分析技术、16S rRNA基因测序技术及ELISA、Western Blot等分子生物学技术，系统研究白术芍药散炮制效应物质基础、血液、尿液及粪便外源性代谢产物、肠道菌群分布、粪便内源性代谢物、胃肠激素和神经递质血管活性肽、P物质、生长抑素等）及TLR4/NF-κB信号通路调控，从整体、组织及分子水平阐明白术芍药散炮制增效的多靶点作用内涵。

项目首先研究了炒制对于白术芍药散化学成分的影响，发现针对非挥发性成分，炒品组和生品组间异性化学成分34种；针对挥发性成分，炮制前后样品中供鉴定出145种成分，其中炒制后消失的成分15种，新出现成分17种。考察生品和炒品大鼠血浆中25种原型成分行为学特征，发现16种化学成分炒制后具有显著性差异：其中14种成分在炒品组大鼠血浆中含量响应高，2种成分在生品组大鼠血浆中含量响应高。从生理指标、病理指标、行为学指标等方面全面评价白术芍药散对大鼠D-IBS模型治疗作用，结果说明饮片经过炒制后组方对IBS-D治疗作用更佳，可显著改善IBS-D大鼠脑-肠轴异常和炎症反应症状。通过16S rRNA测序技术对大鼠粪便样品进行肠道菌群测序分析，发现造模后IBS-D大鼠菌群组成和丰度发生改变，而饮片经炒制后组方能够更有效地调节IBS-D大鼠肠道菌群紊乱的状况，从而改善腹痛、腹泻等IBS-D相关症状。