基于超高效液相色谱-四极杆飞行时间质谱结合网络药理学探讨杜仲治疗糖尿病肾病的有效成分

Active ingredient analysis of Eucommia ulmoides Oliv. in treating diabetic nephropathy by using UPLC-Q-TOF-MS and network pharmacology

目的采用超高效液相色谱-四极杆飞行时间质谱(UPLC-Q-TOF-MS)技术分析传统中药杜仲水提液中主要化学成分,并结合网络药理学方法预测其中治疗糖尿病肾病(DN)的有效成分,探讨其潜在的作用机制。方法根据水提液色谱信息和质谱信息,并结合对照品的质谱碎片裂解规律和文献数据,对杜仲水提液化学成分进行鉴定;利用化合物靶点数据库和疾病靶点数据库,预测杜仲活性成分作用的潜在靶点,并筛选出治疗DN的相关靶点;通过String等网络工具,确定核心成分与靶点;将上述关键核心靶点数据导入到DAVID数据库中进行基因本体(GO)和基因功能(KEGG)通路注释分析,分析靶点关联的主要生物代谢通路;最后采用Cytoscape 3.7.0软件构建中药杜仲"活性成分-靶点-DN-通路"网络关系图。结果通过UPLC-Q-TOF-MS技术共筛选出杜仲水提取液中42个化学成分,其中34个为活性成分,预测出190个潜在的作用靶点;利用网络公共数据平台和网络药理学分析工具检索到224个与DN相关的疾病靶点,筛选出69个核心靶点,3个潜在的核心成分,其中包括桃叶珊瑚苷和咖啡酸;并通过DAVID数据库进行富集分析,找出了治疗DN的10条核心通路,涉及增殖、凋亡、炎症反应、纤维化等生物学过程。结论本研究初步预测了杜仲水提液中治疗DN的主要有效成分,其作用机制可能与细胞增殖分化、氧化应激、炎症反应、纤维化等方面有关。

AIM To study and analyze the main chemical constituents of Eucommia ulmoides Oliv. water extract by ultra performance liquid chromatography quadrupole time of flight mass spectrometry(UPLC-Q-TOF-MS), to predict the effective substances for treating diabetic nephropathy(DN)according to network pharmacology, and to explore the potential mechanism. METHODS According to the chromatographic information of retention time of chromatographic peaks and ultraviolet absorption characteristics, accurate relative molecular weight, secondary mass spectrometry fragments and other mass spectrometry information, and combined with the fragmentation law of mass spectrometry and literature data, the chemical constituents of Eucommia ulmoides Oliv. water extract were identified. The potential targets of Eucommia ulmoides Oliv. were predicted using database of target for compound to predict potential targets of the active ingredients, and the action targets related with DN were screened from disease-associated data. Identify core components and core targets through network tools such as String. GO and KEGG pathways of key targets were enriched by DAVID database, and the main metabolic pathways associated with targets were analyzed. Finally, the network diagram of "active ingredient-target-disease-pathway" of Eucommia ulmoides Oliv. was constructed by Cytoscape software(version 3.7.0). RESULTS Forty-two chemical constituents of Eucommia ulmoides Oliv. water extract were analyzed by UPLC-Q-TOFMS technology, among which thirty-four active constituents were identified. A total of one hundred and ninety potential targets were predicted. Two hundred and twenty-four disease targets related to DN were retrieved through network public database. Sixty-nine core targets were screened by analysis tool of network pharmacology, and three core components, including aucubin and caffeic acid. Through the DAVID database for enrichment analysis, we concluded that Eucommia ulmoides Oliv. had 10 core non-extensive signaling pathways in the treatment of DN, involving biological processes such as proliferation, apoptosis, inflammatory response and fibrosis. CONCLUSION The main effective components of Eucommia ulmoides Oliv. water extract in the treatment of DN are preliminarily predicted in this study. The mechanism of action may be related to cell proliferation and differentiation, oxidative stress, inflammatory response, fibrosis and so on.