Black fungus degraded polysaccharide as well as preparation method and application thereof

Abstract

The invention discloses a black fungus degraded polysaccharide, a preparation method and application of the black fungus degraded polysaccharide in preparation of products for enhancing immune function and / or immunoregulation, monosaccharide of the black fungus degraded polysaccharide comprises mannose and glucuronic acid, and the molar ratio of the mannose to the glucuronic acid is 9.46: 1. The auricularia auricula-judae degraded polysaccharide disclosed by the invention has relatively low molecular weight and viscosity, the biological activity is remarkably improved, and experiments prove that the auricularia auricula-judae degraded polysaccharide can be used for increasing the thymus index, increasing the spleen index, relieving oxidative stress injury, increasing the content of cell factors, reducing intestinal tissue injury and regulating the intestinal flora structure; and safe and reliable active ingredients which can be taken for a long time are provided for development of immune function enhancing and / or immune regulation products.

Description

Technical Field

[0001] The invention belongs to the technical field of medicines, and in particular relates to a black fungus degraded polysaccharide and a preparation method and application thereof. Background Art

[0002] Immunomodulation is an important system for the body to maintain physiological balance and stability, and to maintain health and vitality. Cyclophosphamide, as a drug for the treatment of cancer and other diseases, plays an important role, but it has side effects that damage organs such as the intestines, causing digestive tract diseases and subsequently causing low immunity. Previous studies have shown that immunomodulatory drugs or dietary interventions have a profound effect on human immune regulation, by improving the host's intestinal function, creating a good intestinal homeostasis, and preventing low immunity.

[0003] Black fungus (Auricularia auricula), also known as black vegetable, fungus, and cloud fungus, belongs to the genus Auricularia of the family Auriculariaceae. It is a precious colloid fungus used for both medicine and food in my country, and is also recognized as a health food in the world. Black fungus is rich in nutrients and is known as the "king of vegetarians" and the "black treasure of Chinese food." The content of protein and iron in its dried product is relatively high, with a protein content of 20g / 100g and an iron content of 97.4mg / 100g. In addition, black fungus is also rich in calcium, phosphorus, carotene, and vitamin B 1 , Vitamin B 2 , cellulose and other ingredients. Black fungus polysaccharide is considered to be the most important biological component in black fungus. With the continuous deepening of modern pharmacological research, it has been confirmed that it has biological activities such as anti-oxidation, anti-coagulation, anti-tumor, hypoglycemic, immune regulation, lipid-lowering, antibacterial and anti-radiation, and has broad development prospects in the fields of food and medicine. However, natural black fungus polysaccharide has a large molecular weight and high viscosity, resulting in low utilization rate. Therefore, it is urgent to explore new methods for modifying black fungus polysaccharide, discover highly effective active substances, and then expand the application of black fungus polysaccharide. Summary of the invention

[0004] In order to solve the above technical problems, the present invention provides a novel black fungus degraded polysaccharide, a preparation method and its application in the preparation of products for enhancing immune function and / or immunomodulation. Experimental verification shows that the black fungus degraded polysaccharide provided by the present invention can increase the thymus index, increase the spleen index, reduce oxidative stress damage, increase the cytokine content, reduce intestinal tissue damage, and regulate the intestinal flora structure, providing a safe, reliable, and long-term active ingredient for the development of products for enhancing immune function and / or immunomodulation.

[0005] In a first aspect of the present invention, the present invention provides a degraded black fungus polysaccharide, wherein the monosaccharide composition of the degraded black fungus polysaccharide comprises mannose and glucuronic acid, and the molar ratio of mannose to glucuronic acid is 9.46:1.

[0006] In some embodiments of the present invention, the molecular weight of the degraded black fungus polysaccharide is 83.56 kDa.

[0007] In some embodiments of the present invention, the protein concentration of the degraded black fungus polysaccharide is 7.51±0.21%, and the uronic acid concentration is 27.07±5.51%.

[0008] In a second aspect of the present invention, the present invention provides a method for preparing black fungus degraded polysaccharide, the preparation method comprising the following steps:

[0009] S1, drying and crushing black fungus, extracting with hot water to obtain a polysaccharide extract, concentrating the polysaccharide extract, and sequentially subjecting the extract to alcohol precipitation, deproteinization, dialysis and drying to obtain black fungus crude polysaccharide;

[0010] S2. Dissolving the black fungus crude polysaccharide in water to form a black fungus crude polysaccharide suspension, adding hydrogen peroxide and ascorbic acid to the black fungus crude polysaccharide suspension to form a reaction system, reacting the reaction system at 40° C. to 60° C. for 1 h to 3 h, and then concentrating, dialyzing and drying in sequence to obtain the black fungus degraded polysaccharide.

[0011] In some embodiments of the present invention, in step S1, the extraction temperature of hot water extraction is 95°C to 100°C.

[0012] In some embodiments of the present invention, the extraction time of hot water extraction is 1 h to 2.5 h.

[0013] In some embodiments of the present invention, the deproteinization uses a sevage reagent, which is a mixed solution of n-butanol and chloroform in a volume ratio of 1:4.

