Plant lactobacillus and mannose type-extracellular polysaccharide produced by the plant lactobacillus are applied to relieve colitis

Abstract

This invention discloses a strain of *Lactiplantibacillus plantarum* and its produced mannose-type extracellular polysaccharide in the relief of colitis, belonging to the fields of microbial technology and pharmaceutical technology. The *Lactiplantibacillus plantarum* CCFM 1393 screened in this invention possesses a mannosyltransferase gene and can directionally synthesize CCFM 1393 mannose-type extracellular polysaccharide. When the mannose-type extracellular polysaccharide prepared using *Lactiplantibacillus plantarum* CCFM 1393 of this invention was applied to mice with colitis, it significantly alleviated colitis symptoms, increased intestinal short-chain fatty acid levels, and enhanced intestinal flora species diversity, variability, and the abundance of beneficial *Dubosiella* bacteria. Therefore, *Lactiplantibacillus plantarum* CCFM 1393 and its mannose-type extracellular polysaccharide have great application potential in the preparation of products for the prevention and / or treatment of colitis.

Description

Technical Field

[0001] This invention relates to a strain of *Lactobacillus plantarum* and the application of its produced mannose-type extracellular polysaccharide in relieving colitis, belonging to the fields of microbial technology and pharmaceutical technology. Background Technology

[0002] Ulcerative colitis (UC) is a multifactorial disease, caused by a variety of factors including irregular eating habits, immune dysfunction, and genetic factors. Persistent UC can lead to abdominal pain, diarrhea, rectal bleeding, intestinal colic, and fatigue. Since the occurrence and development of UC are closely related to the high expression of pro-inflammatory cytokines, inflammatory cytokine neutralizing antibodies can be used to treat UC. Researchers have used various biological agents, immunomodulators, and Janus kinase inhibitors to treat UC; however, these drugs may produce side effects, including headache, dizziness, and acute systemic anaphylactic reactions. Studies have shown that extracellular polysaccharides (ECPs), important secondary metabolites of *Lactobacillus plantarum*, can inhibit UC by modulating the host's immune response. Furthermore, *Lactobacillus plantarum* ECPs have fewer toxic side effects and significant therapeutic effects, making them a novel potential alternative for treating UC.

[0003] Extracellular polysaccharides (EPPs) of *Lactobacillus plantarum* are large glycomolecules produced and secreted by *Lactobacillus plantarum* during its growth process, eventually seeping into the culture medium. Based on their location within the cell, PEPs are generally classified into two types: those that adhere to the cell wall and form a capsule, called capsular polysaccharides, and those that enter the culture medium and form mucus, called mucinous polysaccharides. In recent years, research on *Lactobacillus plantarum* PEPs has received widespread attention. Due to the structural diversity of *Lactobacillus plantarum* PEPs, they exhibit excellent antitumor, antioxidant, and antidiabetic activities, as well as remarkable anti-inflammatory capabilities.

[0004] The ability of *Lactobacillus plantarum* extracellular polysaccharides to alleviate ulcerative colitis is closely related to their monosaccharide composition. Mannose-rich extracellular polysaccharides exhibit a strong effect in alleviating ulcerative colitis. Macrophages possess a mannose receptor on their surface, which is highly specific for mannose. Mannose plays an important role in human metabolism, particularly in regulating protein glycosylation, inhibiting glycolysis, and enhancing the effects of chemotherapeutic drugs. Therefore, mannose-rich extracellular polysaccharides possess strong anti-inflammatory capabilities. Further research has found that mannose-rich *Lactobacillus plantarum* extracellular polysaccharides can increase macrophage activity by activating the TLR4-NF-κB signaling pathway. However, in actual production, mannose is extremely rare in *Lactobacillus plantarum* extracellular polysaccharides, making the screening of mannose-containing *Lactobacillus plantarum* extracellular polysaccharides very difficult. However, adding a certain amount of mannose to the culture medium can synthesize GDP-mannose through the mannose pathway (mannose → mannose-6-P → mannose-1-P → GDP-mannose) and activate mannosyltransferases in *Lactobacillus plantarum*, leading to the directed synthesis of mannose-type *Lactobacillus* extracellular polysaccharides. In conclusion, *Lactobacillus plantarum* mannose-type extracellular polysaccharides (MnEPS) are a promising immunomodulator with excellent application prospects.

[0005] Therefore, developing a mannosyl-extracellular polysaccharide from *Lactobacillus plantarum* that can alleviate colitis may be a safe and harmless way to improve colitis-related symptoms. Summary of the Invention

[0006] To address the issues of low production capacity of mannose-containing extracellular polysaccharides by *Lactobacillus plantarum* in existing technologies and the limited application areas of extracellular polysaccharides derived from *Lactobacillus plantarum*, this invention screens a new strain of *Lactobacillus plantarum* (… Lactiplantibacillus plantarum CCFM 1393 was used to produce mannose-type extracellular polysaccharides, and the use of said mannose-type extracellular polysaccharides in alleviating colitis was provided.

[0007] The first objective of this invention is to provide a strain of *Lactobacillus plantarum* (… Lactiplantibacillus plantarum CCFM 1393, the Lactobacillus plantarum ( Lactiplantibacillus plantarum CCFM 1393 has been deposited at the Guangdong Provincial Center for Microbial Culture Collection, with accession number GDMCC No: 64508.

[0008] In one embodiment, the colony characteristics of the *Lactobacillus plantarum* CCFM 1393 are as follows: the colonies are milky white, round and raised, with neat and smooth edges.

[0009] The present invention also provides a composition containing the aforementioned *Lactobacillus plantarum* CCFM 1393.

[0010] The present invention also provides a method for synthesizing mannose-type extracellular polysaccharides using the aforementioned *Lactobacillus plantarum* CCFM 1393.

[0011] In one embodiment, the *Lactobacillus plantarum* CCFM 1393 is cultured in a culture medium at 30-40°C for 12-36 h.

[0012] In one embodiment, the culture medium is an MRS medium containing 5 g / L mannose.

[0013] In one embodiment, the formulation of the ordinary MRS culture medium is as follows: peptone 5-20 g / L, beef extract 5-20 g / L, yeast extract 1-10 g / L, anhydrous glucose 10-30 g / L, anhydrous sodium acetate 1-5 g / L, magnesium sulfate (MgSO4·7H2O) 0.1-2 g / L, manganese sulfate (MnSO4·H2O) 0.1-1 g / L, diammonium hydrogen citrate 1-4 g / L, dipotassium hydrogen phosphate (K2HPO4·3H2O) 1-5 g / L, and Tween 80 0.5-2 mL / L; preferably, it is: peptone 10 g / L, beef extract 10 g / L, yeast extract 5 g / L, anhydrous glucose 20 g / L, anhydrous sodium acetate 2 g / L, magnesium sulfate (MgSO4·7H2O) 0.5 g / L, and manganese sulfate (MnSO4·H2O) 0.25 mL / L. g / L, diammonium hydrogen citrate 2 g / L, dipotassium hydrogen phosphate (K2HPO4·3H2O) 2.6 g / L, Tween 80 1 mL / L.

