| Mulberry fruit has various functional activities,and are of high nutritional and economic value.Mulberry fruit has been widely used as food material,and the progress of juice industry is one of the important ways which highly promote the healthy and sustainable development of mulberry processing.Polysaccharide is one of the main functional components of mulberry fruit,with a variety of biological activities such as antioxidant,hypoglycemic,hypolipidemic,hepatoprotective,immunomodulatory,and prebiotic effects.Enzymes are widely used in food processing,such as fruit juice manufacturing and extraction of plant-derived ingredients because of their numerous advantages such as being green,safe,and efficient.Therefore,this study investigated the effects of pectinase,pectin lyase,and cellulase treatment on the release of active components such as soluble sugars,phenols,flavonoids,and anthocyanins in mulberry juice.Meanwhile,the digestive characteristics of the whole mulberry juice prepared by enzymatic treatment in the oral and gastrointestinal tracts and the colonic fermentation characteristics were elucidated.In addition,the crude mulberry fruit polysaccharides were prepared with the three enzymes mentioned above.Their chemical components and structural characteristics were systematically investigated;while their colonic fermentation characteristics,effects on gut microbiota and structure-activity relationships were elucidated.The main research results are as follows:(1)The enzymatic treatment enhanced the release of soluble sugars and uronic acids from mulberry fruits,with cellulase having the best effect,followed by pectinase,and pectin lyase being the least effective.Meanwhile,pectinase mainly promoted the release of cyanidin-3-O-glucoside(C3G),pectin lyase promoted the release of phenols,flavonoids,C3G and cyanidin-3-O-rutinoside(C3R),cellulase mainly promoted the release of flavonoids.After in vitro digestion and absorption,the mulberry juice leftover showed decreased levels of polyphenols,flavonoids and C3G,and increased C3R,soluble sugars and uronic acids.Enzyme treatment improved the utilization of sugars in mulberry juice by the gut microbiota.Furthermore,the results of 16 S rDNA sequencing showed that mulberry juice leftover could regulate the composition and abundance of the gut microbiota.At the phylum level,mulberry juice residue increased the relative abundance of Bacteroidetes,Firmicutes,and Proteobacteria(without pectinase treated juice)and decreased the relative abundance of Fusobacteria.At the genus level,mulberry juice residue increased the relative abundance of Bacteroides,Klebsiella,Parabacteroides and Phascolarctobacterium and decreased the relative abundance of Fusobacterium and Escherichia.Among them,enzyme-treated mulberry juice was less effective than enzyme-free mulberry juice in increasing the relative abundance of Bacteroidetes and Firmicutes;Pectinase treatment reduced the increase of Proteobacteria during fermentation;Pectin lyase-treated mulberry juice was more effective than enzyme-free mulberry juice in reducing the relative abundance of Fusobacteria.Moreover,the fermentation of mulberry juice produced large amounts of short-chain fatty acids(SCFA),mainly acetic,propionic and butyric acids.(2)The enzyme treatment could improve the yield of crude black mulberry fruit polysaccharides(BMPs),with the highest yield of cellulase-assisted extraction,followed by pectinase,and a lower yield of pectin lyase-assisted extraction.The crude BMPs obtained with different enzymes varied in carbohydrate,uronic acid,protein,and total phenolic content.Structural analysis showed that the crude BMPs mainly contained water-soluble pectin.The enzyme treatment reduced the molecular weight and branching degrees of crude BMPs,and changed the monosaccharide composition,glycosidic bonds and microscopic morphology of polysaccharides.(3)Crude BMPs could be effectively fermented and utilized by gut microbiota,and the carbohydrates in the polysaccharides were consumed during fermentation,with gradually decreased molecular weight and monosaccharide content.Among them,the gut microbiota more easily consumed and utilized pectin lyase and water-extracted crude BMPs.The 16 S rDNA sequencing results showed that crude BMPs increased the relative abundance of Bacteroidetes and Firmicutes and decreased the relative abundance of Proteobacteria(without cellulase extracted polysaccharides)and Fusobacteria at the phylum level;and increased the relative abundance of Bacteroides and Klebsiella and decreased the relative abundance of Escherichia-Shigella and Fusobacterium at the genus level.Different enzyme treatments exhibited different effects on the structure and abundance of gut microbiota.Enzyme-assisted extracted crude BMPs were less effective than water-extracted polysaccharides in increasing the relative abundance of Bacteroidetes and Firmicutes and decreasing the relative abundance of Proteobacteria;the cellulase-extracted crude polysaccharide was more effective than water-extracted polysaccharides in decreasing the relative abundance of Fusobacteria.In addition,the fermentation of crude BMPs produced large amounts of acetic,propionic and butyric acids;Pectin lyase extracted polysaccharide produced the highest yields of acetic acid,propionic acid and total SCFAs;Meanwhile,pectin lyase and water extracted polysaccharides showed the highest production of butyric acid(no significant difference between the two).Treatments with pectinase and cellulase may not be conducive to the fermentation of extracted polysaccharides;however,polysaccharides extracted from water and pectin lyase exhibited better fermentability and prebiotic potential probably due to their higher molecular weight and more galactose and arabinose in their structure.In conclusion,the polysaccharide is one of the main compounds affecting colonic fermentation of mulberry juice,and the enzyme treatment affected the fermentation characteristics of mulberry juice by changing the structure of mulberry polysaccharides. |
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