Preparation method and application of elaeagnus angustifolia gum water-soluble homogeneous polysaccharide

Abstract

The invention discloses a preparation method and application of elaeagnus angustifolia gum water-soluble homogeneous polysaccharide. According to the method, elaeagnus angustifolia gum is taken as a raw material, and the elaeagnus angustifolia gum water-soluble homogeneous polysaccharide is prepared through extraction, ethanol precipitation, separation and purification. Experiments show that the elaeagnus angustifolia gum water-soluble homogeneous polysaccharide reduces disease activity indexes and improves clinical symptoms by restoring body weight and excrement viscosity and reducing intestinal bleeding; the intestinal tract damaged by inflammation is relieved by repairing the mucosal barrier function of the colon tissue; by inhibiting mucosal lesion and remodeling an intestinal flora structure, dysbacteriosis induced by dextran sodium sulfate is reversed, and overall structure damage, mucosal lesion and colon tissue inflammation infiltration of DSS mice are effectively relieved. The application potential of the elaeagnus angustifolia gum water-soluble homogeneous polysaccharide in treatment of ulcerative colitis is disclosed, and the elaeagnus angustifolia gum water-soluble homogeneous polysaccharide is successfully endowed to the field of modern medicine from traditional application. Experimental research not only verifies the effectiveness of the compound, but also provides a new strategy and direct evidence for developing a novel and safe ulcerative colitis therapy derived from natural products, and has a clear application prospect.

Description

Technical Field

[0001] This invention belongs to the field of biomedicine, specifically relating to a method for preparing and applying a water-soluble homogeneous polysaccharide of jujube gum. Background Technology

[0002] Ulcerative colitis is a subtype of inflammatory bowel disease that primarily occurs in the proximal colon and rectum, manifesting as abdominal pain, diarrhea, weight loss, inflammation of the colonic mucosa, and rectal bleeding. The intestinal barrier is a physical barrier that prevents toxins, pathogens, and allergens from entering the circulatory system. Maintaining the integrity of the intestinal barrier is crucial for overall health. Various medications are available for treating ulcerative colitis, such as aminosalicylate, tumor necrosis factor-alpha agents, and immunosuppressants; however, many of these drugs cause multiple side effects. Therefore, there is an urgent need for a treatment method for ulcerative colitis with fewer side effects.

[0003] Sand dates ( Elaeagnus angustifolia L. Distributed in arid regions of western China, the Elaeagnus angustifolia (Rubus idaeus) exhibits strong adaptability. Numerous studies have shown that the entire Elaeagnus angustifolia tree has development value. However, to date, compared to its resource advantages and utilization value, its gum resources have not been fully developed and utilized. Elaeagnus angustifolia gum is a hard, glassy secretion exuded from the bark, resulting from natural disasters, human damage, or microbial invasion, serving as a protective mechanism after injury. Elaeagnus angustifolia gum is mainly composed of polysaccharides, with monosaccharide components including galactose, arabinose, rhamnose, glucuronic acid, and mannose; its water content (11.63%) is relatively high, exhibiting strong water absorption and retention capabilities. Studies have shown that Elaeagnus angustifolia gum polysaccharides can alleviate barrier function damage in mice with dry skin. Since the main component of most natural gums is polysaccharides, a deeper understanding of the physicochemical properties of natural gums requires research into polysaccharide chemistry.

[0004] Due to their structural diversity and complexity, most polysaccharides cannot be directly digested or absorbed in the upper gastrointestinal tract. However, they are degraded in the gut and used as a carbon source, utilizing the gut microbiota to promote the production of various metabolites related to host health benefits. The structural characteristics of polysaccharides are related to the presence of functional chemical groups and different monosaccharide units (glucose, galactose, arabinose, rhamnose, xylose, fucose, glucuronic acid, and methylglucuronic acid). Furthermore, the interaction mechanisms of target proteins from different gut microbiota indicate that they can degrade multiple monosaccharide chains in polysaccharides obtained from gums. The structural complexity of polysaccharides directly affects their fermentation characteristics in the gut, enabling them to serve as specific microbial carbon sources, selectively regulate the colonization and abundance of different bacteria, and ultimately exert beneficial regulatory effects on host physiological functions through metabolite regulation.