[0014] In one embodiment, the formulation of the MRS medium containing 5 g / L mannose is as follows: peptone 5-20 g / L, beef extract 5-20 g / L, yeast extract 1-10 g / L, mannose 5-10 g / L, anhydrous glucose 10-30 g / L, anhydrous sodium acetate 1-5 g / L, magnesium sulfate (MgSO4·7H2O) 0.1-2 g / L, manganese sulfate (MnSO4·H2O) 0.1-1 g / L, diammonium hydrogen citrate 1-4 g / L, dipotassium hydrogen phosphate (K2HPO4·3H2O) 1-5 g / L, and Tween 80 0.5-2 mL / L; preferably, peptone 10 g / L, beef extract 10 g / L, yeast extract 5 g / L, mannose 5 g / L, and anhydrous glucose 15 g / L. g / L, anhydrous sodium acetate 2 g / L, magnesium sulfate (MgSO4·7H2O) 0.5 g / L, manganese sulfate (MnSO4·H2O) 0.25 g / L, diammonium hydrogen citrate 2 g / L, dipotassium hydrogen phosphate (K2HPO4·3H2O) 2.6 g / L, Tween 80 1 mL / L.

[0015] In one embodiment, the culture temperature is 37°C.

[0016] In one embodiment, the culture time is 24 hours.

[0017] The present invention also provides a method for producing mannose-type extracellular polysaccharides, wherein the method comprises inoculating the *Lactobacillus plantarum* CCFM 1393 into a fermentation medium, fermenting, and collecting the supernatant.

[0018] In one embodiment, the fermentation medium comprises MRS medium containing 5 g / L mannose.

[0019] In one embodiment, the fermentation temperature is 30-40°C, preferably 37°C.

[0020] In one embodiment, the fermentation time is 12-36 h, preferably 24 h.

[0021] In one embodiment, the inoculation amount of *Lactobacillus plantarum* is 2%-8% (v / v), preferably 4% (v / v).

[0022] In one embodiment, after collecting the supernatant, the supernatant is further subjected to crude separation of non-mannose-type extracellular polysaccharides and mannose-type extracellular polysaccharides.

[0023] In one implementation, the coarse separation step includes:

[0024] (1) Remove proteins from the supernatant;

[0025] (2) Precipitation of polysaccharides in the supernatant obtained in step (1);

[0026] (3) Dialysis step (2) yields polysaccharide, which is mannose-type extracellular polysaccharide.

[0027] In one embodiment, step (1) uses trichloroacetic acid to remove proteins.

[0028] In one embodiment, anhydrous ethanol is used to precipitate polysaccharides in step (2).

[0029] In one embodiment, dialysis is performed in step (3) using a 1000 Da dialysis bag.

[0030] The present invention also provides a mannose-type extracellular polysaccharide prepared using the aforementioned *Lactobacillus plantarum* CCFM 1393.

[0031] In one embodiment, the monosaccharide composition of the mannose-type extracellular polysaccharide is: glucose, galactose, mannose, xylose, glucosamine and galactosamine, wherein the molar percentage of glucose:galactose:mannose:xylose:glucosamine:galactosamine is 24.9:4.9:7:44.3:7.1:11.8.

[0032] The present invention also provides a pharmaceutical composition containing the mannose-type Lactobacillus plantarum extracellular polysaccharide.

[0033] The present invention also provides the use of the aforementioned *Lactobacillus plantarum* CCFM 1393 and / or mannose-type *Lactobacillus plantarum* extracellular polysaccharide in the preparation of medicaments for the prevention and / or treatment of colitis.

[0034] In one embodiment, the prevention and / or treatment of colitis includes at least one of the following (a) to (e):

[0035] (a) Reduce the intestinal disease score index in individuals with colitis;

[0036] (b) Reduce the expression level of pro-inflammatory factors in the colonic tissue of individuals with colitis, said pro-inflammatory factors including at least one of TNF-α, IL-1β, IL-6, IFN-γ or IL-17;

[0037] (c) Increase the expression level of tight junction protein ZO-1 in the colonic tissue of individuals with colitis;

[0038] (d) Reduce inflammatory cell infiltration in the colonic tissue of individuals with colitis, and / or alleviate crypt loss, and / or alleviate goblet cell loss;

[0039] (e) Reduce the number of total macrophages and M1 macrophages in the tissues of individuals with colitis;

[0040] (f) Increase the content of short-chain fatty acids in the intestines of individuals with colitis;

[0041] (g) Enhance the diversity and variability of gut microbiota species and increase the number of beneficial bacteria. Dubosiella Abundance.

[0042] In one embodiment, the short-chain fatty acid includes, but is not limited to, acetic acid and propionic acid.

[0043] This invention also provides the application of the aforementioned *Lactobacillus plantarum* CCFM 1393 and / or mannose-type *Lactobacillus plantarum* extracellular polysaccharide in the preparation of health products that regulate intestinal flora.

[0044] In one implementation, the application specifically involves: enhancing the diversity and variability of gut microbiota species and increasing the number of beneficial bacteria. Dubosiella Abundance.

[0045] Beneficial effects:

[0046] (1) This invention provides a strain of *Lactobacillus plantarum* capable of directionally synthesizing mannose-type extracellular polysaccharides. Lactiplantibacillus plantarum CCFM 1393, when cultured in a mannose-containing medium, was able to synthesize a mannose-type extracellular polysaccharide with the ability to alleviate colitis.

