The Antimicrobial Activity Of Soy Whey Fermented By L.Plantarum D1501 And Its Use On Bean Curd As Preservation

It has been found that Lactobacillus plantarum D1501 isolated from Dong sour meat is a bacteriocin-producing bacteria,which can produce macromolecule bacteriocin when inoculated in MRS.In this study,L.plantarum D1501 was inoculated in soy whey to make fermented soy whey,whose neutral fermented supernatant had broad-spectrum antimicrobial activity.By optimizing the fermentation conditions,the yield of antimicrobial activity was maximized,and the antimicrobial substances was determined to be polypeptide bacteriocins in nature.Then bacteriocin was separated and purified by various separation methods.The molecular weight and amino acid sequence of bacteriocin were analyzed by mass spectrometry.The physicochemical properties of bacteriocin were determined and the fermented soy whey was used to extend the shelf-life of bean curd.It provides theoretical basis for using fermented soy whey to product new natural Lactobacillin,and provides a feasibility analysis for the application of new Lactobacillin in food preservation.The reuse of soy whey resources also provides a new idea for environment protection.The main research contents are given as follows:1?Optimization of fermentation conditionsAfter eliminating the influence of hydrogen peroxide,acid products and residual bacteria,the agar diffusion Oxford cup method was used to determine the antimicrobial activity of neutral fermentation supernatant produced by L.plantarum D1501.It could produce antimicrobial substances which could inhibit many Gram-positive and Gram-negative bacteria.Through single factor and orthogonal experiments,it was determined that L.plantarum D1501 was inoculated in soy whey liquid medium with initial pH 6.0,at 3%(v/v)inoculation,and incubated at 37? for 24 hours.The antimicrobial activity of L.plantarum D1501 in neutral broth reached the highest level.2?The antimicrobial characteristics of soy whey fermented by L.plantarum D1501 and the purification and identification of its bacteriocinThe antimicrobial spectrum of soy whey fermented by L.plantarum D1501 was tested under the optimum fermentation condition after freezedrying and concentration.It was shown that it had autoimmunity and a broad antimicrobial spectrum.It could not only inhibit lactic acid bacteria with close relationship,but also inhibit common Gram-positive and Gram-negative spoilage bacteria and pathogenic bacteria in food.The antimicrobial substances produced by L.plantarum D1501 was treated with various enzymes,which was determined to be polypeptide bacteriocins without carbohydrates and lipid groups in the structure.The bacteriocin was purified to homogeneity by ultrafiltration,RESOURCE-Q anion-exchange chromatography,reverse chromatography and secondary reverse chromatography,which was designted as Lp100.The final yield of bacteriocin was 1%and.the specific activity was 7247.00IU/mg,which was increased 79.94 fold.The molecular mass of purified bacteriocin was determined by LC-MS/MS to be approximately 1434.90Da,and its N-terminal sequence was MCCKVLLLLSRR.Through NCBI database comparison and literature search,no complete matching sequence was found,which proved that the bacteriocin was a new type of Lactobacillins that had not been reported.3?Physical-chemical properties and mode of action of bacteriocin LP100Bacteriocin LP100 was sensitive to protease,but lipase,amylase and catalase did not affect the antimicrobial activity.The antimicrobia activity of LP100 was stable in a wide pH range between 2 to 9,which showed strong antimicrobial activity under acidic and neutral conditions,decreased antimicrobial activity under alkaline conditions,and lost activity at pH>9.LP100 had good thermal stability and low temperature tolerance.It can retain 90.04%antimicrobial activity after exposuring to 121? for 15 minutes,and 97%antimicrobial activity after 2 hours of treatment at-20?.The antimicrobial activities did not change after UV radiations.Different chemicals had no effect on the antimicrobial activity of bacteriocin Lp100,but EDTA could significantly increase the antimicrobial activity to 137.7%.The action of bacteriocin on Staphylococcus aureus and Escherichia coli was bacteriostation instead of sterilization.4?Application of soy whey fermented by L.plantarum D1501 as preservation in bean curdAfter spray drying,the fermented soy whey solution with 5%,10%and 20%were obtained.Four strains of spoilage bacteria were isolated from spoilage bean curd,and their growth were inhibited by 10%fermented soy whey solution.The bean curd were immersed in the fermented soy whey solution for 10min,whose concentration were 5%,10%and 20%.Then Stored at 37? for 7 days.The effect of different concentration of fermented soy whey on the fresh-keeping effect of bean curd was determined daily by measuring the total number of colonies,the number of coliforms,Salmonella and Staphylococcus aureus and combining with sensory score.It was found that fermented soy whey with 10%concentration had the best preservation effect,and could extend the shelf-life for at least 5 days on the basis of maintaining the original quality of bean curd.