Effects Of Polysaccharide From Fagopyrum Esculentum Moench Bee Pollen On Instinal Microflora

Fagopyrum esculentum Moench bee pollen was formed by storage and fermentation of pollen collected from Fagopyrum esculentum Moench anthers by bee.Fagopyrum esculentum Moench bee pollen was rich in nutrients such as protein,vitamins,and mineral,with the function of health care,and softening blood vessels,lowering blood pressure and lipids,making it a typical drug and food homologous substances.Polysaccharide was one of the main physiologically active components of Fagopyrum esculentum Moench bee pollen,but the structural characteristics,in vitro fermentation,and the effects on the intestinal microecology of the polysaccharide were not clear.In this paper,Fagopyrum esculentum Moench bee pollen was used as a raw material.Firstly,polysaccharides were extracted,and their physical and chemical properties and structural characteristics were analyzed and identified after further isolation and purification.Secondly,the effect of the polysaccharides on the metabolites of human fecal microbial flora was analyzed by establishing an in vitro fermentation model.Finally,a mouse intestinal dysbiosis model was established to explore the effects of polysaccharide from Fagopyrum esculentum Moench bee pollen on intestinal flora of mice and its repair effects on intestinal barrier,laying a theoretical foundation for the development and application of Fagopyrum esculentum Moench bee pollen.The main research results were as follows:1.The isolation,purification and structure dentification of polysaccharides from fagopyrum esculentum moench Bee Pollen were studied in this paper.The polysaccharide was extracted from fagopyrum esculentum moench bee pollen with extraction in water and alcohol precipitation.The crude polysaccharide was purified using papain treatment combined with Sevag method and DEAE-52 cellulose column chromatography,three polysaccharide fractions(WFPP-N?WFPP-1.WFPP-2)were obtained.The chemical structures of main components were characterized using technologies ultraviolet spectroscopy(UV),gas chromatography-mass spectrometry(GC-MS),high performance liquid chromatography(HPLC),infrared spectroscopy(IR).The results showed that the three fractions did not contain proteins and nucleic acid.etc.All polysaccharide fractions were composed of arabinose,glucose,galactose,rhamnose and xylose.The monosaccharide components in different polysaccharide fractions had different molar ratio.The molecular weights of the WFPP-1,WFPP-2 were 1.96×104 Da and 2.24×104 Da,and WFPP-2 contained two polysaccharide components the molecular weight of 2.40×104 Da and 4.38×103Da,respectively.The infrared spectrum of the polysaccharide showed that all fractions had polysaccharide characteristic absorption peak.2.By establishing an in vitro fermentation model,the pH value,total sugar content,NH3-N content and SCFAs content of fermentation broth were determined.The fermentation behavior of WFPP was analyzed to explore the beneficial effects of WFPP on human intestinal flora.The pH value and content of total sugar in the fecal culture declined as fermentation time increased.After 24h of fermentation,the pH value of WFPP-H group decreased from 7.26 to 4.64,the percentage of the total carbohydrates consumed in WFPP groups reached up to 90%.The accumulation of NH3-N content was reduced significantly by adding WFPP in culture broth,there are the higher NH3-N content in WFPP groups,which had the better effect;Acetic,propionic and n-butyric acid were the dominant short-chain fatty acids(SCFAs),which got a significantly rise in fermentation.The content of SCFAs from high to low is WFPP-H,WFPP-M,WFPP-L in the culture broth of polysaccharide groups.3.A 250 mg/kg/bw ceftriaxone sodium was used to establish a model of intestinal dysbiosis in Balb-c mouse.At the same time,the low,medium and high dose of MFPP(100,200 and 400 mg WFPP per1kg of weight,respectively)were fed to Balb-c mouse.Based on high-throughput sequencing technology,it was found that the OUT,Chao1,Ace index,and Shannon index in the polysaccharide group increased,and the Simpson index decreased.The changes in the NR group and WFPPL group were similar,while the correlation indexes in the WFPPM and WFPPH groups were basically restored to the level of Group C.Through the analysis of the histogram of the flora composition,different levels of WFPP could change the intestinal flora composition,which was beneficial to the recovery of intestinal resident bacteria and inhibited the multiplying of conditional pathogenic bacteria caused by intestinal disorder.The recovery effect of WFPPL group on intestinal disorders in mice was similar to that of NR group on the whole,the intervention effect was not obvious,The WFPPM and WFPPH groups were similar to group C,and the medium and high dose polysaccharides had better remodeling effect,the high dose polysaccharides had the best effect.According to the analysis of LEfSe diagrams,the different doses of WFPP could affect the strains that had significant effects in the changes of flora structure.4.After the intervention of WFPP,the food intake,weight,thymus index and spleen index of the mouse increased,and the LPS content in peripheral blood decreased.At the same time,HE and AB-PAS staining of the colon were found that colonic mucosa in WFPPM?WFPPH groups returned to be normal.The inflammatory cell infiltration basically disappeared,the crypt structcure basically returned to normal,the length of the villi was similar to that of the normal control group,and the number of goblet cells increased,Measurement of SIgA in intestinal mucosa revealed that SIgA was significantly increased in the WFPPH group.Analysis of TNF-??1L-6?1L-1?proinflammatory factors in intestinal mucosa showed,that the contents of TNF-??IL-6?1L-1? in different doses of polysaccharide groups were significantly reduced,which effectively reduced the inflammatory response caused by intestinal dysbiosis.