[0047] (2) This invention provides a novel use of mannose-type extracellular polysaccharides produced by *Lactobacillus plantarum* CCFM 1393 in alleviating colitis, specifically in comparison with the DSS group and the blank group:

[0048] (a) The disease activity index score of colitis mice decreased from 7.75±0.46 to 4.88±0.83;

[0049] (b) The colon length of colitis mice increased from 4.86±0.34 cm to 65.89±0.47 cm;

[0050] (c) The infiltration of inflammatory cells in the colon tissue of colitis mice was significantly reduced, and the loss of crypts and goblet cells was alleviated;

[0051] (d) The concentration of the inflammatory factor TNF-α in the colon of colitis mice decreased from 37.53±5.95 pg / mL to 25.37±4.07 pg / mL;

[0052] (e) The concentration of the inflammatory factor IL-1β in the colon of colitis mice decreased from 3103.99±154.6 pg / mL to 1926.22±460.76 pg / mL;

[0053] (f) The concentration of the inflammatory factor IL-6 in the colon of colitis mice decreased from 412.69±143.43 pg / mL to 76.12±35.29 pg / mL, and there was no significant difference in the concentration of the inflammatory factor IL-6 compared with the control group mice;

[0054] (g) The concentration of the inflammatory factor IFN-γ in the colon of colitis mice decreased from 26.35±3.25 pg / mL to 13.44±2.68 pg / mL;

[0055] (h) The concentration of the inflammatory factor IL-17 in the colon of colitis mice decreased from 54.91±10.53 pg / mL to 33.87±14.7 pg / mL;

[0056] (i) The mRNA expression level of ZO-1 in the colon of colitis mice increased from 0.63±0.15 to 1.03±0.34, and there was no significant difference in the mRNA expression level of ZO-1 compared with the blank group mice;

[0057] (j) The concentration of acetic acid in the colonic contents of colitis mice increased from 1.56±0.39 μmol / g to 4.8±1.03 μmol / g; the concentration of propionic acid in the colonic contents of colitis mice increased from 0.28±0.15 μmol / g to 1.58±0.23 μmol / g.

[0058] (k) The total number of colonic macrophages in colitis mice decreased from 29.54±0.79 to 28.31±1.01; the number of M1 macrophages decreased from 10.27±1.06 to 9.3±0.6, and there was no significant difference compared with the control group.

[0059] (l) The Shannon index of the gut microbiota in colitis mice increased from 5.46±0.63 to 6.03±0.45, with no significant difference compared to the control group; the number of species observed increased from 270.57±47.85 to 388.86±53.51, with no significant difference compared to the control group; the β-diversity of gut microbiota was significantly different from that in DSS mice. P <0.05), and enhances beneficial bacteria. Dubosiella Abundance.

[0060] Therefore, *Lactobacillus plantarum* ( Lactiplantibacillus plantarum CCFM 1393 mannose-type extracellular polysaccharide has great application potential in products for the prevention and / or treatment of colitis.

[0061] Preservation of biological materials

[0062] Lactobacillus plantarum ( Lactiplantibacillus plantarum (CCFM 1393, categorized as follows) Lactiplantibacillus plantarum It was deposited on May 15, 2024 at No. 5, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with collection number GDMCC No: 64508. Attached Figure Description

[0063] Figure 1 Lactobacillus plantarum ( Lactiplantibacillus plantarum (CCFM 1393 extracellular polysaccharide gene cluster diagram)

[0064] Figure 2 Lactobacillus plantarum ( Lactiplantibacillus plantarum CCFM 1393 non-mannose-type extracellular polysaccharide molecular weight and monosaccharide composition.

[0065] Figure 3 Lactobacillus plantarum ( Lactiplantibacillus plantarum CCFM 1393 Mannose-type extracellular polysaccharide molecular weight and monosaccharide composition.

[0066] Figure 4Disease activity index scores of experimental mice in different groups.

[0067] Figure 5 Colon length of mice in different experimental groups.

[0068] Figure 6 H&E staining results of colon tissue from different groups of experimental mice.

[0069] Figure 7 : Levels of the inflammatory factor TNF-α in the colon tissue of mice in different experimental groups.

[0070] Figure 8 Levels of the inflammatory factor IL-1β in the colon tissue of mice in different groups.

[0071] Figure 9 Levels of the inflammatory factor IL-6 in the colon tissue of mice in different experimental groups.

[0072] Figure 10 Levels of the inflammatory factor IFN-γ in the colon tissue of mice in different groups.

[0073] Figure 11 Levels of the inflammatory factor IL-17 in the colon tissue of mice in different groups.

[0074] Figure 12 : The mRNA expression level of ZO-1 in the colon tissue of mice in different groups.

[0075] Figure 13 : Levels of short-chain fatty acids (SCFAs) in the colon tissue of mice in different groups.

[0076] Figure 14 The total number of macrophages and M1 macrophages in the colon tissue of mice in different groups.

[0077] Figure 15 α-diversity and β-diversity of gut microbiota in different groups of mice.

[0078] Figure 16 Differences in gut microbiota among different groups of mice.

[0079] In the images above: *, **, ***, and **** represent comparisons between different groups. P Values ​​less than 0.05, 0.01, 0.001, and 0.0001; ns indicates no significant difference compared to each group. Detailed Implementation

[0080] The C57BL / 6 male mice used in the following examples were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. TNF-α, IL-1β, IL-6, IFN-γ, IL-17, and IL-10 ELISA kits were purchased from RD.

[0081] Definition and scoring method of intestinal disease scoring index: The intestinal disease activity index combines the patient's (sick animal's) percentage of weight loss (0 for no change in weight, 1-5 for 1 point, 5-10 for 2 points, 10-15 for 3 points, and greater than 15 for 4 points), stool viscosity (0 for normal, 2 for loose stool, and 4 for diarrhea), and stool bleeding (0 for normal, 2 for occult blood, and 4 for overt bleeding). The total score of the three results is the DAI value.

[0082] The culture media involved in the following examples are as follows:

[0083] Standard MRS liquid culture medium (L): 10 g peptone, 10 g beef extract, 5 g yeast extract, 20 g anhydrous glucose, 2 g anhydrous sodium acetate, 0.5 g magnesium sulfate (MgSO4·7H2O), 0.25 g manganese sulfate (MnSO4·H2O), 2 g diammonium hydrogen citrate, 2.6 g dipotassium hydrogen phosphate (K2HPO4·3H2O), 1 mL Tween 80, pH 6.2-6.4.

[0084] Standard MRS solid medium (L): 10 g peptone, 10 g beef extract, 5 g yeast extract, 20 g anhydrous glucose, 2 g anhydrous sodium acetate, 0.5 g magnesium sulfate (MgSO4·7H2O), 0.25 g manganese sulfate (MnSO4·H2O), 2 g diammonium hydrogen citrate, 2.6 g dipotassium hydrogen phosphate (K2HPO4·3H2O), 1 mL Tween 80, 15 g agar, pH 6.2-6.4.

[0085] MRS liquid medium (L) containing 5 g / L mannose: 10 g peptone, 10 g beef extract, 5 g yeast extract, 5 g mannose, 15 g anhydrous glucose, 2 g anhydrous sodium acetate, 0.5 g magnesium sulfate (MgSO4·7H2O), 0.25 g manganese sulfate (MnSO4·H2O), 2 g diammonium hydrogen citrate, 2.6 g dipotassium hydrogen phosphate (K2HPO4·3H2O), 1 mL Tween 80, pH 6.2-6.4.

