基于超高效液相色谱-电喷雾-质谱、天然产物词典和活性筛选的艾纳香内生真菌次生代谢产物

Screening of bioactive secondary metabolites of endophytes associated with Blumea balsamifera(L.) DC. by UPLC-QTof-MS, DNP and bio-assay

【背景】植物内生真菌是天然活性小分子的重要来源,但由于种类繁多,导致寻找结构新颖、活性强的化合物非常困难,重复分离已成为制约新型药源小分子发掘的瓶颈。【目的】综合各种技术,快速寻找目标活性次生代谢产物。【方法】通过对菌株分子鉴定、天然产物词典(dictionary of natural products,DNP)数据库检索、超高效液相色谱-电喷雾-质谱(ultra performance liquid chromatographyquadrupole time-of-flight mass spectrometry,UPLC-QTof-MS)分析、滤纸片法抑菌实验和色谱技术跟踪获得活性单体化合物。运用质谱和单晶衍射技术对化合物结构进行鉴定,96孔板法对单体化合物进行活性评价。【结果】分离鉴定出11株艾纳香内生真菌,筛选出一株各方面表现良好的艾纳香内生菌菌株Diaporthe sp.,从其大米培养基中获得一个单体化合物Cytochalasin H,活性评价显示其对枯草芽孢杆菌具有很好的抑制活性,MIC值为32μg/mL。【结论】将多种筛选技术相结合的方法应用于艾纳香内生真菌活性代谢产物的发掘,为快速寻找活性先导化合物提供了很好的借鉴。

[Background]Endophytic fungi is a reservoir of bioactive small-molecular natural compounds,but it is difficult to obtain structurally novel compounds and pronouncedly active analogues because of enormous species dispersed in nature,and repeated isolation became an bottleneck in the mining process.[Objective]The objective is to rapidly find new biologically active metabolites with combination of LC-MS,DNP and bioactivity evaluation.[Methods]Pure metabolite was isolated through silica gel and recrystallization.The structure was elucidated by spectroscopic and X-Ray techniques.The biological activity was evaluated through 96-well dilution method.[Results]Eleven endophytes were isolated and identified from B.balsamifera(L.)DC.A targeted endophyte identified as Diaporthe sp.was selected and an active compound Cytochalasin H was afforded from the rice extract of this fungus,its inhibitory effects against Bacillus subtilis was assayed with a MIC value of 32μg/mL.[Conclusion]This study introduced a new extensive method in mining for bioactive secondary metabolites from endophytes of B.balsamifera(L.)DC.,which provides references for rapidly exploration of effective lead agents.