| Inflammatory bowel disease is a chronic intestinal disease,which is easy to recur and difficult to treat.Current treatments for inflammatory bowel disease are expensive and have side effects.Natural products have the advantages of low cost,less side effects and high safety.Citrus peels,as an important by-product of processing,are rich in pectin.Numerous studies have shown that gut microbiota disorder is one of the important mechanisms to induce inflammatory bowel disease.Short chain fatty acids,as an important metabolite of gut microbiota,play critical role in regulating intestinal immunity and anti-inflammatory.Citrus pectin can not be decomposed and utilized in the stomach and small intestine,but can be fermented by gut microbiota in the colon,producing short chain fatty acids.Studies have shown that citrus pectin can regulate gut microbiota and plays an important role in maintaining intestinal homeostasis and alleviating colitis.However,most of the citrus pectins reported in the literature are commercial pectin.The primary structure of pectin determines its higher structure,which ultimately affects its function.Citrus pectin is a complex heteropolysaccharide with varied structure and different structure of citrus pectin has great difference in health functions.It is necessary to investigate the primary structure and spatial conformation of citrus pectin.Therefore,it is of great significance to study the relationship between extraction,structure(primary structure and spatial conformation),and function of citrus pectin.In this study,citrus peel was used as the raw material to study the extraction method,structural characteristics,in vitro fermentation and the effect of alleviating colitis.The main findings are as follows:(1)Study on structure and in vitro fermentation characteristics of citrus pectin with different extraction methods: Firstly,the effects of acid extraction and alkali extraction on pectin yield of grapefruit peel were studied.The results showed that the yield of grapefruit pectin GP2 was the highest under p H 2 for acid extraction,and GP10 was the highest under p H 10 for alkali extraction.For enzyme extraction,cellulase was used to separate and release pectin.The results showed that under the optimal temperature and p H value,the yield of pectin was still low,and the yield of orange pectin and grapefruit pectin were only 11.7% and 10.4% after 4 h of extraction.In order to improve the yield of citrus pectin of cellulase extraction,p H 2,p H 10,and cellulase were selected for combined extraction,namely acid+cellulase extraction and alkali+cellulase extraction.The results indicated that the production of citrus pectin obtained by the two combined extraction methods was higher than that of the corresponding single extraction.In addition,the yield of citrus pectin extracted by alkali+cellulase was higher than that extracted by acid+cellulase.The primary structure and spatial conformation of orange and grapefruit pectin obtained from acid extraction(p H 2),alkali extraction(p H 10),cellulase extraction,acid+cellulase extraction and alkali+cellulase extraction were characterized.The results showed that the orange pectin and grapefruit pectin obtained by acid extraction,enzyme extraction,and acid+cellulase extraction were all high methoxyl pectin,and the RG-I region content was low,and the surface structure was lamellar and compact.The spatial conformation is extended and disordered,with large radius of gyration Rg and small cross-sectional radius Rc of pectin.Orange pectin and grapefruit pectin extracted by alkali and alkali+cellulase were low methoxyl pectin,with high content of RG-I region and loose and porous surface structure.Orange pectin(OP10+C)extracted by alkali+cellulase had the highest content of RG-I(61.80%),the most compact conformation and the largest Rc(4.78 nm).Based on the above results,an in vitro anaerobic fermentation model was established to simulate the metabolism of pectin in vivo.It was found that OP10+C had the optimal short chain fatty acids production capacity.It can increase the relative abundance of Bifidobacterium,Faecalibacterium,Roseburia,Subdoligranulum,Blautia and Phascolarctobacterium.It also inhibited the proliferation of Escherichia-Shigella,a potential pathogen.In conclusion,OP10+C extracted from orange pectin by alkali+cellulase can be used as a potential prebiotics and play a positive role in regulating intestinal health.The results can provide a reference for elucidating the relationship between the extraction,structure and in vitro fermentation characteristics of citrus pectin,which is conducive to the efficient preparation of citrus pectin with intestinal regulation function.(2)Study on alleviating effects and structure-activity relationship of citrus pectin on colitis in mice:The alleviating effects of orange pectin and grapefruit pectin on DSS induced acute colitis mice were studied by using citrus pectin with different structures obtained from acid extraction,alkali extraction,enzyme extraction,acid+enzyme extraction,and alkali+enzyme extraction.The data indicated that the extracted citrus pectin with different structure could relieve acute colitis in mice to a certain extent,but the alleviating effects was different.Among them,OP10+C extracted from orange pectin by alkali+cellulase exhibited the most obvious alleviating effect.The orange pectin OP10+C significantly reduced disease activity index,spleen index,colon redness and atrophy in mice with colitis.It was found that OP10+C significantly improved the inflammatory infiltration and crypt destruction of the colon,promoted the proliferation and mucus secretion of the colon goblet cells,and effectively protected the normal function of the intestinal barrier.OP10+C increased the α-diversity of gut microbiota.It also promoted the proliferation of anti-inflammatory bacteria Akkermansia,and bacteria associated with the production of short-chain fatty acids(Clostridium＿sensu＿stricto＿1,Lachnospiraceae＿NK4A136＿group,Lactobacillus,Ruminococcaceae＿UCG-014,and Prevotellaceae＿UCG-001).It effectively protected the balance of intestinal microecology in mice with colitis and promoted the production of colonic short chain fatty acids.Meanwhile,OP10+C promoted the expression of short chain fatty acid receptors GPR43 and GPR109 A and tightly connected proteins claudin-1,ZO-1 and Occludin.Moreover,the secretion of anti-inflammatory factor IL-10 was promoted,and the secretion of pro-inflammatory factors TNF-α,MPO,IL-6 and IL-1β was inhibited.By analyzing the correlation of extraction method,the structure characteristics,and colitis of citrus pectin,we found that the compact conformation and the larger Rc of OP10+C promoted its fermentation characteristics in mice.To be specific,the compact conformation and the larger Rc of OP10+C facilitated the bacteria to produce pectin enzyme hydrolysis,thus hydrolyzing pectin effectively,and producing the high content of short chain fatty acids,and alleviating colitis in mice.The results of this chapter elucidated the mechanism and structure-activity relationship of citrus pectin in alleviating colitis in mice,which provided a reference for further understanding of the extraction,structure,properties and function of citrus pectin,and also provided a scientific basis for the targeted preparation and nutrition of citrus pectin.This study is helpful to deeply understand the relationship between citrus pectin extraction method,structural characteristics,fermentation characteristics,and alleviating effects on colitis.Our study can provide a new idea for the targeted development of citrus pectin that alleviates colitis,provide reference for precise nutrition and health,and help to realize the high-value utilization of citrus pectin. |
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