[0086] MRS solid medium (L) containing 5 g / L mannose: 10 g peptone, 10 g beef extract, 5 g yeast extract, 5 g mannose, 15 g anhydrous glucose, 2 g anhydrous sodium acetate, 0.5 g magnesium sulfate (MgSO4·7H2O), 0.25 g manganese sulfate (MnSO4·H2O), 2 g diammonium hydrogen citrate, 2.6 g dipotassium hydrogen phosphate (K2HPO4·3H2O), 1 mL Tween 80, 15 g agar, pH 6.2-6.4.

[0087] Example 1: Lactobacillus plantarum ( Lactiplantibacillus plantarum Screening and identification of CCFM 1393

[0088] (1) Lactobacillus plantarum ( Lactiplantibacillus plantarum CCTV 1393 screening:

[0089] Using fecal samples from healthy individuals in Wuxi, Jiangsu Province, one spoonful of the sample was added to 5 mL of PBS (with 0.05% cysteine ​​added), mixed well, and serially diluted. A 10⁻⁶ dilution was selected. -5 ~10 -7 The serially diluted solutions were spread onto the above-mentioned MRS solid medium and incubated at 37 ℃ for 48 h. Typical colonies were picked and streaked onto MRS solid medium for purification, and incubated upside down in a 37 ℃ incubator for 48 h. Single colonies were picked and inoculated into 5 mL of MRS liquid medium, and incubated at 37 ℃ for 16-18 h. 1.5 mL of bacterial culture was centrifuged at 6000 r / min for 3 min to remove the supernatant, and 1 mL of 30% sterile glycerol was added for preservation. At the same time, 1.5 mL of bacterial culture was centrifuged, the supernatant was removed, and the culture was resuspended in sterile water for bacterial identification.

[0090] (2) Identification of strain species

[0091] The resuspended bacterial cells were subjected to genomic DNA extraction according to the FastDNA SPIN Kit for Feces instructions, which was then used as a PCR template. The genomic DNA was then amplified using the 16S rDNA amplification method described in Table 1 below.

[0092] Table 1 16S rDNA amplification system

[0093]

[0094] Sequencing and Analysis: After confirmation by nucleic acid electrophoresis, the PCR products were sent to a sequencing company for sequencing. The obtained assembled sequences were then used for species confirmation via NCBI's BLAST (https: / / blast.ncbi.nlm.nih.gov / Blast.cgi). Based on the results, the purified strain was named *Lactobacillus plantarum* (…). Lactiplantibacillus plantarum (CCFM1393)

[0095] Example 2: Lactobacillus plantarum ( Lactiplantibacillus plantarum Extracellular polysaccharide gene cluster prediction of CCFM 1393

[0096] The *Lactobacillus plantarum* CCFM 1393 screened in Example 1 was cultured in MRS medium, and the DNA was extracted from *Lactobacillus plantarum* using a Fast DNA spin assay kit. Lactiplantibacillus plantarum Genomic DNA was extracted from CCFM 1393. The Illumina Hiseq platform was used to analyze *Lactobacillus plantarum* (Lactobacillus plantarum). Lactiplantibacillus plantarum Paired-end sequencing was performed using a CCFM1393 platform. This platform generated 2 × 150 bp paired-end libraries, providing at least 100 short sequences with genomic coverage for each sample. SOAPdenovo v2.0.4 software was used to assemble contigs from the paired ends de novo, and the assembly results were optimized to form a scaffold based on the overlap relationships between the short sequences. Glimmer was used to predict protein-coding DNA sequences (CDS), which were then translated into CDS for subsequent homologous protein sequence retrieval. The BLASTP function from DAIMOND was used to search for *Lactobacillus plantarum* (…). Lactiplantibacillus plantarum Homologous sequences of the extracellular polysaccharide gene cluster in CCFM 1393 were analyzed and their identity was calculated. The parameters were: identity, 30%; e-value, 1e-15; --ultra-sensitive. The sequence with the highest homology was used to represent *Lactobacillus plantarum* (…). Lactiplantibacillus plantarum Predictive results of extracellular polysaccharide-related synthases from CCFM 1393.

[0097] Lactobacillus plantarum ( Lactiplantibacillus plantarum A notable feature of CCFM 1393 is its ability to produce extracellular polysaccharides. One essential condition for extracellular polysaccharide production is the presence of an extracellular polysaccharide gene cluster. This cluster includes regulatory genes (phosphate regulatory modules EPS B, EPS C, and EPS D), polysaccharide assembly mechanism genes (initiator EPS E, polymerase Wzy, flipper enzyme Wzx, and attachment EPS A), and genes encoding glycosyltransferases required for repeat unit assembly. Results of BlastP prediction of the extracellular polysaccharide gene cluster are shown below. Figure 1 Lactobacillus plantarum (Lactiplantibacillus plantarum CCFM1393 contains 24 genes, including 7 glycosyltransferase genes (glucosyltransferase, galactosyltransferase, rhamnosyltransferase, fructosyltransferase, mannosyltransferase, glucosamine transferase, and galactosamine transferase), of which 19 are forward-stranded and 5 are reverse-stranded. Mannose is a potent immunomodulator that can activate immunity and inhibit glycolysis to reduce host inflammation by binding to the NF-κB pathway of macrophages. Therefore, identifying mannose-containing extracellular polysaccharides is beneficial for the development of functional polysaccharides. In *Lactobacillus plantarum* (… Lactiplantibacillus plantarum CCFM 1393 detected mannosyltransferase, indicating that *Lactobacillus plantarum* (…) Lactiplantibacillus plantarum CCFM 1393 can produce mannosyltransferase, which has the potential to synthesize mannosyl-extracellular polysaccharides.

[0098] Example 3: Lactobacillus plantarum ( Lactiplantibacillus plantarum Cultivation of CCFM 1393 and isolation of non-mannose extracellular polysaccharides

[0099] Lactobacillus plantarum ( Lactiplantibacillus plantarum CCFM 1393 was inoculated into ordinary MRS liquid medium with glucose as the carbon source and cultured at 37°C for 24 h. Then, the bacterial culture was transferred into fresh MRS liquid medium at an inoculation rate of 4% (v / v) and cultured under the same conditions for 24 h.

[0100] After culturing, the cells were removed by centrifugation at 8000 r / min for 20 min, and the fermentation supernatant was collected. 4% (m / v) trichloroacetic acid was added to the supernatant, and the mixture was reacted at 4℃ for 24 h. Then, the mixture was centrifuged at 10000 r / min for 20 min to remove proteins, and the supernatant was collected. Next, 2.5 times the volume of anhydrous ethanol was added to the supernatant, and the mixture was precipitated for 24 h. The precipitate was collected by centrifugation at 10000 r / min for 20 min. The precipitated extracellular polysaccharides were dialyzed using a 1000 Da dialysis bag, with water changed every 6 h, for 3 days. After dialysis, the liquid in the dialysis bag was collected, and the liquid was freeze-dried for 3 days to obtain the dried extracellular polysaccharides.

