Optimization Of Fermentation Process Parameter In Shaking Flask On And Antibiotics Purification Of Streptomyces Sp. LA5 Strain

The Streptomyces sp. LA5 strain was isolated from the capsicum plant in Hainan Province, China, and strongly inhibitory to Phytophthora capsici, Fusarium oxysporum f. sp. cubense, Collectotrichum musae, etc. LA5's fermentation liquid of carrot gave good control effects against capsicum blight, banana Panama disease and banana anthracnose. The antifungal active component is the antibiotics produced by LA5 strain, but its production is low in LA5 fermentation liquid of carrot. In order to understand the antibiotic LA5 better, we optimized the medium and process parameter in shaking flask for LA5 fermentation, and purified the antifungal active component from LA5 fermentation liquid.A total of 11 factors were tested their function of improving antibiotics yield of LA5 by single-factor test methods. The results showed that glucose is the best carbon source for the antibiotics productivity of LA5, other are sucrose, maltose, corn flour; bean flour is the best nitrogen sources, others are peptone, yeast flour, yeast extract, beef extract; divalent ions as Ca2+, Mg2+ stimulated LA5 antibiotic biosynthesis. Hereby, an uniform design test was applied to optimize above 4 factors. The follow regression model equation and optimized medium were obtained according to regression analysis of the data from uniform design experiment: Y=-5.7512 + 0.5515X1 +0.7811X2 +26.6954X3 +26.8706X4 -0.0066X12 - 0.0115X22 -24.0747X32 -22.5914X42 -0.01302X1X2 -0.3516X2X4 -21.4559X3X4. The optimum culture medium includes bean flour 28.00 g/L, glucose 13.78 g/L, CaCO3 0.42 g/L, MgSO4 0.28 g/L. The model was validated experimentally by the value of 16.10mm, and the deviation was 4.62% compared with theoretical value. The results also indicated that the optimum process parameter in shaking flask on fermentation consist of initial pH of 6.8, 28℃of culture, 5% of inoculation amount, medium loadage 100ml/300ml, 200 r/min of rotation, and 120h of fermentation time.The purification procedure of the fatsoluble antibiotic LA5 was elucidated by research on fermentation liquid pretreatment, rotary evaporator concentrating, organic solvent extracting, reversed phase column isolating, silica gel column chromatography isolating, TLC test etc. The crude antibiotic LA5 was well purified and six pure component were gained by this purification procedure.