Isolation,Purification And Structure Eculidation Of Novel Lipopeptides From Streptomyces Griseus Subsp. Griseus HYS31

The strain HYS31 was isolated from soil.The methanol extract of HYS31 biomass exhibited antifungal activity against Alternaria longipes.Therefore,the secondary metabolites as agricultural antibiotics were studied by bio-guided isolation used Alternaria longipes as indicator.The antifungal metabolites in HYS31 fermentation broth were isolated and purified by methanol extraction,silica gel column chromatography and Sephadex LH-20 column chromatography.Two monomeric compounds with agriculturally antifungal activity,A3-1 and A3-2,were prepared by semi-preparative HPLC.The chemical structure was identified by HR-ESI-MS,amino acid analysis,1H-NMR,13C-NMR,2D-NMR,single-crystal x-ray diffraction and so on.The results of structural analysis showed that both two target compounds are lipopeptides.These two compounds are homologs with molecular weights of 1468 and 1482,respectively.Their molecular formulas were C73H92N14O19 and C74H94N14O19,respectively.These two compounds contained some unusual amino acids including hydroxyglycine,3-hydroxyleucine,3-phenylserine,?-hydroxy-O-methyltyrosine,which determined the novelty of their molecular structures.The HPLC analysis method was established to quantify the contents of two compounds in fermentation and purification processes.The reliability of analysis method was verified by statistical calculation.The results of antimicrobial experiments in vitro showed that the two compounds performed similar antifungal activity against plant pathogenic fungi,but the bioactivity was not significant,and their minimum inhibitory concentration(MIC)against Alternaria longipes was 250 mg/L.The strain HYS31 was identified by its 16S rDNA gene sequence,combined with its physiological and biochemical characteristics on different media.The results showed that the strain HYS31 was Streptomyces griseus subsp.griseus classified into Streptomyces family.