The Screening Of High-yielding Strains Of Natamycin

Natamycin is one kind of broad-spectrum macrolide antibiotic antifungal which is used to inhibit mold, yeast growth and other fungistatic and fungicid growth. It can be produced by Streptomycesna talensis, Streptomyces chatanoogensis and Streptomyces gilvosporeus and so on. Compared with other antiseptic reagent, the usage of natamycin is only 10-6 of magnitude and can be applied to a wide range pH values. Natamycin is known as an efficient, safe biological preservatives, which is non-toxic, and not mutations, non-carcinogenic, non-teratogenic, non-allergenic, and does not affect the flavor of product. In this experiment, Streptomyces gilvosporeus, ATCC 13326 was used as the original strain which′s genetic background is relatively clear and output is low and stable. In this study the following methods were taken to conduct a preliminary study of its.The results are as follows:The method of high-temperature domestication was applied to improvement of the strain of Streptomyces gilvosporeus,ATCC13326.After several times of temperature gradient domestication, HS-5 strain was obtained which can adapt to a condition of 36℃. And its spores grow fast with a state of repletion. Isolated plates were used to culture HS-5 which was screened out by high-temperature domestication. The result showed that morphological characteristics of the colonies significantly changed compared with the original strain. The growth curve of seed and kinetics of natamycin fermention were studied with the strain of HS-5.The results were that seed age was ahead of 6h and both stationary phase of bacterial growth and synthesis phase of natamycin extended compared with the original strain. The yield of natamycin production was increased by 41.3 %.HS-5 strain which was screened by high-temperature domesticated was treated with DES in series, a mutant strain DS-C-13 with high natamycin productivity was screened out by using the medium with selective pressure of erythromycin and genetic stability of the mutation strains. Its production reached 219.42±2.82 mg/L. The culture conditions and the fermentation medium for natamycin production were optimized with strain DS-C-13. The culture conditions and the fermentation process were optimized. The effects of cultivation conditions and fermentation process such as inoculum size, medium volume, carbon and nitrogen sources, on namycin production were evaluated. Under the optimized conditions, which was inoculum size 4 %, medium volume 30 mL/250 mL, Glucose 50 g/L, Soya Peptone 20 g/L, Yeast Extract 5 g/L, natamycin production reached 256.32 mg/L which was higher 16.82% than that of initial conditions. To further enhance natamycin production of the strain, based on a single factor experiments, Response Surface Method was used to optimize natamycin fermentation. According to Box-Benhnken of the central composite experimental design principles,carbon source glucose, medium volume and inoculum size which have significant influence on natamycin fermentation were selected to design experiments of three factors and three levels. The optimized cultivation conditions were as follows: inoculation size 2.7%, medium volume 29 mL/250 mL, Glucose 44.63 g/L, Soya Peptone 20 g/L, Yeast Extract 5 g/L. Verification experiment was conducted under the theoretical optimum fermentation conditions, the result showed that the yiled of natamycin reached 256.32 mg / L with the theoretical value of the difference of 1.48%. Natamycin production was increased by 27.17% with this method.