Screening Of Broad-spectrum Bacteriocin Producing Lactobacillus From Fermented Cucumber And The Bacteriocin

Food security has become an increasing concern with the improvement of living standards and the increase of health awareness.People start looking for safe natural Preservatives and antibacterial agents.Bacteriocins were metabolited products of food-grade microorganisms,there has been a recent explosion in the interest of bacteriocins as natural biopreservatives in food systems and they have been considered to be the most effective alternative to antibioties.This paper main researched the isolation and identification of broad-spectrum bacteriocin producing lactobacillus,the purification,antimicrobial spectrum and biological stability of the bacteriocin.The results offered a theoretical base for the development and ultilization of bacteriocin as a kind of natural food preservative of antibacterial bacteriocin.1.A new board-spectrum bacteriocin producing strain LG34 was screened from fermentation cucumber by agar diffusion assay method with Staphylococcus aureus and Bacillus coliindicator as indicator.Using observation of morphological characteristics,physiological and biochemical tests and combining with the molecular method of 16 S rDNA sequence analysis,the strain was confirmed to be Lactococcus garvieae.2.The bacteriocin produced by LG34 was designated as garviecin LG34.The purification and the molecular weight of garviecin LG34 was researched.The results showed that the optimum conditions of Sephadex G50 gel column chromatography was determined as follows: elution speed 1.0 mL/min,sample volume 3 mL and concentration 0.3 g/mL,at these conditions the bacteriocin activity reached 592 AU/mL;Then purified by SP Sepharose Fast Flow cation exchange chromatography,the optimum conditions was determined as follows: equilibrium buffer pH 3.0,ionic strength of the eluate was 1.0 mol/L,and the bacteriocin activity reached 2206 AU/mL.The molecular weight of the bacteriocin determined by Tris-Tricine SDS-PAGE was 5.4kDa.3.Research on the antibacterial spectrum and the stability of garviecin LG34.The antimicrobial spectrum assays showed that garviecin LG34 could inhibit the growth of both Gram-negative bacteria and Gram-positive bacteria,but does not inhibit mold and yeast;test for the stability of garviecin LG34 showed that it has a strong thermal stability,the antibacterial activity only have 10.5% loss suffered under the temperature of 121℃ for 30min;it was sensitive to pepsin and trypsin;the antibacterial activity is quite stable on the condition of pH 2.0~8.0,and the loss of antibacterial activity is less than 20%;inorganic salts have a significant effect on antibacterial activity(P<0.01),with the increase in the concentration of inorganic salts,the loss of antibacterial activity increased gradually;organic solvents and surfactants in the added amount of 10% ~50% didn’t have significant effect on the antibacterial activity(P>0.05).