[0101] HPSEC analysis of Lactobacillus plantarum ( Lactiplantibacillus plantarum The molecular weight of extracellular polysaccharides from CCFM 1393 was analyzed using HPIC (Hydrocotyle vulgaris plantarum). Lactiplantibacillus plantarum The extracellular polysaccharide monosaccharide composition of CCFM 1393. (e.g.) Figure 2 As shown in A, the molecular weight of CCFM 1393 non-mannose-type extracellular polysaccharide consists of two parts: 97920 Da (35.38%) and 4670 Da (64.62%). Figure 2In B, numbers 1-10 represent fucose, galactosamine, rhamnose, arabinose, glucosamine, galactose, glucose, xylose, mannose, and fructose, respectively. For example... Figure 2 As shown in B, the monosaccharide composition of CCFM 1393 non-mannose-type extracellular polysaccharide is: glucose: galactose: fructose: arabinose: glucosamine: galactosamine, with molar percentages of 17.8%: 8.1%: 61.1%: 4.1%: 2.9%: 6.1%.

[0102] Analysis revealed that the target mannose was not expressed in the extracellular polysaccharide, and the gene expression of *Lactobacillus plantarum* was affected by the culture environment, with the structure of its extracellular polysaccharide varying with culture conditions. Studies have shown that GDP-mannose is a precursor to mannose with multiple synthetic pathways, synthesized via the mannose pathway (mannose → mannose-6-P → mannose-1-P → GDP-mannose). In standard MRS medium (with glucose as the sole carbon source), GDP-mannose synthesis follows a single pathway. Glucose-6-P is converted to fructose-6-P, which is then catalyzed by mannose-6-phosphate isomerase, phosphogmannanase, and mannose-1-phosphate guanylate transferase to synthesize GDP-mannose. However, adding a certain proportion of mannose to the culture medium can activate the mannose pathway, stimulating *Lactobacillus plantarum* to synthesize mannose-type extracellular polysaccharides. Based on this background, this patent proposes adding a certain proportion of mannose to standard MRS medium to stimulate *Lactobacillus plantarum* (… Lactiplantibacillus plantarum CCFM 1393 synthesizes mannose-type extracellular polysaccharides.

[0103] Example 4: Lactobacillus plantarum ( Lactiplantibacillus plantarum Cultivation of CCFM 1393 and isolation of mannose-type extracellular polysaccharides

[0104] Lactobacillus plantarum ( Lactiplantibacillus plantarum CCFM 1393 was inoculated into MRS liquid medium containing 5 g / L mannose and cultured at 37°C for 24 h. Then, the bacterial culture was transferred into fresh MRS liquid medium containing 5 g / L mannose at an inoculation rate of 4% (v / v) and cultured under the same conditions for 24 h.

[0105] After culturing, the cells were removed by centrifugation at 8000 r / min for 20 min, and the fermentation supernatant was collected. 4% (m / v) trichloroacetic acid was added to the supernatant, and the mixture was reacted at 4℃ for 24 h. Then, the mixture was centrifuged at 10000 r / min for 20 min to remove proteins, and the supernatant was collected. Next, 2.5 times the volume of anhydrous ethanol was added to the supernatant, and the mixture was precipitated for 24 h. The precipitate was collected by centrifugation at 10000 r / min for 20 min. The precipitated extracellular polysaccharides were dialyzed using a 1000 Da dialysis bag, with water changed every 6 h, for 3 days. After dialysis, the liquid in the dialysis bag was collected, and the liquid was freeze-dried for 3 days to obtain the dried extracellular polysaccharides.

[0106] HPSEC analysis of Lactobacillus plantarum ( Lactiplantibacillus plantarum The molecular weight of mannose-type extracellular polysaccharides (CCFM 1393) was analyzed using HPIC (Hydrogenase Insectidae) and Lactobacillus plantarum was determined. Lactiplantibacillus plantarum CCFM 1393 is composed of mannose-type extracellular polysaccharides and monosaccharides. (For example...) Figure 3 As shown in A, the molecular weight of CCFM 1393 mannose-type extracellular polysaccharide is 40951 Da (100%). Figure 3 In section B, numbers 1-10 represent fucose, galactosamine, rhamnose, arabinose, glucosamine, galactose, glucose, xylose, mannose, and fructose, respectively. The calculated monosaccharide composition of the mannose-type extracellular polysaccharide in CCFM 1393 is: glucose:galactose:mannose:xylose:glucosamine:galactosamine, with molar percentages of 24.9%:4.9%:7%:44.3%:7.1%:11.8%. Analysis of the results showed that adding mannose to the culture medium can promote the growth of *Lactobacillus plantarum* (…). Lactiplantibacillus plantarum Expression of mannosyltransferase in CCFM 1393 to synthesize CCFM 1393 mannose-type extracellular polysaccharide.

[0107] Example 5: Lactobacillus plantarum ( Lactiplantibacillus plantarum Effects of CCFM 1393 mannose-type extracellular polysaccharide on the disease activity index score in colitis mice

[0108] Four-week-old SPF-grade male C57BL / 6J mice were randomly divided into three groups: a control group, a DSS group, and an experimental group. The experimental group was administered 200 mg / kg of *Lactobacillus plantarum* via gavage. Lactiplantibacillus plantarum CCFM 1393 mannose-extracellular polysaccharide group. Eight animals per group were housed in the Experimental Animal Center of Jiangnan University at a constant temperature of 21-26℃, humidity of 40-70%, noise level of less than or equal to 60 dB, and animal illumination of 15-20 LX (all animal experimental procedures were reviewed and approved by the Animal Welfare and Ethics Management Committee of Jiangnan University).

[0109] The experiment lasted a total of 15 days.

[0110] Days 1-7 are the adaptation period. During this period, each cage of mice has free access to water and is given a normal growth and reproduction diet.

[0111] After the adaptation period ends, days 8-15:

[0112] DSS group: 3% DSS drinking water was given daily;

[0113] CCFM 1393 mannose-extracellular polysaccharide group: administered 3% DSS drinking water daily + 0.2 mL of *Lactobacillus plantarum* via gavage. Lactiplantibacillus plantarum CCFM 1393 Mannose-type extracellular polysaccharide; the oral dose of mannose-type extracellular polysaccharide is 200 mg / kg;

[0114] Control group: Provided with normal drinking water daily.

[0115] On the last day of the experiment, the disease activity index score and body weight of the mice were measured. The final disease activity index score and body weight are as follows: Figure 4 As shown.