[0005] Currently, research on polysaccharide compounds shows a trend of multidisciplinary integration. Numerous studies have demonstrated that polysaccharides can protect the intestinal barrier from various factors and maintain bodily health. Antibiotic, dextran sulfate sodium, and cyclophosphamide-induced mouse models of inflammatory bowel disease are widely used to study the protective effects of oligosaccharides, polysaccharides, and flavonoids on mice, and are recognized as reliable animal models in gut-related research. Based on the reliability and applicability of this model, this study further systematically evaluated the role and mechanism of polysaccharides in intervening in dextran sulfate sodium-induced colitis, focusing on their regulatory functions on intestinal flora structure and short-chain fatty acid metabolism. The results not only help to reveal the potential mechanisms by which polysaccharides alleviate intestinal inflammation but also provide theoretical basis and experimental support for the development of functional foods or drugs targeting the gut microbiota, possessing significant scientific value and application prospects. Summary of the Invention

[0006] The present invention discloses a method for preparing and applying a water-soluble homogeneous polysaccharide from jujube gum. The method uses jujube gum as raw material, extracts it, and then precipitates it with ethanol before separating and purifying it to prepare the water-soluble homogeneous polysaccharide. Experiments show that the water-soluble homogeneous polysaccharide improves clinical symptoms by restoring body weight and fecal viscosity, reducing bleeding, and lowering the disease activity index; it also protects the intestinal tract from inflammation by repairing the mucosal barrier function of colonic tissue; the water-soluble homogeneous polysaccharide prepared by this invention alleviates intestinal damage caused by sodium dextran sulfate by inhibiting mucosal damage and regulating intestinal flora imbalance. This invention reveals for the first time the application potential of water-soluble homogeneous polysaccharide from jujube gum in the treatment of ulcerative colitis, successfully extending its application from traditional uses to the modern medical field. Experimental studies not only verify its effectiveness but also provide new strategies and direct evidence for developing novel and safe ulcerative colitis therapies derived from natural products, demonstrating clear application prospects.

[0007] The preparation method of the water-soluble homogeneous polysaccharide of jujube gum according to the present invention is carried out according to the following steps: a. Water extraction and alcohol precipitation: Prepare a 0.5%-2% aqueous solution of jujube gum, extract for 8-12 hours, centrifuge at 5000 rpm for 10 minutes, take the supernatant, slowly add 95% ethanol at a volume ratio of 1:3-5, carry out alcohol precipitation and leave overnight, centrifuge again, collect the precipitate, reconstitute with deionized water, dialyze, freeze dry to obtain water-soluble crude polysaccharide of jujube gum; b. Anion exchange column separation: The crude polysaccharide of jujube gum obtained in step a is prepared into a 2-4 mg / mL solution with pure water and loaded onto a DEAE agarose anion exchange column. It is eluted sequentially with purified water and 0.1-0.8 mol / L NaCl solution. The eluent is monitored by the anthrone-sulfuric acid method, and the elution curve is plotted. The effective fraction eluted with water is collected, dialyzed, and freeze-dried to obtain the crude polysaccharide of jujube gum. The aqueous polysaccharide fraction is then separated. c. Purification by cross-linked glucosamine gel column: The aqueous polysaccharide obtained in step b was prepared into a solution of 5-10 mg / mL, loaded onto a cross-linked glucosamine gel column, eluted with water, and the eluent was monitored by the anthrone-sulfuric acid method. The elution curve was plotted, the peak top was collected, dialyzed and freeze-dried to obtain water-soluble homogeneous polysaccharide of jujube gum.

[0008] Application of the water-soluble homogeneous polysaccharide obtained by the method in the preparation of a treatment for ulcerative colitis.

