Separation, Purification, And Biological Activity Of Antifungal Antibiotics In A Preliminary Study

So far,80%of antibiotics which have been found are from actinomycetes, actinomycetes mainly exist in soil and account for great proportion.The strain FL₁ used in this paper was supplied by the cell engineering group of IEA,who firstly screened 20 active strains from 2000 different actinomycetes,and then we screened again and got one high efficiency strain named FL₁.With the Bipolaris maydis as a test microorganism,applied the agar solid fermentation method combined with shaking flask liquid fermentation method,we investigated the bioactivity of the broth and found that the broth which was produced by FL₁ had good bacteriostasis effect to Fusarium.moniliforme,Sphacelotheca reiliana,Exserohilμm turcicμm,Botrytis cinerea,Fusariμm oxysporum f.sp.cucμmerinμm, Alternaria longipes Tobacco pathotype,Colletotrichum orbicμlare,alternaria leaf spot of egg plant,so it belongs to broad-spectrum antibiotic.Besides,in the experiment of seed soaking of corn,wheat and cucumber,we also found that the broth had good effect to the growth promotion of the crops.For improving the productivity of bioactive substance of strain FL₁,both the fermentation medium and the fermentation conditions were optimized.Determining the bacteriostasis effect to Fusariμm.moniliforme in different conditions,the suitable fermentation conditions were as follows:cultivation temperature,28℃;loading liquid volume,70mL/250mL;fermentation period,4d.Strain FL₁ was identified to be the cinerogriseus group of actinomycetes through the classical and molecular biology categorization method.Blasted in NCBI,Strain FL₁ was most closely related to Streptomyces albogriseus,sharing 16S rDNA similarity value of 100%.However,the morphological,cultural and phy-biochemical characteristics between the two strains revealed some extent difference.Based on the polyphase taxonomica data, Strain FL₁ was primarily identified as Streptomyces albogriseus.n.Var.Bioactive substance was obtained from extracts of fermentation supernatant.We also conducted the preliminary study on the phy-chemical properties of the bioactive products,basing on that,we did the stability experiment,paper chromatography and thin layer chromatography,provided enough evidences for further isolation and purification. The result of paper chromatography showed that the antibiotic may belong to aminoglycosides,or multi-peptid,but there were big differences between them.We applied silica gel column chromatography to separate bioactive substance, bioactive substance was eluted under the condition of mobile phase(CH₃OH:CHCl₃ =1:5).Through The whole ultraviolet wavelength scanning,it made sure that the maximum absorption wavelength was 277nm.Chromatographic condition:detection wavelength:277nm,mobile phase:acetonitrile:KH₂PO₄ buffer(0.02mol/L)=40:60,flow rate:1.5mL/min.when the retention time came to 12.723,chromatographic peak came out, which had great resolving power and good peak form.By means of ESI-MS,the molecular weight(MW)of active substance was conceived as 495.