The Isolation, Identification Of Bacillus Cereus Producing Bacteriocin And The Optimization Of Medium

Due to the using of chemical preservatives in food industry caused more and more security issues, people have begun to look for other preservatives which are provided with security and efficiently. In recent years, natural preservatives including plant source, animal origin and microbial source have become a focus issues in the food industry. The natural preservatives of microbial source have especially attracted increasing attention which bacteriocin was the research priorities. Bacteriocins which produced by bacteria is a kind of antibacterial substances that possess many advantages including efficientl, innocuity, nonhazardous effect, without residue and none drug-resistance. Therefore, the aim of this project was to screen strains of producing bacteriocin, study the biological features of bacteriocin and optimize the production of bacteriocin substance using response surface methodology.In this paper, the bacteriocin-producing strain which named XH25was screened from soil by the agar well diffusion method using Stapoylococcus aureus as the indicator. The supernatant of XH25strain can inhibit indicator strain excluding inhibition effect of organic acid, H2O2and the sensitive test of proteinase K. The results confirned that this inhibition substance was a kind of protein, could be classed as bacteriocin. The physiological and biochemical features of XH25strain were gram-positive, catalase reaction, starch hydrolysis, nitrate reaction, esterase reaction, M.R reaction, V.P reaction, lecithin enzyme hydrolysis, casein hydrolysis and sugar alcohol fermentation were positive except H2S reaction. The strain XH25was identified as Bacillus cereus by molecular identification of16S rRNA sequence analysis which combined with physiological and biochemical tests.Response surface methodology was employed to optimize medium composition for the production of bacteriocin substance by Bacillus cereus XH25. The Plackett–Burmen design was used to evaluate the effects of different components in the culture medium. The initial pH value of medium, soluble starch and (NH4)2SO4were found to significantly influence on the bacteriocin substance production. The steepest ascent method was undertaken to access the optimal region of the above three significant factors, and then their concentrations were optimized by the central composite design and the response surface methodology. The results indicated that the quadratic term of the three factors and the interactions between soluble starch and initial pH value,(NH4)2SO4and initial pH value had a significant effect on the production of bacteriocin substance. The optimal medium composition to achieve the optimal production was soluble starch15.32g/L,(NH4)2SO46.02g/L and initial pH value6.15, which tamped a21.7%increase of antibacterial activity against Staphylococcus aureus when compared with the original medium. The predicted value12.76mm was verified subsequently by validation experiments value12.90mm. The excellent correlation between predicted and observed values further confirmed the adequacy and accuracy of the model obtained. The optimal culture medium obtained in current work provided a basis for further study on the food preservation of bacteriocin substance by Bacillus cereus XH25.