[0009] The present invention relates to a method for preparing and applying a water-soluble homogeneous polysaccharide called jujube gum, wherein the ulcerative colitis model is a C57BL / 6 mouse induced by sodium dextran sulfate. Attached Figure Description

[0010] Figure 1 Elution curves for the separation of water-soluble polysaccharides of jujube gum using a DEAE agarose coagulation ion exchange column in this invention; Figure 2 Elution curves for the purification of water-soluble homogeneous polysaccharides of jujube gum using cross-linked glucono gel column in this invention; Figure 3 This is a high-performance liquid chromatography-gel chromatogram of the water-soluble homogeneous polysaccharide of jujube gum in this invention; Figure 4 This is a diagram showing the monosaccharide composition of the water-soluble homogeneous polysaccharide of jujube gum in this invention; Figure 5 This invention relates to a water-soluble homogeneous polysaccharide derived from jujube gum to alleviate colitis induced by sodium dextran sulfate. The anus represents the mouse; body weight changes; mouse colon length; and mouse spleen size are shown in the figure. P<0.05 ; Figure 6 The water-soluble homogeneous polysaccharide of jujube gum in this invention can alleviate intestinal tissue damage in mice with colitis (H&E staining of distal colon tissue). Figure 7 The present invention reconstructs the principal component analysis (PCA) diagram of the gut microbiota in DSS-induced colitis mice using the homogeneous polysaccharide of jujube gum. Figure 8The present invention describes the analysis of the bacterial composition at the Bar chart phylum level in the gut microbiota of DSS-induced colitis mice using homogeneous polysaccharide remodeling of the intestinal microbiota by jujube gum homogeneous polysaccharide. Figure 9 This invention provides a Bar diagram of bacterial composition at the genus level in the gut microbiota of DSS-induced colitis mice, based on the remodeling of the gut microbiota by the homogeneous polysaccharide of jujube gum. The diagram shows the bacterial composition at the genus level. (Note: The x-axis / y-axis represents the sample name, and the y-axis / x-axis represents the proportion of the species in the sample. Different colored bars represent different species, and the length of the bars represents the proportion of that species.) Figure 10 The present invention reconstructs the LEfSe multi-level species hierarchy tree diagram of the gut microbiota in DSS-induced colitis mice using the homogeneous polysaccharide of jujube gum. Figure caption: Different colored nodes represent microbial groups that are significantly enriched in the corresponding group and have a significant effect on the differences between groups; light yellow nodes represent microbial groups that have no significant differences in different groups or have no significant effect on the differences between groups. Figure 11 The linear discriminant analysis (LDA) effect (LEfSe) of the gut microbiota in mice with DSS-induced colitis was analyzed by the homogeneous polysaccharide of jujube gum in this invention. Figure caption: The LDA discriminant bar chart statistically analyzes the microbial groups with significant effects in multiple groups. The LDA score obtained by LDA analysis (linear regression analysis) indicates that the larger the LDA score, the greater the influence of species abundance on the differential effect. Figure 12 This invention relates to a comparative analysis of multiple groups of gut microbiota in DSS-induced colitis mice using homogeneous polysaccharide remodeled from jujube gum. The analysis shows the average relative abundance differences of the same species among different groups. (Figure caption: The vertical axis represents the species name at different taxonomic levels, and the horizontal axis represents the percentage abundance of a specific species in the sample. Different colors represent different groups.) The rightmost figure shows... P Value, * 0.01< P ≤ 0.05, ** 0.001< P ≤ 0.01, *** P ≤ 0.001. Detailed Implementation

[0011] The present invention will be further described in detail below with reference to the embodiments. However, the uses and purposes of these exemplary embodiments are only for illustrating the present invention and do not constitute any limitation on the actual protection scope of the present invention, nor are they intended to limit the protection scope of the present invention to this. Example 1

[0012] Preparation of water-soluble homogeneous polysaccharide from jujube gum a. Water extraction and alcohol precipitation: Prepare a 0.5% aqueous solution of jujube gum, extract for 8 hours, centrifuge at 5000 rpm for 10 minutes, take the supernatant, slowly add 95% ethanol at a volume ratio of 1:3, carry out alcohol precipitation and leave overnight, centrifuge again, collect the precipitate, reconstitute with deionized water, dialyze, freeze dry to obtain water-soluble crude polysaccharide of jujube gum; b. Anion exchange column separation: The crude polysaccharide of jujube gum obtained in step a was prepared into a 2 mg / mL solution with pure water and loaded onto a DEAE agarose anion exchange column. It was eluted sequentially with purified water and 0.1-0.8 mol / L NaCl solution. The eluent was monitored by the anthrone-sulfuric acid method, and the elution curve was plotted. The effective fraction eluted with water was collected, dialyzed, and freeze-dried to obtain the crude polysaccharide of jujube gum. The aqueous polysaccharide fraction was then separated. c. Purification by cross-linked glucosamine gel column: The aqueous polysaccharide obtained in step b was prepared into a 5 mg / mL solution, loaded onto a cross-linked glucosamine gel column, eluted with water, and the eluent was monitored using the anthrone-sulfuric acid method. The elution curve was plotted, the peak tip was collected, dialyzed, and freeze-dried to obtain water-soluble homogeneous polysaccharide of jujube gum. Example 2