[0116] like Figure 4 As shown, after 7 days of drinking water containing 3% DSS, the disease activity index score of the DSS group increased to 7.75±0.46. In contrast, the CCFM 1393 mannose-extracellular polysaccharide group, which also drank 3% DSS-containing water for 7 days, had a disease activity index score of 4.88±0.83, a decrease of 37.03% compared to the DSS group. Therefore, *Lactobacillus plantarum* CCFM 1393 mannose-extracellular polysaccharide has the effect of reducing the disease activity index score of colitis in mice.

[0117] Example 6: Lactobacillus plantarum ( Lactiplantibacillus plantarum Effects of CCFM 1393 mannose-type extracellular polysaccharide on colon length in colitis mice

[0118] The method for constructing the animal model is the same as in Example 5.

[0119] After the experiment, the mice were dissected in accordance with ethical requirements, and their colons were taken and their length measured.

[0120] Colon length as Figure 5 As shown, compared to the control group (7.56±0.54 cm), the colon length in the DSS group was significantly reduced to 4.86±0.34 cm. With the intervention of CCFM 1393 mannose-type extracellular polysaccharide, the colon length significantly increased to 5.89±0.47 cm, a 21.19% increase compared to the DSS group. Therefore, CCFM 1393 mannose-type extracellular polysaccharide can effectively slow down the shortening of colon length in mice.

[0121] Example 7: Lactobacillus plantarum ( Lactiplantibacillus plantarum Effects of CCFM 1393 mannose-type extracellular polysaccharide on inflammatory cell infiltration and crypt and goblet cell morphology in colitis mice

[0122] The method for constructing the animal model is the same as in Example 5.

[0123] After the experiment, the mice were dissected in accordance with ethical requirements. A 1 cm distal end of the colon was taken and fixed in 4% paraformaldehyde solution for 36 h. After being dehydrated by ethanol gradient, cleared with xylene, embedded in paraffin, stained with H&E, and the sections were scanned to observe the cross-sectional condition of the colon.

[0124] Colon slices Figure 6 As shown, the control group mice had normal crypts, uniformly distributed goblet cells, and no inflammatory cell infiltration. The DSS group mice showed crypt loss, partial loss of goblet cells, depletion of mucin, and severe inflammatory cell infiltration. Compared to the DSS group, mice treated with CCFM 1393 mannose-extracellular polysaccharide showed restored colonic crypts, restored goblet cell numbers, increased mucin, and almost no inflammatory cell infiltration, significantly alleviating the colonic pathology in these mice. Therefore, CCFM 1393 mannose-extracellular polysaccharide can effectively slow the progression of histopathological changes in mice with colitis.

[0125] Example 8: Lactobacillus plantarum ( Lactiplantibacillus plantarum Effects of CCFM 1393 mannose-type extracellular polysaccharide on the level of TNF-α, a pro-inflammatory factor in colitis, in mice.

[0126] The method for constructing the animal model is the same as in Example 5.

[0127] After the experiment, the mice were dissected in accordance with ethical requirements. Colonic tissue was weighed and added to an appropriate volume of a mixture of RIPA lysis buffer and a protease phosphatase inhibitor. The tissue was then ground, and the supernatant was extracted. The concentration of TNF-α in the colonic tissue was determined according to the ELISA kit instructions.

[0128] TNF-α concentration in colon tissue is shown in Figure 7 .Depend on Figure 7 Quantitative results showed that the TNF-α concentration in the blank group was 4.36±1.27 pg / mL, while the TNF-α concentration in the DSS group was 37.53±5.95 pg / mL. The TNF-α concentration was significantly reduced to 25.37±4.07 pg / mL under CCFM 1393 mannose-type extracellular polysaccharide intervention, a decrease of 32.40% compared to the DSS group. Therefore, CCFM1393 mannose-type extracellular polysaccharide can effectively inhibit TNF-α levels in colonic tissue and alleviate colonic inflammation.

[0129] Example 9: Lactobacillus plantarum ( Lactiplantibacillus plantarum Effects of CCFM 1393 mannose-type extracellular polysaccharide on the level of IL-1β, a pro-inflammatory factor in colitis, in mice.

[0130] The method for constructing the animal model is the same as in Example 5.

[0131] After the experiment, the mice were dissected in accordance with ethical requirements. A measured amount of colon tissue was taken and mixed with an appropriate volume of RIPA lysis buffer and a mixture of protease phosphatase inhibitors. The tissue was then ground, and the supernatant was extracted. The concentration of IL-1β in the colon tissue was determined according to the ELISA kit instructions.

[0132] Colon tissue IL-1β concentrations are shown in the figure. Figure 8 .Depend on Figure 8 Quantitative results showed that the IL-1β concentration in the blank group was 42.09±11.51 pg / mL, while the IL-1β concentration in the DSS group was 3103.99±154.6 pg / mL. The IL-1β concentration was significantly reduced to 1926.22±460.76 pg / mL under CCFM 1393 mannose-extracellular polysaccharide intervention, a decrease of 38.58% compared to the DSS group. Therefore, CCFM 1393 mannose-extracellular polysaccharide can effectively inhibit IL-1β levels in colonic tissue.

[0133] Example 10: Lactobacillus plantarum ( Lactiplantibacillus plantarum Effects of CCFM 1393 mannose-type extracellular polysaccharide on the level of IL-6, a pro-inflammatory factor in colitis, in mice.

[0134] The method for constructing the animal model is the same as in Example 5.

[0135] After the experiment, the mice were dissected in accordance with ethical requirements. Colonic tissue was weighed and added to an appropriate volume of a mixture of RIPA lysis buffer and a protease phosphatase inhibitor. The tissue was then ground, and the supernatant was extracted. The concentration of IL-6 in the colonic tissue was determined according to the ELISA kit instructions.

[0136] Colon tissue IL-6 concentrations are shown in Figure 9 .Depend on Figure 9 Quantitative results showed that the IL-1β concentration in the control group was 8.56±3.43 pg / mL, while the IL-6 concentration in the DSS group was 412.69±143.43 pg / mL. The IL-6 concentration was significantly reduced to 76.12±35.29 pg / mL under CCFM 1393 mannose-extracellular polysaccharide intervention, a decrease of 81.39% compared to the DSS group, and there was no significant difference compared to the control group. Therefore, CCFM 1393 mannose-extracellular polysaccharide can effectively inhibit IL-6 levels in colonic tissue.

[0137] Example 11: Lactobacillus plantarum ( Lactiplantibacillus plantarum Effects of CCFM 1393 mannose-type extracellular polysaccharide on the pro-inflammatory factor IFN-γ in colitis mice

[0138] The method for constructing the animal model is the same as in Example 5.

