Study On The Secondary Metabolites Of Insect Endophytic Fungus Aspergillus Lentulus

Endophytic fungi are an important class of microorganisms that live in the host plants and animals.As time goes on,endophytic fungi with their hosts have established an inseparable relationship.The secondary metabolites of endophytic fungi and their hosts have significantly promoted further research on their microorganisms.Their secondary metabolites are expected to become an important source of different biologically active compounds due to huge development potential.An endophytic fungus isolated from insect intestines was identified as Aspergillus lentulus by ITS sequence.After confirming the species of the endophytic fungus and consulting the related literature,nine kinds of culture media were selected for fermentation.Then the rice medium was selected to be the most suitable fermentation condition by high performance liquid chromatography analysis.The strain was cultured at 28℃ for 30 days.After the fermentation,the metabolites were extracted with ethyl acetate and a total of 72 g crude extract was obtained.Macroporous adsorption resin,TLC chromatography,silica gel column chromatography,ODS chromatography,HPLC and Pre-HPLC were applied to separate the crude extract.Finally a total of 18 compounds were obtained.The planar structure of 18 compounds was identified with the application of NMR,MS,UV and IR techniques to analyze the physical and chemical properties of the separated compounds.The relative configurations of the new compound were determined by ROESY spectrum and the absolute configuration was confirmed by ECD calculation and ECD induced by Rh2(OCOCF3)4.Finally,18 compounds were identified including three prenylated indole alkaloids(compounds C1-C3),five other alkaloids(compounds C4,C6-C9),and five polyketone dimer(C11-C15)and five other types of compounds(C5,C10,C16-C18).C1 has a 6/5/6/5/5 pentacyclic ring system structure and C2 has a 6/5/6/7 tetracyclic new skeleton structure.In addition,the anti-tumor activity of these compounds was measured by the MTT method and the anti-bacterial activity against agricultural pathogenic bacteria was measured by the micro-dilution method.C2,C4,C5,C13 and C15 showed moderate anti-tumor activity against A549,and their IC50 values were in the range of 17.92-48.29 μM.During the experiment to evaluate the antibacterial activity,compounds C1,C2 and C13-C15 showed obvious anti-bacterial activity against agricultural pathogenic bacteria Xanthomonas oryzae pv.Oryzae and the MIC value was ranging between 25-100 μg/ml.In summary,this study explored mainly about the fermentation metabolites of the endophytic fungus A.lentulus and three new alkaloid compounds and five polyketone dimers were isolated from the metabolites,two of which were the alkenylated indole alkaloid with a new 6/5/6/5/5 and 6/5/6/7 ring system structure respectively.Finally the anti-tumor activity and anti-bacterial activity of the isolated compounds are determined,which provides the basis for finding lead compounds.