Studies On The Screening, Identification And The Metabolite Of The Strain Producing Bioactive Substance Against Fungi

In this paper, the agar column method combined with diffusion method of fermentation broth was applied, with the penicillium as a test microorganism, to screen the actinomycetes which can produce anti-fungi bioactive substance from 1, 436 different actinomycetes. At last, the Actinomycetes-1356 was obtained as our objective strain. Antibiogramic research showed that the strain 1356 had strong activity against Fusarium vasin fectum and Baker's yeast et al, and had activity against other pathogenic fungi to some extent, such as Cladosporium fulvum, but had no inhibition activity against bacteria.Strain 1356 was identified to be Aureus group of genus Streptomyces through the classical and molecular-biological methods. In the phylogenetic tree strain 1356 was most closely related to Streptomyces griseochromogenes subsp, suitaensis NBRC 13776, sharing 16SrDNA similarity value of 99.9%. However, the morphological, cultural and physiol-biochemical characteristics between the two strains revealed obvious difference. Based on the polyphase taxonomical data, strain 1356 was primarily identified as Streptomyces griseochromogenes subsp, suitaensis, n. Var.For improving the productivity of bioactive substance by strain 1356, the fermentation conditions were studied. Antibiotic content of the fermentation broth was determined by cylinde-plate method, the suitable fermentation conditions were as follows: seed age, 32h; inoculation volume, 10.0(v/v); loading liquid volume, 50 ml (in 250ml flask); fermentation period, 4 d.Bioactive substance was obtained from extracts of fermentation supernatant and from extracts of mycelium of the strain 1356. We also conducted the preliminary study on the physico-chemical property of the bioactive products: the stability inspection, paper chromatograph and pH chromatograph experiments. These resules showed the basic physico-chemical property of the bioactive products and founded the preliminary techniques of separation.The method of isolating active substance from fermentation supernatant by macroporous adsorbent resin was studied. DC2 resin was selected as the best adsorbent, and the desorption conditions were optimized: water was used for washing-out polar impurities, 20% ethanol was used for removing some other impurities and pigment, and finally 80% acetone was used for eluting active substance. Furthermore, 11 kinds of organic solvent were investigated to extract the bioactive substance from mycelium, the best extractant was acetone.