Extraction And Purification And Functional Characteristics Of Exopolysaccharides From Lactobacillus Pantheris TCP102

Lactic acid bacteria (LAB) exopolysaccharides (EPSs) are capsular polysaccharide slime polysaccha-rides which secreted by LAB into the extracellular environment during growth, is the secondary metabolites of LAB. A number of studies have proved that as a kind of macromolecule material, LAB EPS has no cytotoxicities, can enhance the immune cell function, restrain the proliferation of cancer cell, antioxidant, antivirus, and other functions, having a broad application in the future.In this study, we isolated and purified the EPSs of Lactobacillus pantheris TCP 102 strain firstly. We further investigated the effects of purified EPSs on proliferation in macrophage and inhibition of cancer cell lines. The results were as follows:1. We optimized the factors affecting EPSs production of Lactobacillus pantheris TCP 102 which were culture medium and fermentation time, temperature, initial pH value and inoculation amount. The preferable culture conditions were inoculation amount of 4%,37?, and pH 7 for 30h, cultured with MRS liquid medium. The EPS production was about 900 mg/L.2. The crude EPS were isolated by DEAE-Sepharose Fast Flow ion-exchange chromatography column and Sepharose CL-6B gel filtration chromatography, three pure EPSs were abtained, which were named EPS1, EPS2, and EPS3. EPS1 was neutral polysaccharide while EPS2 and EPS3 were acidic polysaccharides. All these three EPSs almost had not nucleic acids and proteins by UV scanning. The average molecular weight of EPS1, EPS2 and EPS3 were 20292 Da,22971 Da and 19301 Da respectively through GPC method. EPS1 was composed of D-mannose, D-glucose and D-lyxose in an approximate molar ratio of 14.44:2.94:1. EPS2 was composed of D-galactose, D-glucose, D-mannose and D-lyxose in an approximate molar ratio of 8.39:3.51:1.80:1. EPS3 was composed of D-galactose, D-glucose, D-mannose and D-lyxose in an approximate molar ratio of 6.61:4.67:1.23:1. EPS1, EPS2 and EPS3 had characteristic absorption peak of polysaccharides and a-pyranose ring configurations after the analysis of the infrared scanning.3. We found that Ana-1 macrophage proliferated after stimulated with EPS1, EPS2 and EPS3. Meanwhile, the production levers of NO, TNF-a and IL-6 were also enhanced in EPSs-stimulated Ana-1 cells. In addition, the same results were got in mice peritoneal macrophages stimulated with EPS1, EPS2 and EPS3. These results suggested that EPSs were capable of activating macrophages.4. We found that all these three EPS imhibited the proliferation on HCT-116 cells (human colon cancer cell line), Caco-2 cells (human colon cancer cell line), A-2780 cells (human ovarian cancer cell line) and BGE-803 cells (human gastric cancer cell line) after stimulated with EPS1, EPS2 and EPS3 during different time (24 h,48 h and 72 h). Moreover, the inhibition ability increased both in EPS concentration does and time dependent manner while the inhibition rate was EPS3> EPS2>EPS1. It was mentioned that the inhibition rate on A-2780 human ovarian cancer A-2780 cells were 72% after treating with EPS3 72 hours as a concentration of 500 ?g/L.