[0013] Preparation of water-soluble homogeneous polysaccharide from jujube gum a. Water extraction and alcohol precipitation: Prepare a 1% aqueous solution of jujube gum, extract for 10 hours, centrifuge at 5000 rpm for 10 minutes, take the supernatant, slowly add 95% ethanol at a volume ratio of 1:4, carry out alcohol precipitation and leave overnight, centrifuge again, collect the precipitate, reconstitute with deionized water, dialyze, freeze dry to obtain water-soluble crude polysaccharide of jujube gum. b. Anion exchange column separation: The crude polysaccharide of jujube gum obtained in step a was prepared into a 3 mg / mL solution with pure water and loaded onto a DEAE agarose anion exchange column. It was eluted sequentially with purified water and 0.5 mol / L NaCl solution. The eluent was monitored by the anthrone-sulfuric acid method, and the elution curve was plotted. The effective fraction eluted with water was collected, dialyzed, and freeze-dried to obtain the crude polysaccharide of jujube gum. The aqueous polysaccharide fraction was then separated. c. Purification by cross-linked glucosamine gel column: The aqueous polysaccharide obtained in step b was prepared into an 8 mg / mL solution, loaded onto a cross-linked glucosamine gel column, eluted with water, and the eluent was monitored using the anthrone-sulfuric acid method. An elution curve was plotted, the peak tip was collected, dialyzed, and freeze-dried to obtain water-soluble homogeneous polysaccharide of jujube gum. Example 3

[0014] Preparation of water-soluble homogeneous polysaccharide from jujube gum a. Water extraction and alcohol precipitation: Prepare a 2% aqueous solution of jujube gum, extract for 12 hours, centrifuge at 5000 rpm for 10 minutes, take the supernatant, slowly add 95% ethanol at a volume ratio of 1:5, carry out alcohol precipitation and overnight, centrifuge again, collect the precipitate, reconstitute with deionized water, dialyze, freeze dry to obtain water-soluble crude polysaccharide of jujube gum. b. Anion exchange column separation: The crude polysaccharide of jujube gum obtained in step a was prepared into a 4 mg / mL solution with pure water and loaded onto a DEAE agarose anion exchange column. It was eluted sequentially with purified water and 0.8 mol / L NaCl solution. The eluent was monitored by the anthrone-sulfuric acid method, and the elution curve was plotted. The effective fraction eluted with water was collected, dialyzed, and freeze-dried to obtain the crude polysaccharide of jujube gum. The aqueous polysaccharide fraction was then separated. c. Purification by cross-linked glucosamine gel column: The aqueous polysaccharide obtained in step b was prepared into a 10 mg / mL solution, loaded onto a cross-linked glucosamine gel column, eluted with water, and the eluent was monitored using the anthrone-sulfuric acid method. An elution curve was plotted, the peak tip was collected, dialyzed, and freeze-dried to obtain water-soluble homogeneous polysaccharide of jujube gum. Example 4

