卤盐胁迫盐生药用植物内生真菌的群体感应抑制活性产物研究

Searching for efficient quorum sensing inhibitors (QSIs) is a new way to solve the problem of drug resistance in bacterial infection. Because of the ecological specificity of the medicinal plants in the coastal mudflat, the metabolic pathway of endophytic fungus is not only different from that of the common plant , but also that of the general salt tolerant plant. Under the coastal mudflat circumstances, the endophytic fungus may produce new compounds of potential bacterial quorum sensing inhibitory activity. Previous studies have shown that halogen salt stress can enrich the structure of secondary metabolites of salt tolerant fungi and enhance the antibacterial activity. Based on this, in order to study the secondary metabolites of fungi, 3 medicinal plants on Yangkou-Beach of Qingdao are selected as the research objects, 7 kinds of halogen salt stress are chosen to enrich the metabolites, QS inhibition activity test of Chromobacterium violaceum and Pseudomonas aeruginosa are taken as the test index of activity. During our early study, 25 endophytic fungi have been obtained from one plant, and two “talented strains” with abundant metabolites and good activity of QS inhibiting have been obtained. In order to get strong activity QSIs of new structure, we will study the endophytic fungi and the metabolites of the other two plants in the following work, so as to obtain lead compounds for resisting durg-resistant bacteria. Not only does this study have important practical significance to solve the problem of bacterial resistance, but also provide the basic data for the protection and development of biological resources in the coastal beach.

寻找高效群体感应抑制剂（QSIs）是解决细菌感染耐药性问题的新途径。沿海滩涂抗菌药用植物生态的特异性，使其内生耐盐真菌形成既不同于普通植物内生真菌，又迥异于一般耐盐真菌的代谢途径，是潜在的细菌QS抑制化合物产生者。前期研究表明卤盐胁迫可丰富耐盐真菌次生代谢产物结构类型，增强抑菌活性。基于此，本研究以青岛仰口沿海滩涂3种抗菌药用植物内生真菌为研究对象，7种卤盐胁迫为丰富代谢产物手段，以紫色杆菌和铜绿假单胞菌的QS抑制为活性指标，对真菌次生代谢产物进行研究。本研究已从其中一株植物获得25株内生真菌，并筛选得到两株代谢产物丰富，抑制QS活性良好的“Talented菌株”。接下来，拟对另两株植物内生真菌及代谢产物进行研究，以期得到结构新颖的强活性QSIs，为寻找抗耐药细菌药物提供先导化合物。本研究不仅对解决细菌耐药性的问题有重要现实意义，而且能为沿海滩涂生态系统中生物资源的保护与开发提供基础资料。

沿海滩涂药用植物来源真菌是研究群体感应抑制剂分子的重要来源。本研究从4株耐盐植物中华补血草、野生大豆、蔓荆、罗布麻及1株陆地植物桑树的根、茎、叶、果实获得内生真菌和根际土共附生真菌167株。对这些真菌资源进行了保藏和HPLC指纹图谱库构建，为后续开展药用先导化合物挖掘相关工作提供了资源微生物。.本研究从筛选得到的6株真菌，青霉菌Penicillium sp. L129、哈茨木霉Trichoderma harzianum L130、间座壳属菌Diaporthe goulteri L17、黄柄曲霉Aspergillus flavipes L36、变红镰刀菌Fusarium incarnatum L107、支顶孢属菌Acremonium citrinum SS-g13的次生代谢产物中分离鉴定单体化合物64个，其中新化合物18个，新天然产物1个。对纯化得到的化合物进行活性测试。实验结果获得强细胞毒活性新化合物2个，QS群体感应抑制活性化合物3个，抗农业致病菌活性化合物5个。本研究为开发抗细菌、抗肿瘤、抗农业致病菌活性先导化合物提供了侯选化合物和模式结构。.在研究化合物结构的过程中，基于1D和2D核磁共振波谱、旋光、圆二色谱（CD）、计算ECD、X-Ray单晶衍射（铜靶）精确阐明了大部分新化合物的立体绝对构型。利用化学衍生手段（钼试剂法）确定含有仲醇碳的手性构型。另外，本研究对6株海洋来源真菌（L36、L6、L115、L130、L46、XY）进行OSMAC策略研究。研究发现发酵培养基中碳氮源，或卤盐的变化可改变耐盐真菌次生代谢产物的种类和活性的变化。研究结果整理发表天然产物药物化学相关 SCI 论文 7 篇，中文核心论文1篇，授权第一发明人专利1项。