[0139] After the experiment, the mice were dissected in accordance with ethical requirements. A measured amount of colon tissue was taken and mixed with an appropriate volume of RIPA lysis buffer and a mixture of protease phosphatase inhibitors. The tissue was then ground, and the supernatant was extracted. The concentration of IFN-γ in the colon tissue was determined according to the ELISA kit instructions.

[0140] The concentration of IFN-γ in colon tissue is shown in the figure. Figure 10 .Depend on Figure 10 Quantitative results showed that the IFN-γ concentration in the blank group was 7.79±1.4 pg / mL, while the IFN-γ concentration in the DSS group was 26.35±3.25 pg / mL. The IFN-γ concentration was significantly reduced to 13.44±2.68 pg / mL under CCFM 1393 mannose-type extracellular polysaccharide intervention, a decrease of 49.00% compared to the DSS group. Therefore, CCFM 1393 mannose-type extracellular polysaccharide can effectively inhibit the IFN-γ level in colonic tissue.

[0141] Example 12: Lactobacillus plantarum ( Lactiplantibacillus plantarum Effects of CCFM 1393 mannose-type extracellular polysaccharide on the pro-inflammatory factor IL-17 in colitis mice

[0142] The method for constructing the animal model is the same as in Example 5.

[0143] After the experiment, the mice were dissected in accordance with ethical requirements. Colonic tissue was weighed and mixed with an appropriate volume of RIPA lysis buffer and a mixture of protease phosphatase inhibitors. The tissue was then ground, and the supernatant was extracted. The concentration of IL-17 in the colonic tissue was determined according to the ELISA kit instructions.

[0144] Colon tissue IL-17 concentrations are shown in the figure. Figure 11 .Depend on Figure 11 Quantitative results showed that the IL-17 concentration in the blank group was 5.9±2.13 pg / mL, while the IL-17 concentration in the DSS group was 54.91±10.53 pg / mL. The IL-17 concentration was significantly reduced to 33.87±14.7 pg / mL under CCFM 1393 mannose-extracellular polysaccharide intervention, a decrease of 38.32% compared to the DSS group. Therefore, CCFM1393 mannose-extracellular polysaccharide can effectively inhibit IL-17 levels in colonic tissue.

[0145] Example 13: Lactobacillus plantarum ( Lactiplantibacillus plantarum Effects of CCFM 1393 mannose-type extracellular polysaccharide on tight junction protein ZO-1 in colonic tissue of colitis mice

[0146] The method for constructing the animal model is the same as in Example 5.

[0147] After the experiment, mice were dissected according to ethical requirements. Colon tissue was weighed and extracted using FreeZol reagent for total RNA extraction and reverse transcription. The relative cDNA content was analyzed using a ChamQ SYBR qPCR Master Mix quantitative real-time PCR thermal cycler. (The last sentence appears to be incomplete and requires further context.) -ΔΔCT Calculate the ZO-1 content standardized by GAPDH.

[0148] Tight junction proteins are an important component of the intestinal barrier, and ZO-1 is currently the most studied tight junction protein. The mRNA expression level of ZO-1 in colonic tissue is shown in [reference needed]. Figure 12 .Depend on Figure 12 Quantitative results showed that the mRNA expression level of ZO-1 in the DSS group was 0.63±0.15, while the mRNA expression level of ZO-1 under CCFM1334 extracellular polysaccharide intervention was significantly increased to 1.03±0.34, which was 63.49% higher than that in the DSS group, and there was no significant difference compared with the blank group. Therefore, CCFM 1393 mannose-type extracellular polysaccharide can effectively promote the mRNA expression level of the tight junction protein ZO-1 in colon tissue.

[0149] Example 14: Lactobacillus plantarum ( Lactiplantibacillus plantarum Effects of CCFM 1393 mannose-type extracellular polysaccharide on short-chain fatty acids (SCFAs) in the colon of colitis mice

[0150] The method for constructing the animal model is the same as in Example 5.

[0151] After the experiment, the mice were dissected according to ethical requirements. The contents of the mouse colon were weighed and added to 500 μL of saturated NaCl solution. After homogenization, 1 mL of anhydrous diethyl ether was added to extract short-chain fatty acids. The supernatant was then treated with 0.5 g of anhydrous sodium sulfate to remove excess water. The content of short-chain fatty acids in the colon contents was determined using GC-MS.

[0152] Short-chain fatty acids (SCFAs) are effective anti-inflammatory agents and have been shown to alleviate the development of colitis by modulating gut microbiota, influencing host metabolism, and repairing the intestinal barrier. The content of SCFAs in colonic contents is shown in [link to relevant data]. Figure 13 .Depend on Figure 13Quantitative results for A showed that the acetic acid concentration in the blank group was 3.38 ± 0.64 μmol / g, while the acetic acid concentration in the DSS group was 1.56 ± 0.39 μmol / g. Under the intervention of CCFM 1393 mannose-type extracellular polysaccharide, the acetic acid concentration significantly increased to 4.8 ± 1.03 μmol / g, an increase of 207.69% compared to the DSS group and 42.01% compared to the blank group. Figure 13 Quantitative results from B showed that the propionic acid concentration in the blank group was 0.84±0.15 μmol / g, while the propionic acid concentration in the DSS group was 0.28±0.15 μmol / g. Under the intervention of CCFM1393 mannose-type extracellular polysaccharide, the propionic acid concentration significantly increased to 1.58±0.23 μmol / g, an increase of 464.29% compared to the DSS group and 88.10% compared to the blank group. Therefore, CCFM 1393 mannose-type extracellular polysaccharide can effectively increase the content of short-chain fatty acids in colonic contents. Increased short-chain fatty acid concentration can help regulate the intestinal flora, regulate host metabolism, and repair the intestinal barrier in colitis mice to alleviate colitis.

[0153] Example 15: Lactobacillus plantarum ( Lactiplantibacillus plantarum Effects of CCFM 1393 mannose-type extracellular polysaccharide on colonic macrophage infiltration and morphology in colitis mice

[0154] The method for constructing the animal model is the same as in Example 5.

[0155] After the experiment, mice were dissected according to ethical requirements. Colon tissue was fixed in 4% paraformaldehyde solution, dehydrated, embedded, and sectioned. The sections were then placed in sodium citrate buffer (pH 6.0, 0.01 mol / L) and blocked with goat serum for 1 h. Samples were incubated with the primary monoclonal antibody (iNOS, F4 / 80) and the secondary antibody, and DAPI was added to the samples. Sections were sealed with 50% glycerol. The expression of iNOS and F4 / 80 was detected using confocal laser scanning microscopy. ImageJ was used to count the number of F4 / 80+ and iNOS+ cells.

