The Study Of Active Secondary Metabolites From Two Strains Of Fungi Under The Guidance Of Genome

Natural products derived from fungi have complex structures,novel skeletons,and diverse biological activities,making them the main source of small molecule drugs.The discovery of active natural products is to a large extent an accidental process,which hinders the rapid excavation of drug lead compound.In recent years,with the progress of genome sequencing technology,the cost of sequencing has been reduced,and the secondary metabolite gene cluster has been found rapidly,which has promoted the discovery process of natural products to a certain extent.Mining bioactive substances from fungi under the guidance of genomic information is of great significance for the development of new drug lead compound.The HPLC fingerprints of the endophytic fungus Aspergillus micronesiensis H39and the terrestrial endophytic fungus Collariella virescens 148.68 from sea cucumbers indicate their potential to catalyze the enrichment of secondary metabolites,and their potential bioactive molecules can be explored under the guidance of genomic information.First,two strains of fungi were cultured in small-scale fermentation.Modern chromatographic separation technologies such as silica gel column chromatography,gel column chromatography,semi preparative high-performance liquid chromatography and thin layer chromatography were used to separate and purify the secondary metabolites.Three new compounds and 24 known compounds were isolated,including butyrolactones,cytochalasins and chromones.The chemical structures of 27 compounds were identified using methods such as one-dimensional nuclear magnetic resonance(1D NMR),two-dimensional nuclear magnetic resonance(~1H-~1H COSY,HMQC,HMBC,and NOSEY),and mass spectrometry.Bioactivity testing and biosynthetic pathway speculation were conducted on 27 compounds.Genomic information display A micronesiensis H39 has 45 secondary metabolite biosynthesis gene cluster,including 12 type I polyketide synthase(T1PKS)gene cluster,1 type III polyketide synthase(T3PKS)gene cluster,10 non ribosome peptide synthase(NRPS)gene cluster,14 NRPS like gene cluster,1 T1PKS/NRPS like gene cluster,1 NRPS like/NRPS gene cluster,1 indole gene cluster 4 terpenes and 1betalactone gene cluster.From Eleven compounds were systematically isolated and identified from the crude extract of A.micronesiensis H39 fermentation.This includes one new compound,aspernolide M(1),and ten known compounds,aspernolide C(2),aspochalasin I(3),aspochalasin H(4),aspochalasin J(5),aspochalasin M(6),rosellichalasin(7),asperphenamate(8),ergosterol(9),epicocolide B(10),and epicocone(11).Through bioinformatics analysis,the NRPS like gene cluster located in scaffold 000002F is responsible for the biosynthesis of lipid compounds 1 and 2 in butene.Compounds 1-11 did not exhibit inhibitory activity against drug-resistant bacteria and fungi,and compounds 1 and 2 did not exhibit angiogenic activity in the zebrafish model.C.Virescens 148.68 secondary metabolite biosynthesis gene cluster has 26,including 12 T1PKS gene cluster,1 T3PKS gene cluster,7 NRPS gene cluster,2 NRPS like gene cluster,and 2 betalactone gene cluster.From 16 compounds were isolated from the crude fermentation extract of C.virescens 148.68,mainly natural products of chromones,including 2 new compounds chaetosemin F(1)and chaetosemin G(3),14known compounds chaetosemin E(2),aminochrome C(4),chaetoquatrin D(5),eugenitin(6),6-hydroxymethylaugenin(7),12.4 6-methoxymethylaugenin(8),chaetoquatrin G(9),chaetoquatrin H(10),sterigmatocystin(11),Chaetochromin A(12),3-methylglutaconic acid(13),3-[(1-carboxyvinyl)oxy]benzoic acid(14),p-hydroxybenzoic acid(15),and 5-hydroxy-2-hydroxymethyl-4H-pyran-4-one(16).Through bioinformatics analysis,the biosynthetic pathways of thiochromone compounds 3,4,and 5 were speculated.Compounds 7,9 and 12 showed weak to moderate inhibitory activity against common bacteria,especially compound 12 against Micrococcaceae with MIC value of 80μg/m L.