Studies On The Secondary Metabolites From Eight Soil Actinomycetes Strains

Actinobacteria are important sources of bioactive natural products,especially antibiotic.Among those antibiotic,ansamycins have special structures and biological activities.The aim of our research is to discover new ansamycins from soil actinomycetes.Traditional methods for natural products screening from fermentation broths are time-consuming.inefficient and aimless.Based on the necessity of the AHBA synthase in the biosynthetic pathway of ansamycins.as well as the high conservation of AHBA synthase genes in different actinomycetes,we obtained thirty-three AHBA synthase genes positive strains via soil sampling,actinomycete isolation and PCR screening.After twice strain screenings,and in view of the yield of strains and genetic evolutionary tree,eight strains with the potent of producing ansamycins were selected for further research.We obtained different types of ansamycins and other compounds from their secondary metabolites.Meanwhile,the biological activities of the new ansamycins were evaluated.Seven compouds were isolated from the oatmeal medium of Streptomyces sp.S011,S013,S014 and S027,including one octaketide-type ansamycin（17-O-demthylgeldanamycin）.two nonaketide-type ansamycins（hygrocin C and hygrocin D）and one new compound belonging to naphthol.Nineteen compouds were isolated from the oatmeal medium of Streptomyces sp.S015,S045 and Kitasatospora sp.S023,vincluding two new compounds.One was furanone derivative and the other was quinoline derivative.No ansamycins were isolated.We deduced that ansamycin gene cluster was in low expression or not expressed.Twenty-two compouds were isolated from the oatmeal medium of Streplomyces sp.S012,including twelve new compounds.The new compounds included three precursors of ansamycins and nine rich-structured streptovaricins,ansavaricins A-I（34-38 and 42-45）.which belonged to undecaketide-type ansamycins along with rifamycins.Part of the new streptovaricins（34,35 and 42-45）that ansa chain disconnected with C-5 were open chain ansamycins which were reported once before.Moreover,we evaluated the biological activities of the new ansamycins and found that new compound 38 could inhibit the secretion of SPI-1 effectors（SipA/B/C/D）and had no influence on the growth of Salmonella Typhimurium strain,which reduced risks of drug resistance.Ansavaricin H（44）was confirmed to potently inhibit both Topo Ⅰ and Topo Ⅱα.What’s more,it was confirmed that streptovaricins with acyclic ansa chains（42-45）were more effective in topoisomerases inhibitory activities than the cyclized compounds.The results proved that Ansavaricin H（44）was more cytotoxic against HeLa and MDA-MB-453 cells than other tested cell lines(IC₅₀=50μM).Western Blot and Immunofluorescence experiments proved that ansavaricin H（44）appeared to induce DNA damage and apoptosis in HeLa cells.