基于血清药物化学和代谢组学的蒙药地格达-4味汤抗急性肝损伤的物质基础及作用机制分析

Digeda-4 decoction with the significant hepatoprotective efficacy is the classic Mongolian Medicine, but its potential basis and action mechanism are not yet understood. In the previous study, when the rats with acute injury in liver were intervened by the Digeda-4, their blood can be measured the medicine prototype ingredients and metabolites.Besides,and also there are unusual creature maker peaks of chromotagraph.It shows that changes of creature maker related to intervention of bloob component that Digeda-4 decoction was absorbed in. Therefore, based on the preliminary studies, the project to be declared treats rats with acute liver injury by D-GlaN as the object in experiment and intervened by the Digeda-4 ,and then establishes the hypothesis that bloob component that Digeda-4 decoction was absorbed in is the material basis of creature maker changes and hepatoprotective efficacy in creasures with acute liver injury .With the apparatus such as liquid chromatography-mass spectrometry and nuclear magnetic resonance chromatographic techniques, uniting chemicals group and metabolomics research model, hepatoprotective active ingredient groups were screened. Also researching the model animal liver tissue, serum and urine metabolomics were operated and those all contribute to build metabolic networks of potential biological markers and sketch map of adjusting the network diagram with Digeda-4 decoction. At the same time, it clears Digeda-4 decoction's protective material basis for optimization on multi-link, multi-channel, multi-target studies and laid the foundation for future study on Digeda-4 decoction.

地格达-4味汤是具有显著保肝功效的蒙医经典方。但仍未弄清其药效物质基础和作用机制。在前期研究中发现，地格达-4味汤水提物干预急性肝损伤大鼠后，血液中出现了地格达-4味汤原型成分和代谢产物以外，还有异常的生物标示物色谱峰。推测生物标示物变化与地格达-4味汤入血成分的干预有关。因此，拟申报课题在前期研究基础上，以D-GlaN致急性肝损伤模型大鼠为研究对象，以地格达-4味汤水提物为示例药物，建立"地格达-4味汤入血成分是使急性肝损伤动物体内生物标示物发生变化和产生保肝作用的物质基础"的工作假说，利用液质联用和核磁共振色谱技术，联合化学物质组和代谢组学的研究模式，筛选保肝活性成分群，并进行模型动物肝组织、血清和尿液代谢组学研究，构建急性肝损伤潜在生物标示物代谢网络和地格达-4味汤调节该网络的示意图。同时，明确地格达-4味汤多环节、多途径、多靶点的保肝物质基础，为地格达-4味汤优化研究奠定了基础。

蒙药地格达-4味汤由肋柱花、栀子、胡黄连、瞿麦等四味药组成，具有保肝利胆作用，是临床常用的保肝药物，体现药物临床应用特点，以“体内化学成分”为中心，结合机体疾病状态、生物学效应“两要素”，观察蒙药地格达-4味汤的保肝作用的基础上分析了在模型大鼠体内移行成分及保肝作用机制。.地格达-4味汤除了能抑制D-GlaN和CCL4致大鼠急性肝损伤之外还能显著降低 ANIT 致大鼠胆汁淤积大鼠模型血清 的 AST、ALT、ALP、TBIL 的活性，并且能有效缓解肝组织的病理改变。地格达-4 味汤能明显降低血清 TNF-α,IL-6、 水平，有显著性差异（P<0.O1 或 P<0.05）。地格达-4 味汤低能明显升高 IL-10 水平，能明显提升血清和肝组织的 GSH-PX水平，有显著性差异（P<0.O1 或 P<0.05）。地格达-4 味汤还能明显降低肝组织 MDA 水平，提高血清和肝组织 SOD 水平，但没有统计学意义（P>0.05）。初步认为地格达-4 味汤可以通过炎性反应及抑制氧化应激这一机制来治疗肝内胆汁淤积，改善 ANIT 诱导的肝内胆汁淤积大鼠模型的生化指标，起到保肝作用。.通过液质联用仪（LC-MS）和核磁共振谱（NMR）分析地格达-4味汤在急性肝损伤状态下大鼠血清、肝、肺、心、肾、脾、尿液中的移行成分，结果在体内共检测到地格达-4味汤来源的20个移行成分，其中直接来源于地格达-4味汤的原型成分有9个，分别为草夹竹桃苷、獐牙菜苦苷、β-谷甾醇、棕榈酸、京尼平苷、胡黄连苷Ⅱ、4-0--glucopyranosylacetoPhenone2,3,4-Trihydroxy-6-[2-(3,4,5-trihydroxy-6-hydroxymethyl-tetrahydro-pyran-2-yloxycarbonyl)-vinyl]-benzoic acid methyl este、Sweroside；地格达-4味汤在体内新产生的代谢物为11个。进而利用RT-PCR法、Western blot法测定蒙药地格达-4味汤对目标基因的影响，分析其该对幽门结扎性肝损伤的保护作用机制。结果蒙药地格达-4味汤各给药组与模型组比较地格达-4味汤对AST、ALT、BILD2、BILT3、TBA、ALP明显降低（P < 0.01或P < 0.05），GSH-PX明显增高（P < 0.01）；外