Study On The Mechanism Of The Polysaccharidefrom Rosa Roxburghii Tratt Fruit In Regulating Intestinal Flora Composition And Intestinal Mucosal Barrier Function

Rosa roxburghii Tratt,also known as Shanwang fruit and Wenxian fruit,is a small perennial deciduous shrub of the genus Rosaceae in the Rosaceae family.It is also a kind of wild fruit that can be used as both medicine and food.It is distributed in Shaanxi,Gansu,Jiangxi,Yunnan and Guizhou,and is known as the "king of vitamin C" for its richness in polysaccharides,amino acids,vitamins and other nutrients and unique flavour.In our previous studies,we extracted a water-soluble polysaccharide(named RTFP)from Rosa roxburghii Tratt fruit,and found that RTFP had many biological activities such as anti-tumor,antioxidant,hypoglycemic and so on,but its prebiotic activity and related applications are rarely reported.Therefore,in our study,normal mice and obese mice were used as research objects to analyze intestinal flora metabolites by chemical analysis.And the mechanism of RTFP’s regulation of intestinal flora was revealed by microbiome analysis,and the molecular mechanism of RTFP’s protective effect on intestinal mucosal barrier of obese mice was explored.The protective effect of RTFP against intestinal mucosal barrier damage was then verified by Fecal Microbiota Transplantation(FMT)through regulation of intestinal flora.Finally,a kind of prebiotic polysaccharide buccal tablet was prepared with RTFP as the main raw material,which provided a theoretical basis for the high value of Rosa roxburghii Tratt.The main conclusions of this paper are as follows:1.A model of intestinal barrier damage was established by feeding mice on a high-fat diet(HFD),and RTFP was given by gavage to investigate its protective effect on the intestinal barrier of obese mice.The results showed that after 7 weeks of RTFP intervention,the symptoms of weight gain,fat accumulation and dyslipidemia in obese mice were improved;serum levels of D-lactic acid(D-LAC)and lipopolysaccharide(LPS)were reduced and diamine oxidase(DAO)activity was decreased;the levels of inflammatory factors TNF-α,IL-6 and IL-1β in serum,colon and ileum were reduced;the expression of TLR4/NF-κB pathway proteins was downregulated,reducing intestinal inflammatory responses;symptoms of intestinal crypt lesions,intestinal shortening and increased intestinal permeability were alleviated;the expression of tight junction proteins ZO-1,Claudin-1 and Occludin in the colon and ileum were upregulated and oxidative stress was inhibited,protecting the structural integrity of the intestinal barrier and alleviating oxidative damage caused by HFD.2.16 S r RNA high-throughput sequencing technology and chemical analysis were used to explore the mechanism of RTFP regulating intestinal flora and its metabolites.The results showed that RTFP improved the diversity of intestinal flora in mice.RTFP could improve intestinal barrier damage and inflammation and regulate immune response by regulating gut microbiota composition,including reducing the ratio of Firmicutes to Bacteroides,increasing the abundance of beneficial bacteria and decreasing the abundance of harmful bacteria.RTFP also increased the concentration of SCFAs in colon contents and fecal water content,reduced fecal ammonia content and promoted intestinal health in mice.3.Feces of mice in the control group,model group and RTFP treatment group were collected to verify the protective effect of RTFP on the intestinal mucosal barrier of obese mice by regulating gut microbiota through FMT.The results showed that after 9 weeks of FMT treatment,fecal flora of donor mice treated with RTFP significantly alleviated weight gain,abnormal lipid metabolism,liver fat accumulation,intestinal inflammation and enhanced intestinal permeability of the mice compared with the model control group.In addition,intestinal barrier dysfunction can be improved and intestinal function can be maintained by increasing fecal moisture content and reducing fecal ammonia content,upregulating tight junction protein expression and inhibiting oxidative stress response.Finally,based on 16 S r RNA high-throughput sequencing technology,it was found that fecal bacteria transplanted from donor mice treated with RTFP could also regulate the intestinal flora of HFD mice,reduce the ratio of Firmicutes to Bacteroides,increase the abundance of beneficial bacteria and reduce the abundance of harmful bacteria in the intestine.4.Using RTFP as the main ingredient and sorbitol,microcrystalline cellulose,citric acid and magnesium stearate as the auxiliary ingredients,a combination of single-factor sensory evaluation test and four-factor three-level orthogonal test was used to determine the optimal formulation of the main and auxiliary ingredients of this tablet.The results showed that the optimum formulation of prebiotic tablets was 60% RTFP,25% sorbitol,12% microcrystalline cellulose,2% citric acid and 1% magnesium stearate.The tablets obtained according to the optimum formulation have a smooth surface without cracks,are sweet and sour and have the characteristic flavour of Rosa roxburghii Tratt fruit.The tablet weight varies within ± 5%.The hardness of the tablets was in the range of 40-50 N.The average time of incubation of the tablets exceeded 10 min and the fragility of the tablets was less than 1%.Therefore,the tablets meet the requirements of the Chinese Pharmacopoeia for tablet quality inspection.