[0156] Infiltration of macrophages in the lamina propria and M1 macrophages exacerbates colitis in mice. Using F4 / 80 + Cells replace total macrophages in the lamina propria, iNOS + Cells replace M1 macrophages. (By...) Figure 14Quantitative results for A showed that the total number of macrophages in the blank group was 25.26 ± 0.86, while the total number of macrophages in the DSS group was 29.54 ± 0.79. Under CCFM 1393 mannose-type extracellular polysaccharide intervention, the total number of macrophages was 28.31 ± 1.01, a decrease of 1.28 compared to the DSS group. Figure 14 Quantitative results from B showed that the number of M1 macrophages in the control group was 9.05±0.39, while the number of M1 macrophages in the DSS group was 10.27±1.06. Under CCFM 1393 mannose-extracellular polysaccharide intervention, the number of M1 macrophages was 9.3±0.6, a decrease of 0.97 compared to the DSS group, and there was no significant difference compared to the control group. Therefore, CCFM 1393 mannose-extracellular polysaccharide can effectively reduce the number of total colonic macrophages and M1 macrophages.

[0157] Example 16: Lactobacillus plantarum ( Lactiplantibacillus plantarum Effects of CCFM 1393 mannose-type extracellular polysaccharide on gut microbiota in colitis mice

[0158] The method for constructing the animal model is the same as in Example 5.

[0159] After the experiment, mice were dissected according to ethical requirements, and the colon contents were stored at -80°C. Total DNA was isolated from mouse feces using a rapid DNA spin kit. The 16S rRNA V3-V4 region was amplified using 341F / 806R PCR. The purified DNA was sequenced using the Illumina MiSeq platform. The raw sequences were processed using the Qiime2 platform. α-diversity was represented by the Shannon's Diversity Index (SHDI) and the number of species observations; β-diversity was based on the Bray-Curtis distance and displayed using principal component analysis. Multivariate analysis of permutation variance was used to calculate the significance of clustering differences in the PcoA community. Species differences were screened using linear discriminant analysis (LDA Effect Size, LEFSE) (Wilcoxon rank-sum test; LDA score > 2.0, P < 0.05). An interaction network was constructed using Spearman correlation coefficients.

[0160] Gut microbiota dysbiosis is a crucial factor in the pathogenesis of colitis. Mannose-type extracellular polysaccharides are promising prebiotic candidates for alleviating colitis, possessing the potential to modulate the gut microbiota. Figure 15As shown in Figure A, the Shannon index in the control group was 6.27±0.44, and in the DSS group it was 5.46±0.63. The Shannon index in mice treated with CCFM 1393 mannose-type extracellular polysaccharides was 6.03±0.45, which was 10.44% higher than the DSS group, and there was no significant difference compared to the control group. Figure 15 As shown in Figure B, the number of species observed in the control group was 407.5±53.74, while that in the DSS group was 270.57±47.85. The number of species observed in mice treated with CCFM1393 mannose-extracellular polysaccharide was 388.86±53.51, which was 43.72% higher than that in the DSS group, and there was no significant difference compared to the control group. Both the aroma index and the number of species observed indicate that CCFM 1393 mannose-extracellular polysaccharide significantly improved the bacterial diversity of DSS mice.

[0161] β-diversity revealed the differences in microbial communities among sample groups. For example... Figure 15 As shown in C, there was a significant difference in β-diversity between the blank group mice and the DSS group mice. P <0.05), indicating that DSS-induced colitis alters the distribution of gut microbiota in mice. Subsequently, as... Figure 15 As shown in D, there were significant differences in gut microbiota between CCFM 1393 mannose-extracellular polysaccharide-treated mice and DSS group mice. P The value <0.05 indicates that CCFM 1393 mannose-type extracellular polysaccharide can alter the species composition of the mouse gut microbiota.

[0162] To analyze the differences in gut microbiota between DSS group mice and CCFM 1393 mannose-extracellular polysaccharide group mice, LEFSE analysis was used. Figure 16 As shown, CCFM 1393 mannose-type extracellular polysaccharide can improve... Dubosiella Abundance. Dubosiella It can alleviate colonic inflammation in mice by increasing the level of the anti-inflammatory factor IL-10 and decreasing the concentrations of inflammatory cytokines IL-1β, IL-6, and TNF-α through butyrate production, thus being a beneficial bacterium. In conclusion, CCFM 1393 mannose-type extracellular polysaccharide can enhance the species diversity and variability of the gut microbiota in DSS mice and increase the number of beneficial bacteria. Dubosiella Abundance can alleviate colitis in mice.

[0163] Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Anyone skilled in the art can make various modifications and alterations without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention should be determined by the claims.

Claims

1. Lactobacillus plantarum ( Lactiplantibacillus plantarum CCFM 1393, characterized in that, It is deposited at the Guangdong Provincial Center for Microbial Culture Collection, with accession number GDMCC No: 64508.

2. A composition containing the *Lactobacillus plantarum* CCFM 1393 as described in claim 1.

3. A method for synthesizing mannose-type extracellular polysaccharides, characterized in that, The *Lactobacillus plantarum* CCFM 1393 described in claim 1 was cultured in a culture medium at 30-40°C for 12-36 h.

4. A pharmaceutical composition, characterized in that, Contains the mannose-type extracellular polysaccharide prepared by the method of claim 3.

5. The pharmaceutical composition according to claim 4, characterized in that, It also contains drug carriers and / or drug excipients.

6. The use of the *Lactobacillus plantarum* CCFM 1393 as described in claim 1 and / or the mannose-type extracellular polysaccharide prepared by the method described in claim 3 in the preparation of medicaments for the prevention and / or treatment of colitis.

7. The application according to claim 6, characterized in that, The prevention and / or treatment of colitis includes at least one of the following (a) to (e): (a) Reduce the intestinal disease score index in individuals with colitis; (b) Reduce the expression levels of pro-inflammatory factors in the colonic tissue of individuals with colitis; (c) Increase the expression level of tight junction protein ZO-1 in the colonic tissue of individuals with colitis; (d) Reduce inflammatory cell infiltration in the colonic tissue of individuals with colitis, and / or alleviate crypt loss, and / or alleviate goblet cell loss; (e) Reduce the number of total macrophages and M1 macrophages in the tissues of individuals with colitis; (f) Increase the content of short-chain fatty acids in the intestines of individuals with colitis; (g) Enhance the diversity and variability of gut microbiota species and increase the number of beneficial bacteria. Dubosiella Abundance.

8. A health product containing *Lactobacillus plantarum* CCFM 1393 as described in claim 1 and / or mannose-type extracellular polysaccharides prepared by the method described in claim 3.

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