基于AMPK激活作用的黎药-野菠萝双四氢呋喃型木脂素降脂活性成分及其构效关系研究

The water extract of the fruits of Pandanus tectorius Soland was confirmed to possess obvious lipid-lowering activity in our previous study. Further pharmacological investigations have been successfully carried out by which the active fraction was determined. Tentatively chemcial investigation indicated that the active fraction is rich in Furofuran lignans (FFL). Further more, constituents of FFL obtained from PTF showed significant lipid-lowering activities and the hypolipidemic mechanism was found to be closely associated with activating the AMP-activated protein kinase (AMPK). On the basis of previous work, we aim to systematically investigate furofuran lignans (FFL) with hypolipidemic effects in the aqueous extractions of Pandanus tectorius fruits. Based on the methodology integrated LC-MS dereplication, a variety of chromatography technology (i.e. medium-pressure preparation and semi-HPLC) and spectroscopic approaches including NMR, CD and X-ray, FFL will be rapidly screened, isolated and elucidated. Meanwhile, the mechanism and structure-activity relationships of FFL will thoroughly analyzed using methods of molecular biology. In addition, the binding degree between bioactive FFL and AMPK of the obtained compounds will be in vitro evaluated on HepG2 cell combined with computer aided drug design (CADD). Studies on activation against intracellular AMPK or p-AMPK of the effective constituents will also be carried out. Activators of AMPK or p-AMPK from the FFL constituents with the significant lipid-lowering effects will be screened using hamster model of hyperlipidemia. In this way, our findings will establish solid scientific foundation, facilitating not only illustrate the therapeutic basis of this Li medicine but also find new lead compounds with independent intellectual properties and development of new drugs.

课题组研究证实黎药-野菠萝具有显著的降血脂作用，并进一步确定其有效部位。深入研究发现有效部位中富含新颖的双四氢呋喃型木脂素（FFL），且此类成分可通过激活腺苷活化蛋白激酶（AMPK）起到降脂作用。本项目拟在前期基础上，整合中药化学、分子药理学、分子生物学及药理学等多学科的先进方法与技术，针对有效部位中FFL类成分的组成、降脂活性及构效关系进行研究。包括对FFL类成分快速识别后，实现目标化合物的分离与鉴定；以AMPK为靶标，采用FlexX程序将化合物与受体进行分子对接，评价它们相互作用的强弱；利用细胞模型对结合较好的成分进行降脂活性评估，并应用蛋白印迹方法检测活性成分对细胞内AMPK的激活作用，进而深入剖析野菠萝FFL类成分的构效关系；优选活性较强、作用机制基本清晰的成分，经足量制备后，利用金黄地鼠高脂血症模型观察其降脂功效，阐明黎药-野菠萝的药效物质基础，发现具有新药开发前景的先导化合物。

野菠萝为露兜树科（Pandanaceae）露兜树属（Pandanus）植物露兜树（Pandanus tectorius Soland.）的果实，由于其形态奇特，形似菠萝，在黎族民间习称“野菠萝”。野菠萝资源十分丰富，在海南黎族作为药物治疗高脂血症疗效显著。课题组前期研究发现，野菠萝富含的双四氢呋喃型木脂素类成分（FLL）降脂作用显著，并且其功效与激活腺苷活化蛋白激酶（AMPK）密切相关。基于此，本项目开展基于AMPK激活作用的黎药-野菠萝中降脂活性双四氢呋喃型木脂素发现及其作用机制研究。. 通过系统的研究，本项目从野菠萝中发现40个化合物，新化合物2个，其中FFL类成分10个，并首次发现咖啡酰奎宁酸类成分是野菠萝中另一主要成分。利用体外模型完成化合物的降脂活性评价，确定FFL类成分及咖啡酰奎宁酸类成分为活性成分。通过体外模型深入探索活性最佳FFL成分的降脂作用，并进行降脂机制探索。进一步利用与人类高脂血症、二型糖尿病相近的动物模型，开展富含咖啡酰奎宁部位的降血脂、降血糖作用，并应用代谢组学、分子生物学等方法深入探索其作用机制。通过以上研究，分别在国际主流期刊PlosOne，Journal of Nutritional Biochemistry上发表学术论文，另外在Records of Natural Products 等期刊上发表学术论文3篇，申请国家发明专利2项，培养博士后1名，硕士研究生1名。. 本项目的研究结果，基本阐明野菠萝降血脂、降血糖作用的药效物质基础，发现具有新药开发价值的先导化合物和有效部位，并为黎药-野菠萝的开发利用提供科学依据，为降血脂、降血糖民族药物的研究提供思路和参考。