[0015] Water-soluble homogeneous polysaccharide of jujube gum and its effect on intestinal damage induced by sodium dextran sulfate: Animal experiments: Eight-week-old female C57BL / 6 mice (17-20 g, SPF) were used at the Animal Center of Xinjiang Medical University. The experiment followed the guidelines of the Animal Protection Committee and the National Research Council's "Guidelines for the Care and Use of Laboratory Animals." Mice were randomly divided into four groups: control group, jujube gum water-soluble homogeneous polysaccharide group, dextran sulfate sodium group, and dextran sulfate sodium + jujube gum water-soluble homogeneous polysaccharide group, with seven mice in each group. The first two groups were given a normal diet for 10 days. The latter two groups had 3% dextran sulfate sodium added to their drinking water for 7 consecutive days (days 4-10) to establish an acute intestinal injury model. Simultaneously, the mice in the dextran sulfate sodium + jujube gum water-soluble homogeneous polysaccharide group were administered 100 mg / kg jujube gum water-soluble homogeneous polysaccharide by gavage for 10 consecutive days. Dextran sulfate sodium-induced colitis presents with symptoms such as weight loss, rectal bleeding, shortened colon length, and splenomegaly, which can be used as an indicator to evaluate the severity of colitis. Record lifestyle symptoms, including weight, stool consistency, and occult blood, and the results are as follows: Figure 5 As shown: The results showed that: Figure 5In the study, compared with the control group, mice induced by sodium dextran sulfate showed obvious pathological phenomena, such as rectal bleeding and weight loss. Mice in the group with water-soluble homogeneous polysaccharide of jujube gum did not show the above phenomena, suggesting that oral administration of water-soluble homogeneous polysaccharide of jujube gum is harmless to C57BL / 6 mice. As expected, oral administration of 100 mg / kg / d water-soluble homogeneous polysaccharide of jujube gum significantly alleviated rectal bleeding and promoted weight gain. The results showed that the treatment with water-soluble homogeneous polysaccharide of jujube gum could effectively control rectal bleeding caused by sodium dextran sulfate and alleviate the apparent symptoms of intestinal damage in mice by significantly increasing their body weight. Histological analysis: The colon length and spleen index of mice in each group were measured, and the results are as follows: Figure 5 As shown: The results showed that the intervention of water-soluble homogeneous polysaccharide of jujube gum significantly alleviated the shortening of the colon. Similarly, water-soluble homogeneous polysaccharide of jujube gum could also significantly inhibit spleen swelling induced by sodium dextran sulfate. The results showed that water-soluble homogeneous polysaccharide of jujube gum could effectively alleviate ulcerative colitis induced by sodium dextran sulfate. The proximal segment of the colon and rectum was removed and rinsed with pre-cooled PBS; colon sections were fixed in formaldehyde, embedded in paraffin, sectioned (3-4 μm), dewaxed, and stained with hematoxylin and eosin (H&E). The results are as follows: Figure 6 As shown: Diseased colon tissue is a characteristic phenotype of mice with ulcerative colitis: the intestinal damage of mice in each group was evaluated by hematoxylin and eosin (H&E) staining; The results showed that the staining results of hematoxylin and eosin (H&E) in the distal colon of the control group and the jujube gum water-soluble homogeneous polysaccharide group were as follows: Figure 3 The results showed that the intestinal structure was normal, with goblet cells, crypts, mucosa, submucosa, and muscularis propria in good condition. Conversely, compared to the control group and the group treated with jujube gum water-soluble homogeneous polysaccharide, induction with 3% sodium dextran sulfate resulted in multifocal ulcers in the intestinal tissue (orange curved polygons), loss of crypt structure from the mucosal epithelium to the lamina propria, a small amount of connective tissue hyperplasia, and damage extending to the submucosa, with a small amount of inflammatory cell infiltration including lymphocytes and granulocytes (yellow arrows). After treatment with jujube gum water-soluble homogeneous polysaccharide, the intestinal tissue surface showed a single layer of columnar epithelium, and the morphology and structure of the mucosal epithelial cells tended to be normal; the number of crypts in the lamina propria gradually increased, significantly alleviating ulcers and other abnormalities. The results showed that the water-soluble homogeneous polysaccharide of jujube gum effectively alleviated intestinal damage induced by sodium dextran sulfate in mice, with similar efficacy to the control group. The water-soluble homogeneous polysaccharide intervention of jujube gum had a significant protective effect against sodium dextran sulfate-induced intestinal mucosal damage. Gut microbiome sequencing: Colonic feces from 5 animals were randomly selected from each group and stored at -80℃. Total bacterial DNA was prepared using the EZNA™ Mag-Bind Soil DNA Kit and analyzed by Shanghai Meiji Biopharmaceutical Technology Co., Ltd. using 16S rRNA gene sequencing. Method ① Principal Component Analysis (PCA) is a technique for simplifying data analysis. This method can effectively identify the most important elements and structures in the data, remove noise and redundancy, reduce the dimensionality of the original complex data, and reveal the simple structure hidden behind the complex data. A two-dimensional scatter plot is used to display the similarity and difference between the control and treatment groups, and the distance between samples (default Euclidean distance algorithm) reflects the clustering