Study On The Preparation,Characterization And Properties Of Biosurfactants Produced By Lactic Acid Bacteria

Biosurfactants are amphiphilic compounds produced by microorganisms that have both hydrophilic and hydrophobic groups.Compared with traditional chemically synthesized surfactants,in addition to the same functions as reducing surface tension,stabilizing emulsions and increasing foam,it also has the advantages of diversified structure,wide selectivity,strong specificity,good biocompatibility,non-toxic or low toxicity,green environmental protection,simple production process and so on.It has become a research hotspot.In this study,69 strains of lactic acid bacteria from the research group were used as the selecting source.The oil-draining circle method and the ring method were used to measure surface tension to screen out the strains with strong surfactant production and identify the strains;the surfactants were separated and purified by high performance liquid phase,and its structure is characterized and analyzed by mass spectrometry(MS),Fourier transform infrared spectroscopy(FTIR),X-ray photoelectron spectroscopy(XPS),and the emulsification and pathogenic bacteria(The antibacterial properties of Candida albicans,Escherichia coli,Enterococcus faecalis,Staphylococcus aureus,Pseudomonas aeruginosa)were tested,and the fermentation performance of the bacteria slurry after the surfactant extraction was compared with the unextracted fermentation performance.Get the following conclusions:1.The production of surface active substances by 69 strains of lactic acid bacteria varies from bacteria to bacteria.The output of extracellular surfactants is concentrated in the range of 0?300 mg/L,and the output of cell surfactants is concentrated in the range of 100?400 mg/L.Strains 69,B25,B82,S73,S75,and S79 were screened out for gas production.The surface tension was measured by the ring method and re-screened.Finally,the strain S73(extracellular surface tension:48.23 mN/m,cell surface tension:46.93 mN/m)was obtained.The strain was identified as Streptococcus thermophilus by 16S rDNA.2.Through single factor,Plackett-Burman test,response surface test and central combination test,the medium for surface active substance production of Streptococcus thermophilus S73 was optimized,and the best medium for surface active substance production was yeast extract powder 0.50 g/L,Glucose 10.20 g/L,beef powder 8.00 g/L,disodium hydrogen phosphate 2.20 g/L,threonine 0.04 g/L,aspartic acid 0.04 g/L,and serine 0.02 g/L.According to the optimal medium,the results are as follows:the extracellular surface tension is 45.74 mN/m,and the cell surface tension is 43.05 mN/m,which are 13.98%and 32.62%higher than before optimization.3.Through single factor,Plackett-Burman test,and response surface test,the fermentation conditions of Streptococcus thermophilus S73 to produce surface active substances were optimized,and the best fermentation conditions for producing surface active substances were obtained:the pH of the medium was 7.7,and the inoculum was 4.00%.Inoculate and incubate at 37? for 39 h.Under these conditions,the extracellular surface tension is 43.48 mN/m and the cell surface tension is 40.72 mN/m.Compared with the basal medium(glucose,peptone),the extracellular surface tension is reduced by 18.24%,and the cell surface tension is reduced by 36.27%.4.Through thin layer chromatography,color reaction,high performance liquid chromatography mass spectrometry,Fourier infrared spectroscopy and X-ray photoelectron spectroscopy(XPS),it is judged that the extracellular surfactant contains 42%di-rhamnolipids and lipopeptide mixture surfactants,cell surface active agents are mainly composed of lipopeptide surfactants.The emulsification experiment results prove that the two surfactants have a strong emulsification effect on edible oil,and the measured CMC value are both 600 mg/L.5.Bacteriostatic test proof test shows that the extracellular and cell surface active agent at a concentration of 2.5 g/L is effective against Candida albicans,Escherichia coli,Enterococcus faecalis,Staphylococcus aureus,and Pseudomonas aeruginosa at a concentration of 2.5 g/L.Both Gram-positive and negative bacteria such as spores have antibacterial effect.As the concentration increases,the antibacterial effect increases,but the antibacterial effect is different.At a concentration of 10 g/L,the growth inhibition rate of extracellular surfactants on E.coli and Staphylococcus aureus reached 94%and 95%,and the growth inhibition rate of cell surfactants on Staphylococcus aureus reached 84%.6.After the cell surface active agent was extracted from Streptococcus thermophilus S73,the sludge before and after the surface cell surface active substance was extracted from the cell surface was freeze-dried and then subjected to scanning electron microscope(SEM).The results showed that there was no significant difference in the structure and fermentation performance before and after extraction.Recyclable freeze-dried bacterial powder can be used to prepare yogurt direct throw starter without significant difference.In this study,a surfactant-producing strain-Streptococcus thermophilus S73 was obtained,and the culture composition and fermentation conditions for the production of surface-active substances were optimized,which effectively increased the production of surfactants,and the effects on the extracellular and cell surface were analyzed.The structure of the obtained different surfactant agents was characterized,and the preparation of emulsifying and antibacterial biosurfactants and their application in food were provided for reference.