| Viili is a kind of traditional fermented dairy product from Finland. It has the same fermentation principle with traditional yogurt and is fermented under effect of a variety of microbial with sugar as substrates. Compared with traditional yogurt in china, viili contains many kinds of lactic acid bacteria,yeast and filamentous fungi, including many strains which are very beneficial for human body. Because of less fermentation strains in china, it is necessary to study the composition of fermentation strains of viili and develop and utilize strains of viili. Studies have shown that the unique viscosity of viili is caused by the exopolysaccharides producted by lactic acid bacteria. In recent years, the exopolysaccharides of lactic acid bacteria has become a hot topic that the EPS of bacteria are researched and developed, caused the wide attention of scholars. Studies have shown that the exopolysaccharides of lactic acid bacteria not only can improve the texture of fermented milk products and the adsorption of strains of the intestinal mucosa, anti-tumor, enhance immunity, lower cholesterol level and so on, so the EPS from lactic acid bacteria have potential application value on food and pharmaceutical industries.Given viili possesses rich lactic acid bacteria and the function of exopolysaccharides, viili as research samples, lactic acid bacteria are isolated and its exopolysaccharides are studied. The main exopolysaccharides contents and results are as follows:1. The three strains isolated from viili were identified by physiological identification with API 50 CHL kits and 16 Sr DNA sequence alignment. The results showed the three strains were the same strain and Lactobacillus paracasei.2. Optimized production of the exopolysaccharides from lactic acid bacteria. Sugar yield as the index,the best nutritional conditions of the EPS from lactic acid bacteria were defined. The best nutritional conditions were 2% of fructose, 2.5% of soy peptone, 0.4% of ammonium citrate and in this conditions exopolysaccharides yield were 193.99mg/L and 2.6 times higher than before optimization. Sugar yield as the index, the best culture conditions of the EPS from lactic acid bacteria were defined. The best culture conditions were 4% of inoculation quantity, 32 h of incubation time and temperature at 32℃,exopolysaccharides yield reached 263.74mg/L and 36% higher than before.3. The exopolysaccharides from lactic acid bacteria were purified. The crude exopolysaccharides were purified on a DEAE-Sepharose Fast Flow column at first and neutral polysaccharides were obtained.Three components of neutral polysaccharide were named EPS1, EPS2 and EPS3. EPS1, EPS2 and EPS3 were further purified by gel filtration chromatography of Sephadex G-100. The results showed each fraction generated only one symmetrical elution peak. The average molecular weight of EPSfractions(EPS1 and EPS3) were determined by high performance gel permeation chromatography(HPGPC) and were 4.97 × 105 Da, and1.10 × 104 Da. The monosaccharide composition of EPS, EPS1, EPS2 and EPS3 were analyzed by pre-column derivatization HPLC method. the results showed that the crude exopolysaccharides consisted of mannose, rhamnose, galacturonic acid, glucose and galactose in an approximate molar ratio of 1:2.05:3.46:49.66:16.74, Fraction EPS1 was composed of mannose, rhamnose, glucose and galactose in an approximate molar ratio of 1:1.72:11.50:5.57, and fraction EPS1 and EPS3 were composed of mannose, glucose and galactose in an approximate molar ratio of 1:27.75:2.77 and 1:13.25:6.19.4. The effects of the crude exopolysaccharides on T lymphocyte proliferation was superior to purified exopolysaccharides EPS1, EPS2 and EPS3 and growth rate increased at first and then decreased. At concentration of 10μg/m L the maximum increment rate could reach 25.06%. EPS1, and EPS3 all promoted T lymphocyte proliferation and had certain immune activity with low concentration while had inhibitory effect at the high concentration.5. The exopolysaccharides from lactic acid bacteria could inhibit lipid peroxidation. With the increase of concentration, the inhibition rate of crude exopolysaccharides would increase and reach 24.52% with200μg/m L. EPS1, EPS2 and EPS3 could inhibit lipid peroxidation, but were not as good as the crude exopolysaccharides. at concentrations of 100μg/m L、100μg/m L and 200μg/m L, EPS1, EPS2 and EPS3 have maximum inhibition rate and reach 16.51%、 20.02% and 25.22%. The exopolysaccharides from lactic acid bacteria could scavenge Hydroxyl radical. The scavenging activity of EPS was much stronger than Vc at concentrations between 50-100μg/m L and reached 41% with 200μg/m L. The scavenging activity on hydroxyl radical of EPS1, EPS2 and EPS3 were all lower than EPS and the maximum scavenging rate reached 12.38% and 17.99%, EPS2 almost had no scavenging activity.6. The exopolysaccharides from lactic acid bacteria could protect the spleen cell of oxidative damage. It was significant that different concentrations of different samples protected cells. Adding the high doses of EPS and EPS3 had stronger protective effect on cells, which reached 24.25% and 23.27%. Adding the mid dose of EPS1 had stronger protective effect on cells, which reached 14.48%. Adding the low dose of EPS2 had stronger protective effect on cells, which reached 20.52%. The crude exopolysaccharides and its components could reduce the MDA value. When the concentrations of EPS, EPS1, EPS2 and EPS3 respectively were 100μg/m L 、80μg/m L、100μg/m L、40μg/m L, the MDA value was lowest and reduced 46%、42%、46% and 45% compared with the injury group. The crude exopolysaccharides and its components could rise the SOD value. When the concentrations of EPS,EPS1, EPS2 and EPS3 respectively were 100μg/m L 、80μg/m L、100μg/m L、40μg/m L, the SOD value was lowest and rised 17%、11%、13% and 14% compared with the injury group.Keyword: viili; lactic acid bacteria; exopolysaccharides; optimization; structural analysis;... |
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