degree of the sample communities. This analysis is statistically applied using ANOSIM / adonis / PERMAVONA to present the differences in community changes between the control and treatment groups. Figure 7 ); Method ② Community Composition Analysis: This method displays the microorganisms present in the disease and healthy groups and their relative abundance, providing a clear visual indication of species variation trends across different groups. This analysis is a primary method for understanding the microbial community structure in samples. Bar diagrams show the microbial communities of all samples / groups; this paper demonstrates the phylum-level distribution. Figure 8 , belongs to horizontal Figure 9 ; Method ③ Lefse multilevel discriminant analysis (multi-level: phylum, class, order, family, genus, species) performs differential testing at multiple levels, analyzes differentially differentiated species at multiple levels, and uses LDA values ​​to measure the magnitude of the species' influence on the differential effect, suggesting that the species may play a key role in the occurrence and development of the disease; this analysis can also be used as one of the methods to find biomarkers between disease groups and healthy groups. Species differences at different taxonomic levels are displayed in the form of a developmental genus diagram, intuitively reflecting the differentially differentiated species at different species levels obtained between different groups; bar charts display the LDA values ​​of different differentially differentiated species, intuitively showing the magnitude of the influence of the identified landmark species between different groups on the differential effect. Figure 10 , Figure 11 ); Method ④ Species Difference Analysis: Using the between-group difference test method, based on the obtained community abundance data, rigorous statistical methods were applied to perform hypothesis testing on species among multiple groups of microbial communities, assess the significance level of species abundance differences, and obtain information on species with significant differences among multiple groups. Bar charts display the differences in the average relative abundance of the same species among different groups, and indicate whether the differences are significant (P-value, asterisks represent significant differences). This visually demonstrates the significance of differences in the same species among multiple different groups. Figure 12 ); The results show that: ① Figure 7 It was observed that the sodium dextran sulfate group and the control group were significantly separated, indicating a significant difference in microbial composition. The distribution of the blank group and the jujube gum water-soluble homogeneous polysaccharide group partially overlapped, indicating similar microbial composition. Ingestion of jujube gum water-soluble homogeneous polysaccharide alone did not cause drastic disturbances or adverse effects on the core structure of the intestinal flora in healthy mice, demonstrating its good biocompatibility. However, the jujube gum water-soluble homogeneous polysaccharide + sodium dextran sulfate group was closer to the control group and farther from the sodium dextran sulfate group than the sodium dextran sulfate group. ② Differences in microbial composition among different groups were compared at the phylum and genus levels. Figure 8 and Figure 9At the phylum level, the gut microbiota in all groups was dominated by Firmicutes (Bacillota) and Bacteroides, a typical characteristic of the gut in healthy mammals. Other phyla included Actinomycetota, Pseudomonadota, and Vernuomicrobiota. The control group and the jujube gum water-soluble homogeneous polysaccharide group showed highly similar microbiota structures, with a stable ratio of Firmicutes (Bacillota) to Bacteroides. In the dextran sulfate sodium group, the abundance of Bacteroides increased sharply, while the abundance of Firmicutes (Bacillota) decreased significantly. This resulted in a significant decrease in the Firmicutes (Bacillota) / Bacteroides (Bacteroides) (F / B) ratio, a typical marker of dysbiosis. The phylum Vernuomicrobiota (containing the important Akkermansia) completely disappeared, while the abundance of Pseudomonadota increased. In the group treated with jujube gum water-soluble homogeneous polysaccharide + sodium dextran sulfate (recovery effect): compared to the sodium dextran sulfate group, the abundance of Bacteroidetes decreased, while the abundance of Bacillota increased, normalizing the F / B ratio; Vernuomicrobiota reappeared; this is a very positive signal, usually indicating the recovery of the intestinal mucus layer and barrier function; the abundance of Pseudomonadota was suppressed. Genus-level analysis can more precisely reveal which specific bacteria changed, and the connection with physiological function is more direct. At the genus level, the flora was mainly dominated by genera related to Mucor (Muribaculaceae) and Lachnospiraceae. The structure of the control group and the jujube gum water-soluble homogeneous polysaccharide group remained very similar. In the polysaccharide group, the relative abundance of Akkermansia was noticeably increased, suggesting that the water-soluble homogeneous polysaccharide of jujube gum may itself promote the growth of these beneficial bacteria. The abundance of Helicobacter spp. was significantly reduced after intervention with sodium dextran sulfate. This depletion is a typical characteristic of sodium dextran sulfate-induced colitis. The abundance of harmful or opportunistic pathogens such as Helicobacter (a conditionally pathogenic bacterium) and Desulfovibrio (which produces cytotoxic hydrogen sulfide) was significantly increased.Akkermansia, a recognized beneficial bacterium, completely disappeared; the abundance of bacteria such as