Extraction,Purification And Protection Of Exopolysaccharides From Lactobacillus Buchneri TCP016 On LPS/D-GalN-induced Liver Injury

Lactic acid bacteria exopolysaccharides?LAB EPS?are important metabolite of LAB,and have many biological activities.In this study,the EPS were separated and purified from probiotic LAB stored in the laboratory.Furthermore,the LPS/D-GalN-induced mouse model was established,which studied the protective effect of EPS016 on liver injury.Meanwhile,it is explored its effect on inflammatory cytokines,anti-oxidative factors,apoptosis,and intestinal flora.The results were as follows:1.The crude EPS produced from L.buchneri TCP016 were precipitated by adding alcohol,and were purified by DEAE-Sepharose Fast Flow and Sepharose CL-6B,then the purified EPS was obtained during the purification steps,named EPS016.The exopolysaccharide fraction had the absence of protein and nucleic acid.The EPS016 was characterized by infrared spectrum and showed that it had characteristic absorption peak of polysaccharides and?-pyranose ring configuration.The molecular weight of the EPS016 was 8.509×10⁴ Da.The EPS016 was mainly composed of rhamnose,glucuronic acid,xylose,galacturonic acid in an approximate molar ratio of 5.4:1.5:1.2:1.2.To research the effect of different concentrations of EPS016 on liver injury,Mouse liver injury model was established by intraperitineal?IP?injection of LPS/D-GalN.The results showed that EPS016 could significantly reduce the activity of ALT and AST in serum and liver homogenate of model group,and attenuated LPS/D-GalN-induced histologic damage of liver and intestine.EPS016 at concentration of 400 mg/kg showed the most significant among all treatment groups.3.After treatment with EPS016,the levels of TNF-?,IL-1?and IL-6 in serum and liver homogenate were significantly decreased,In contrast,the content of MDA in serum and liver homogenate were significantly increased,compared with the model group.In addition,EPS016 treatment could dose-dependently reduced the level of Caspase-3 and Caspase-8 in the liver homogenate.4.To explor the effect of EPS016 on intestinal flora in mice,high-throughput Illumina sequencing technology was used to detect mice colon contents.The results showed,compared with the model group,EPS016 could significantly increased the abundance and diversity of intestinal flora.Meanwhile,after EPS016 treatment,the abundance ofLacnodpiaceae_NK4A136_group,Ruminiclostridium₉ and Escherichia-Shigella were decreased,while the abundances of Ribaceae,Bacteroides,Lactobacillaceae and Prevotaceae were significantly increased,compared with the model group.These results demonstrated that EPS016 had partly protective effect on LPS/D-GalN-induced liver injury,and could regulate mice intestinal flora of LPS/D-GalN-induced liver injury.This study provides experimental basis for the development and application of LAB EPS.