The Structure Characterization And Impacts On Colorectal Cancer Mice Of Jujube Polysaccharides

Jujube(Ziziphus Jujuba Mill.)belongs to the genus Ziziphus(Rhamnaceae family)and has been considered to be functional food by National Health Commission.The valuable therapeutic agents of jujube fruit have been recorded in ancient Chinese books on herbal medicine.Modern pharmacological studies have shown that jujube has sedative and hepatoprotective effects,immunological and antioxidant activities,and anti-inflammatory effect,which has been attracted the attention of researchers in recent years.Polysaccharides are reported to be among the main biologically active constituents of jujube for the mentioned health benefits.However,to date,few studies have purified and structurally characterized jujube polysaccharides.The World Health Organization has estimated that colorectal cancer is the third commonest cancer globally,with a high mortality rate and an increasing incidence among adults under fifty years old.The jujube polysaccharides could strengthen intestinal barrier function,improve intestinal inflammation,promote intestinal peristalsis,and increase short chain fatty acids content in intestine.There is an increasing evidence of the anti-proliferative effects of plant polysaccharides on colon cancer cells and mice.However,to date,little attention has been given to the anti-colon cancer mechanisms in AOM/DSS-treated mice of jujube polysaccharides.A systematic technique for the purification of jujube polysaccharides was established and the physicochemical property,structure characterization,spectral characteristic,and micromorphology of different part purified jujube polysaccharides were explorated.The present study examined whether jujube polysaccharides influenced their ability to protect against Azoxymethane(AOM)/Dextran sulfate sodium(DSS)-induced colitis in mice after three months diet intererence.The fecal microbiota composition was examined by 16 S r DNA sequencing and fecal metabolome profile was evaluated by ultra high performance liquid chromatography tandem mass spectrometry(UHPLC-MS).The research further analyzed the correlation between intestinal flora and the differential metabolites in colon cancer mice through jujube polysaccharides intervention,and we believe that these will provide a theoretical basis for the development of functional food of jujube polysaccharides.The main research contents and results are as follows:(1)The two-step purification technical rout depending on the column chromatography of DEAD-Sepharose Fast Flow and Sephacryl S-300 was successfully established and applied to purify jujube polysaccharides.Only one neutral polysaccharide(PZMP1)and two acidic polysaccharide components(PZMP2 and PZMP3)were obtained from Ziziphus Jujuba cv.Muzao purification.The average molecular weights of them were 19.67 k D、62.73 k D、58.21 k D,respectively,and the purity of each polysaccharide was above 95%,which were satisfied the requirements in quality and quantity of the jujube polysaccharide components used for the further study.(2)Structural and linkage analysis by GC-MS and NMR suggested that the main chain of PZMP1 was composed of 1,3,5-linked Araf,1,5-linked Araf,or 1,4-linked Glcp.The PZMP2 had a linear backbone of(1→4)-linked Glap A and(1→2,4)-linked Rhap residues and PZMP3 was composed of a backbone of(1→4)-linked Glap A,few side chain is attached to the main chain at C-2 position of some residues.(3)The UV-Vis spectrums of three jujube purified fractions showed no optical absorption at 260 or 280 nm,showed that all of them contained no nucleic acids and proteins.It was found only one diffuse peak in PZMP1,and three or four diffuse peaks in PZMP2 and PZMP3 during the X-ray diffraction analysis.The scanning electron micrographs of PZMP1 showed that in occurs in a state of aggregation,with a thin slice shape and smooth surface,but the acidic polysaccharides exhibited special structural characteristics.Planar and 3D AFM images showed that there were many spherical lumps in jujube polysaccharide fractions.Through X diffraction,SEM and AFM detection,we found that neutral sugar components(PZMP1),acidic sugar components(PZMP2 and PZMP3),retained the plant polysaccharides structural form in the process of extraction,purification and purification.(4)An azoxymethane(AOM)/dextran sulfate sodium(DSS)-induced mouse model was established and administered with or without jujube polysaccharides(ZMP)for 13 weeks.High levels of IL-1β,IL-6,and TNF-α in serum were significantly inhibited by ZMP treatment,and ZMP treatment could dramatically decrease m RNA levels of IL-1β,TNF-α in colonic tissues.ZMP inhibited experimental colorectal cancer(CRC),resulting in overall attenuation of colitis and tumorigenesis,regulating the expression of lymphocytes and cytokines and improving survival rates of AOM/DSS induced mice.(5)According to the Alpha and Beta diversity analysis,comparing with the normal control group,the species of intestinal microflora in the jujube polysaccharide-treated group decreased,but there was no significant difference when compared with the model group.In jujube polysaccharide treatment group,the number of beneficial bacteria increased significantly,and the proportion of harmful bacteria have been decreased significantly.The abundance of Clostridium＿sp＿K4410MGS-306,Bifidobacterium,Bacteroides,and Lactobacillus Lactobacillus increased noticeably.(6)Compared to the model group,the amount of acetate,propionate,and butyrate significantly increased after three moths of jujube polysaccharide intake in AOM/DSS treated mice.The jujube polysaccharide intervention had a great modulating effect on metabolite profiles,causing a clear cluster separation and discrimination away from model group,and 39 metabolites had been up-regulated or down-regulated.Through person correlation analysis,some altered specific gut microflora-related metabolites were strongly associated with perturbed gut microbes.These significant perturbed metabolites participated in the citric acid cycle,glycolysis/gluconeogenesis pathway,and the indole-3-acetic acid and indole-3-acetate had been increased significantly,and the 5-hydroxyindole-3-acetic acid content were markedly decreased in faeces.The results in this study showed the jujube polysaccharides had different structural types of plant polysaccharides.The jujube polysaccharide had significant down regulation function in mice treated with AOM/DSS,and could notably alter structure of gut microbiota,and increasing beneficial bacterium,participating in tricarboxylic acid cycle,glycolysis/gluconeogenesis pathway producing SCFAs.This study could provide theoretical basis for the further application of jujube polysaccharides as well as new evidence of their potential in preventing colorectal cancer as functional food.