Dubosiella and Ileibacterium also changed significantly. In the group of water-soluble homogeneous polysaccharide of jujube gum + sodium dextran sulfate (recovery effect): the abundance of Norank_f_Muribaculaceae was effectively restored; the abundance of harmful bacteria such as Helicobacter and Desulfovibrio was significantly inhibited; Akkermansia reappeared and its abundance was considerable; ③ Through LEfSe analysis, the action pathway of water-soluble homogeneous polysaccharide of jujube gum can be clearly outlined: sodium dextran sulfate attack leads to the depletion of healthy baseline flora (such as Christensenellaceae, Romboutsia), while promoting the excessive proliferation of a group of potentially harmful microorganisms (such as hydrogen sulfide-producing Desulfovibrio). Intervention with water-soluble homogeneous polysaccharides from jujube gum inhibited harmful bacteria: Water-soluble homogeneous polysaccharides from jujube gum effectively inhibited harmful bacteria such as *Desulfovibrio* enriched in the sodium dextran sulfate group. Promoting core beneficial bacteria, water-soluble homogeneous polysaccharides from jujube gum specifically and significantly enriched *Akkermansia*. This is the most crucial link in its protective effect, resisting inflammation by strengthening the intestinal barrier. Regulation of immune homeostasis: Even in healthy mice, water-soluble homogeneous polysaccharides from jujube gum could promote the growth of segmented filamentous bacteria (*Candidatus arthromitus*) with immunomodulatory functions. The growth of these bacteria provides a mechanistic explanation for their broad immunomodulatory effects. ④ Results of multi-group comparative analysis: Ruminococcus family, associated with health and homeostasis, is an important fiber-degrading bacterium that produces short-chain fatty acids such as butyrate, crucial for maintaining intestinal barrier health and anti-inflammation; the abundance of Ruminococcus family was significantly reduced in the sodium dextran sulfate group; its abundance was significantly restored in the sodium dextran sulfate + jujube gum water-soluble homogeneous polysaccharide group. Prevostia, associated with the fermentation of dietary fiber... Prevotellaceae increased significantly in abundance after induction with sodium dextran sulfate; however, the intervention of water-soluble homogeneous polysaccharide from jujube gum inhibited the abnormal increase of Prevotellaceae induced by sodium dextran sulfate to some extent. Bacteroidetes, a family associated with inflammation and disease states, showed a sharp increase in abundance in the sodium dextran sulfate group, a typical characteristic of sodium dextran sulfate-induced dysbiosis; water-soluble homogeneous polysaccharide from jujube gum effectively inhibited the abnormal proliferation of Bacteroidetes induced by sodium dextran sulfate, helping to restore the balance of the microbial community.Increased abundance of Enterobacteriaceae was strongly associated with intestinal inflammation and impaired barrier function. Its abundance surged significantly in the sodium dextran sulfate group. Jujube gum water-soluble homogeneous polysaccharide exhibited a strong inhibitory effect, causing its abundance in the sodium dextran sulfate + jujube gum water-soluble homogeneous polysaccharide group to drop sharply to near-normal levels. This provides key and compelling evidence that jujube gum water-soluble homogeneous polysaccharide alleviates colitis. The results showed that: ① Sodium dextran sulfate disrupts the stable structure of the gut microbiota, while water-soluble homogeneous polysaccharide of jujube gum effectively resists this disruption, reshaping the disordered microbiota and restoring it to a near-healthy state, without harming the normal microbiota itself. Intervention with water-soluble homogeneous polysaccharide of jujube gum reversed the microbiota dysbiosis induced by sodium dextran sulfate, restoring the disordered microbiota structure to a healthy state. ② Ingestion of water-soluble homogeneous polysaccharide of jujube gum alone did not disturb the core structure of the gut microbiota in healthy mice, demonstrating its safety. Sodium dextran sulfate intervention successfully induced severe gut microbiota dysbiosis. However, intervention with water-soluble homogeneous polysaccharide of jujube gum effectively reversed the microbiota dysbiosis induced by sodium dextran sulfate, reshaping the microbiota structure back to a near-healthy state. Changes in the microbiota at the genus level provide strong evidence for the protective effect of water-soluble homogeneous polysaccharide of jujube gum. The water-soluble homogeneous polysaccharide of jujube gum also regulated the abundance of bacteria such as the NK4A136 group (beneficial bacteria that produce short-chain fatty acids) of the Trichophyton family. ③ LEfSe analysis statistically confirmed that the water-soluble homogeneous polysaccharide of jujube gum does not generally alter the gut microbiota, but rather selectively promotes beneficial bacteria and eliminates harmful bacteria. It inhibits disease-associated harmful bacteria while promoting core beneficial bacteria that are crucial for barrier function and immune regulation, thereby guiding the dysregulated gut microbiota back to a healthy state. This provides in-depth microbiological evidence for its treatment of ulcerative colitis. ④ The water-soluble homogeneous polysaccharide of jujube gum can reverse the dysbiosis induced by sodium dextran sulfate in the gut. The water-soluble homogeneous polysaccharide of jujube gum reverses the dysbiosis induced by sodium dextran sulfate by reshaping the gut microbiota structure, specifically by inhibiting pro-inflammatory bacteria and restoring beneficial bacteria, which is one of the core mechanisms by which it alleviates ulcerative colitis.

