白刺链霉菌(Streptomyces albospinus)CT205菌株发酵条件优化及其次生代谢产物性质研究

The optimization of fermentation conditions of Streptomyces albospinus CT205 and study of secondary metabolite

[目的]为了获得白刺链霉菌（Streptomyces albospinus）CT205菌株摇瓶发酵最佳配方和适宜的发酵条件,并初步研究其次生代谢产物的性质。[方法]通过碳、氮源单因子试验、正交试验和主要发酵条件优化试验,以菌浓度和抑菌圈大小为指标确定其最佳发酵培养基配方和发酵条件。通过GF254薄板层析、生物测定、紫外检测、稳定性试验,研究菌株CT205发酵产生的次生代谢产物性质。[结果]菌株CT205发酵最佳培养基配方：蔗糖20.0g·L^-1,可溶性淀粉30.0g·L^-1,蛋白胨2.0g·L^-1,黄豆粉8.0g·L^-1,MgSO4·7H2O0.5g·L^-1,K2HPO4·7H2O0.5g·L^-1,NaCl2.0g·L^-1,CaCO33.0g·L^-1;最佳发酵温度为28～30℃,初始pH8.0;CT205发酵产生的抗菌活性成分的Rf值为0.68,其在λ=258nm处有1个强紫外吸收峰。在不同温度（60、80和100℃）、紫外光照射不同时间（5、10和30min）处理下,活性成分的抗菌活性与对照相比变化不大,在用酸碱处理后,其抗菌活性比对照略有下降。[结论]获得了菌株CT205优化的发酵培养基配方及适宜的发酵条件,其次生代谢产物对热和紫外光均表现出较强的稳定性,但偏酸、偏碱的环境对其活性有一定的影响。

[Objective]The aim of this study was to obtain the optimal medium and conditions for the fermentation of Streptomyces albospinus CT205,and to investigate the properties of its secondary metabolites.[Methods]A single factor at a time experiment,orthogonal experiment and optimizing condition experiment were used to determine the optimal ingredients of fermentation medium and suitable fermentation conditions in shake flask with mycelia quantity and inhibition zone as indicators.The thin layer chromatography（TLC,GF254）,ultraviolet detection and bio-monitoring were used to study the properties of antibacterial substances produced by strain CT205.[Results]The sucrose and starch were the optimal carbon sources,and peptone and soybean meal were the optimal nitrogen sources obtained by using the single factor at a time test.The optimal fermentation medium conditions for strain CT205 were obtained by orthogonal experiment.The optimal medium was sucrose 20.0g·L^-1,starch 30.0g·L^-1,peptone 2.0g·L^-1,soybean meal 8.0g·L^-1,Mg SO4·7H2O 0.5g·L^-1,K2HPO4·7H2O 0.5g·L^-1,NaCl 2.0g·L^-1,Ca CO33.0g·L^-1; suitable fermentation temperature was 28-30 ℃,initial pH was 8.0 and fermentation period was 96 h.It was found that the Rf value of the active substance was 0.68 on the thin layer GF254 in the chromatography system of trichloromethane,ethyl acetate,methyl alcohol and ethyl acetate（The ratio of volume was 16∶ 1∶ 2∶ 0.04）.The active substance was scanned by ultraviolet from 200-400 nm wavelength,and a strong UV absorption peak was found at wavelength λ = 258 nm.Little change was observed in size of inhibition zone of the active substance under the treatment of different temperatures（60,80 and 100 ℃）,UV irradiation at different time（5,10 and 30 min） compared with the control,but the size of inhibition zone of the active substance treated with acid or alkali was smaller than that of the control.[Conclusions]The results of stability test showed that the active substance from strain CT205 had a stronger stability to high temperature and ultraviolet,but had a certain extent impact at the partial acidic and partial alkaline conditions.