Screening Of Probiotic Lactic Acid Bacteria And Their Application On Food Processing

Numerous of probiotic functions of lactic acid bacteria and its products, such as regulating the intestinal flora, improving immunity, increasing antioxidative capacity, lowering cholesterol levels and preventing various diseases have been proved. Lacteins produced by lactic acid bacteria have been used widely in food preservation, fermentation, animal husbandry and cosmetics, for their efficiency, non-poisonous, high temperature resistance, no-drug resistance and giving no residue. In this study, lactic acid bacteria strains were isolated from traditional fermented foods, and potential probiotic strains were selected based on their performance of lactein-producing, acid and bile tolerances, adhesion properties to HT-29 cells and β-d-glucosidase activity. After candidate strains were identified and their probiotic functions were verified, their application on food processing was studied. The main results are as follows:(1) Screening and identification of probiotic lactic acid bacteria Among 300 isolated lactic acid bacteria strains,10 strains were verified to have the capacity to produce lactein, which were arranged to be MPE, MP166, MP30, MP85, MP159, MP149, LP4, MPN5, MP170 and MPC by their bacteriostatic activity. These 10 strains could survive at pH 3.0, in which 7 strains MP30, MP85, MP166, MPC, MPN5, MP149 and MP170 could even get growing in this pH conditon. Bile tolerance test indicated that strain LP4, MP30, MP166, MPN5, MP149, MP159 and MP170 could gain a survival rate higher than 50% in bile medium. Cell adhesion assay indicated that stran MP166 had the highest adhesion ability to HT-29 cells, the average adhesion rate to HT-29 for strain MP166 was 100:1, for strain LP4 was 36:1, for strain MP30 was 12:1, for other strains were low than 5:1. All of the 10 strains could produce β-d-glucosidase and LP4, MPC and MP166 had the highest ability to transform isoflavone glycosides to aglycones in fermented soymilk. Based on above-mentioned results, strain MP166 was chosen for further study. After the morphological, physiological and biochemistry characteristics test and 16 S rRNA sequence homology analysis, strain MP166 was classified as Lactobacillus plantarum.(2) Bioactivity detection in mice It showed that the number of Lactobacillus and Bifidobacterium were significantly increased, while that of Escherichia coli and Enterococcus were significantly decreased(P<0.05 or P<0.01) after fed with MP166 cells in mice. Even heat-inactivated MP166 cells could decrease the amount of Escherichia coli and Enterococcus. Contents of SIgA in the two different doses of MP166 groups increased significantly(P<0.01), as well as the immune organ index. IC and CFE of MP166 displayed good reduction activity and scavengeing capacity to DPPH· and ·OH, meanwhile the activities of SOD, CAT in blood evidently increased, on the contrary, the content of MDA decreased significantly(P<0.05 or P<0.01). Further more, MP166 was attested security. Results above proved the favourable beneficial function of MP166.(3) Purification of the lactein and its physicochemical property Fermentation Medium and conditions for production of the lactein were optimized to be MRS medium and pH=6.5, inoculation amount=1%, temperature =36?C and fermention time=24h. Product was extracted, purified and analyzed through ammonium sulfate precipitation, gel filtration chromatography, SP Sepharose Fast flow, SDS-PAGE and ESI-Q-TOF, which had excellent thermostability(120?C, 20min), pH-stability(pH 2-10) and UV-stability, and showed favorable tolerance to surfactants(Tween 20, Tween 80) and NaCl. It was deduced to be non-lantibiotic bacteriocin.(4)Application of the lactein in food preservation Purified lactein was added in milk and meat. Shelf life of raw milk could be extended to 1-3 times at 4?C or 20?C, meanwhile that of pasteurized milk were extended respectively to 5 times and 10 times at 20?C and 4?C. Similarly, meat at 4?C lasted 3-5 times longer. The lactein could inhibit growth of Listeria monocytogenes obviously in both milk and meat.(5) Research of high-yield soy isoflavone aglycones strain in fermented soybean milk The optimum fermentation condition of MP166 in soymilk was set as pH=7.47, inoculation amount=5.01% and fermentation temperature=42.59?C, through the single-factor tests and response surface methodology(RSM), and the procuction was verified to be 69.06μg/m L, 6.14 times as that of soybean milk. Fermentation characteristics of Lactobacillus plantarum MP166 were analyzed by determining pH value, proteolytic activity, viable cells, syneresis and amount of soy isoflavone aglycones in soymilk. Results revealed that L. plantarum MP166 grew well in the fermented soymilk and obtained a viable count of 4.5×108 CFU/m L. Proteolysis activity and water holding capacity of the soymilk were increased in the process of fermentation.