[0016] Conclusion: Oral administration of water-soluble homogeneous polysaccharide from jujube gum can improve ulcerative colitis induced by sodium dextran sulfate by alleviating bloody stools and promoting body weight. Water-soluble homogeneous polysaccharide from jujube gum effectively alleviates overall intestinal structural damage, mucosal loss, and inflammatory infiltration of colonic tissue induced by sodium dextran sulfate in mice. Water-soluble homogeneous polysaccharide from jujube gum regulates the dysbiosis of the microbial community in mice with sodium dextran sulfate-induced colitis by remodeling the microbial composition. These findings indicate that water-soluble homogeneous polysaccharide from jujube gum has a significant alleviating effect on sodium dextran sulfate-induced ulcerative colitis, which is beneficial for its application in the food and pharmaceutical fields.

Claims

1. A method for preparing water-soluble homogeneous polysaccharides of Elaeagnus angustifolia gum, characterized by, The following steps are carried out: a. Water extraction and alcohol precipitation: The Ziziphus jujuba Mill. gum is prepared into a 0.5%-2% aqueous solution, extracted for 8-12 hours, centrifuged at 5000 rpm for 10 minutes, the supernatant is taken, 95% ethanol is slowly added at a volume ratio of 1:3-5, alcohol precipitation is carried out and overnight, centrifugation is carried out again, the collected precipitate is redissolved with deionized water, dialyzed, freeze-dried to obtain the Ziziphus jujuba Mill. gum water-soluble crude polysaccharide; b. Anion exchange column separation: the Ziziphus jujuba Mill. gum water-soluble crude polysaccharide obtained in step a is prepared into a 2-4 mg / mL solution with pure water, loaded onto a DEAE agarose anion exchange column, eluted with pure water and 0.1-0.8 mol / L NaCl solution gradient, the eluate is tracked and detected by anthrone-sulfuric acid method, the elution curve is drawn, the effective part eluted with water is collected, dialyzed and freeze-dried to obtain the Ziziphus jujuba Mill. gum water-soluble crude polysaccharide, which is separated to obtain the water part polysaccharide; c. Cross-linked Sepharose gel column purification: the water part polysaccharide obtained in step b is prepared into a 5-10 mg / mL solution, loaded onto a cross-linked Sepharose gel column, eluted with water, the eluate is tracked and detected by anthrone-sulfuric acid method, the elution curve is drawn, the peak top end is collected, dialyzed and freeze-dried to obtain the Ziziphus jujuba Mill. gum water-soluble homogeneous polysaccharide.

2. The application of the Ziziphus jujuba Mill. gum water-soluble homogeneous polysaccharide obtained by the method of claim 1 in the preparation of a treatment for ulcerative colitis.

Images