Screening And Identification Of A Strain Producing Bacterial Cellolose From Kombuchaand Purification Of Fermentation Process

Bacterial cellulose is a kind of secondary metabolites produced by microorganisms. It is a kind of extracellular polysaccharide which is linked with ?-1,4 glycosides. Compared with lignocellulose, it is the key focus of the current research in society, with a complex three dimensional reticular structure,material-combining ability, good biocompatibility, high purity, outstanding water-holding capacity, which can play an important role in many areas. In this dissertation, we mainly study the producing strain of bacterial cellulose which is isolated from Kombucha, and it is identified as Gluconacetobacter sp., and it carried out preliminary optimization for the fermentation process of Gluconacetobacter sp. producing bacterial cellulose.Firstly the bacterial cellulose producing strains is separated from Kombucha which provided from the laboratory, then carried out an genetic stability experiment on the separated strains, screened the strain with high production and genetic stability, the strain is identified as Gluconacetobacter sp. through the physiological and biochemical characteristics and 16 S rDNA molecular. Scanned the biofilm by electron microscopy and infrared spectroscopy, determined the product as the bacterial cellulose, and operated purity test on the bacterial cellulose produced by laboratory, which purity can reach 98.02%.The fermentation conditions of Gluconacetobacter sp. are optimized. We plotted the growth curve that Gluconacetobacter sp. is in seed culture medium. Fermentation medium carried out inoculation during which it selected bacteria of 30 h logarithmic phase. This experiment examines the effect of different carbon source, nitrogen source, organic acid, phosphorus and salt on the yield of bacterial cellulose.Through this experiment and find out the changes affect the yield of bacterial cellulose is relatively obvious three factors, they are: acetic acid, yeast extract powder, potassium dihydrogen phosphate. We designed three-factors three-levels response surface experiments(RSM) and analyzed response value based on bacterial cellulose yield, and built two dimensional model between the three-factors and bacterial cellulose production, analyzed and predicted extreme value of the quadratic equation by SAS analysis software. It concluded that bacterial cellulose production is the largest when yeast extract is 3.08%, acetic acid 0.2%(v/v), potassium dihydrogen phosphate 0.12%. glucose 0.8%, sucrose 1.5%. After the detection theproduction of bacterial cellulose is about 5.64 g/L which are basically consistent with the response surface prediction values.We optimized parts of the fermentation conditions, carried out single factor analysis on temperature, initial pH, inoculation amount, selected three significantly influenced level, designed three-factors three-levels orthogonal experiment, examined effects on bacterial cellulose production when different temperatures, initial pH, inoculation amount interacted, operated the range analysis according to the experimental results. We can get a conclusion that when it comes to the effect of bacterial cellulose,inoculation amount is greater than the initial temperature and meanwhile is greater than pH, and the optimum fermentation conditions is 30?, inoculation amount is 2.0%, initial pH is 6.0. Above all, when Gluconacetobacter sp. at 30?, inoculation amount 2.0%, initial pH6.0, static culture for 7 d, the production reached 5.73 g/L, which increased 10 times before optimization.