Interaction Of Mulberry Pectin Polysaccharides With Cyanidin-3-O-Glucoside And Its Effect On Fermentation Properties Of Intestinal Microbiota

Interactions between anthocyanins and pectins are a new way to improve the stability of anthocyanins.These interactions are highly variable and complex and strongly depend on the fine structure of the anthocyanin and pectin.The addition of different pectinases resulted in significant differences in the structure of pectin in juice,which in turn influenced the pectin-anthocyanin interactions in juice.In this study,enzyme-assisted and non-enzyme-assisted extracts of mulberry pectin polysaccharides and mulberry cyanidin-3-O-glucoside(C3G)were used to investigate their interactions and their digestive and fermentation properties.The C3 G monomer was firstly removed from the mulberry anthocyanin by microporous resin and strongly acidic cationic resin,and the C3 G monomer was separated from a specific salt concentration of0.2 mol/L Na Cl solution with a purity higher than 95%,and then lyophilized on a C18 column after desalting.Six kinds of mulberry pectin polysaccharides with different yields,molecular weights,glyoxylate contents and monosaccharide compositions were obtained by hot water extraction(HW),high temperature and pressure extraction(HTP),pectinase assisted-extraction(PE),mixed enzyme-assisted extraction(pectinase: pectin lyase:cellulase = 1:1:1)(M),cellulase assisted-extraction(CE)and pectin lyase-assisted extraction(PL).The mulberry pectic polysaccharide-C3 G complexes were prepared as C3G-hot water extracted polysaccharide(HWC),C3G-high temperature and pressure extracted polysaccharide(HTPC),C3G-pectinase assisted-extraction of polysaccharide(PEC),C3G-mixed enzyme-assisted extraction of polysaccharide(pectinase: pectin lyase:cellulase = 1:1:1)(MC),C3G-cellulase assisted-extraction of polysaccharide(CEC)and C3G-pectin lyase assisted-extraction of polysaccharides(PLC).Color analysis,FTIR analysis,SEM analysis and circular dichroism showed that the six pectic polysaccharides bound to C3 G to different degrees and in different ways,with M and CE being the strongest.In addition,in vitro digestion results showed that MC and CEC were better able to resist gastrointestinal digestion and showed better bioaccessibility.Both HW and HTP increased the bioaccessibility of C3 G and helped it reach the colon.By adding cellulase to the supernatant product of non-cellulase assisted extracted mulberry pectin polysaccharides obtained by treating cellulose,the interaction between mulberry pectin polysaccharides extracted with cellulase and C3 G was again demonstrated by color analysis.Further in vitro fermentation experiments showed that C3 G was fermented to produce a series of metabolites,including anthocyanins,protocatechuic acid(PCA),2,4,6-trihydroxybenzaldehyde(TB),2,4,6-trihydroxybenzoic acid(TBA),p-Coumaric acid(PC)and p-Hydroxybenzoic acid(PHA).Fermentation of pectic polysaccharideC3 G complexes with either enzyme-assisted or non-enzyme-assisted extraction could regulate the timing of C3 G metabolite production and increase the yield of metabolites,but the results varied significantly depending on the pectic polysaccharide.The composition and ratio of intestinal microbiota in the pectic polysaccharide-C3 G complex group was significantly different compared to the C3 G alone fermentation group,and significant differences also occurred between the different pectin polysaccharide-C3 G complex groups.At the phylum level,the pectin polysaccharide-C3 G complex group showed an increase in Bacteroides,while the C3 G group decreased the percentage of Bacteroides.Their relationship was as follows: MC > CEC > PEC > HTPC > PLC >HWC > CON > C3 G.In contrast,the percentage of Fusobacterium was significantly reduced in the pectic polysaccharide-C3 G complex group and increased in the C3 G group.At the genus level,there was no significant change in the genus of Fusobacterium in the C3 G group compared to the CON group,a significant increase in the HTPC and PEC groups,and the rest of the pectic polysaccharide-C3 G complex groups showed a nonsignificant but up-regulated trend.The C3 G group tended to up-regulate Escherichia＿Shigella abundance compared to the CON group,while HWC,HTPC and PEC significantly increased their relative abundance and CEC and PLC tended to downregulate their abundance.the C3 G group significantly increased the abundance of Fusobacterium,but all pectic polysaccharide-C3 G complex groups significantly decreased their abundance.In addition,the main short-chain fatty acids produced after fermentation were acetic acid,propionic acid and butyric acid,and the production of SCFAs was generally higher in the enzyme-assisted pectic polysaccharide extraction group than in the non-enzyme-assisted pectic polysaccharide extraction group.In conclusion,mulberry pectin polysaccharides obtained by different extraction methods modulate the gastrointestinal digestive properties and intestinal health of C3 G in a structure-dependent manner.This study may provide a theoretical basis for the interaction mechanism and health benefits of mulberry pectin polysaccharides and C3 G.