[0014] In some embodiments of the present invention, in step S1, the dialysis time is 72±2h.

[0015] In some embodiments of the present invention, in step S2, the solid-liquid ratio of the black fungus crude polysaccharide suspension is 1:(30-35) mg / mL.

[0016] In some embodiments of the present invention, in step S2, the concentration of hydrogen peroxide is 3-100 mM, and the concentration of ascorbic acid is 3-100 mM.

[0017] In some embodiments of the present invention, in step S2, the molar concentration ratio of the hydrogen peroxide to the ascorbic acid is 1:1.

[0018] In some embodiments of the present invention, in step S2, the dialysis uses a 3500Da dialysis bag and the dialysis time is 72±2h.

[0019] In the third aspect of the present invention, the present invention provides the use of the degraded black fungus polysaccharide described in the first aspect or the degraded black fungus polysaccharide prepared by the preparation method described in the second aspect in the preparation of products enhancing immune function and / or immunomodulation.

[0020] In some embodiments of the present invention, the immunity enhancement or immune regulation includes: restoring body weight, increasing thymus index, increasing spleen index, reducing oxidative stress damage, increasing cytokine content, reducing intestinal tissue damage, and regulating intestinal flora.

[0021] In some embodiments of the present invention, the cytokines include: IL-6, IL-10, IL-17, IL-1β, TNF-α.

[0022] In some embodiments of the present invention, regulating the intestinal flora includes: restoring the number of OTU units and regulating the abundance of the intestinal flora.

[0023] In some embodiments of the present invention, the regulating the abundance of intestinal flora includes: inhibiting the decrease in the relative abundance of Bacteroidota and Campilobacterota caused by cyclophosphamide; inhibiting the increase in the relative abundance of Firmicutes, Actinobacteriota and Desulfobacterota caused by cyclophosphamide; increasing the relative abundance of Eggerthellaceae, Coriobacteriales-Incertae-Sedis, and Desulfovibrionaceae; inhibiting the increase in the relative abundance of UCG-010, Bacillus, and Rikenella caused by cyclophosphamide.

[0024] In some embodiments of the present invention, the product is at least one of a medicine, a health product and a functional food.

[0025] In some embodiments of the present invention, the dosage form of the drug is granules, powders, tablets, capsules, oral liquids or pills.

[0026] Beneficial effects:

[0027] 1. The present invention separates degraded polysaccharides from black fungus, analyzes and identifies the molecular weight, monosaccharide composition, and chemical structure, and determines the molecular weight and structural composition. Experimental verification shows that the degraded polysaccharide has good immunomodulation and intestinal homeostasis maintenance capabilities, and can significantly improve cyclophosphamide-induced immunodeficiency and maintain intestinal homeostasis.

[0028] 2. The degradation method of the present invention has the advantages of being cheap and low in ecological consumption, and is suitable for large-scale industrial controlled degradation of crude polysaccharides from black fungus. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 This is the effect of shear rate on viscosity of black fungus crude polysaccharide before and after degradation.

[0030] Figure 2 The changes in molecular weight of black fungus crude polysaccharide before and after degradation.

[0031] Figure 3 The changes in monosaccharide composition of black fungus crude polysaccharides before and after degradation.

[0032] Figure 4 The effects of black fungus polysaccharides on the body weight and organ indexes of immunodeficient mice induced by cyclophosphamide. Among them, (A) mouse feeding flow chart, (B) the effects of crude black fungus polysaccharides and degraded black fungus polysaccharides on the body weight of immunodeficient mice, (C) the effects of crude black fungus polysaccharides and degraded black fungus polysaccharides on the thymus index of immunodeficient mice, (D) the effects of crude black fungus polysaccharides and degraded black fungus polysaccharides on the spleen index of immunodeficient mice.

[0033] Figure 5 The effects of black fungus polysaccharides on the antioxidant and inflammatory responses of immunodeficient mice induced by cyclophosphamide; among them, (AB) are the effects of crude black fungus polysaccharides and degraded black fungus polysaccharides on the antioxidant indicators of immunodeficient mice, and (CD) are the effects of crude black fungus polysaccharides and degraded black fungus polysaccharides on IL-1β and TNF-α of immunodeficient mice.

[0034] Figure 6 To investigate the effects of crude and degraded polysaccharides from Auricularia auricularia on the intestinal morphology and structure in cyclophosphamide-induced immunodeficient mice.

[0035] Figure 7 The effects of crude polysaccharides and degraded polysaccharides from black fungus on the intestinal flora of cyclophosphamide-induced immunodeficient mice, including (A) Venn diagram, (B) petal diagram, (C) PCA, (D) flora composition diagram at the phylum level, (E) heat map of differential bacteria at the family level, (F) differential bacterial abundance map at the genus level, (G) spearman correlation analysis, and correlation analysis between differential bacteria at the genus level and immune characteristics. DETAILED DESCRIPTION

[0036] The present invention is further described in detail below in conjunction with specific embodiments, and the examples provided are only for illustrating the present invention, rather than for limiting the scope of the present invention. The examples provided below can be used as a guide for further improvements by those of ordinary skill in the art, and do not constitute a limitation of the present invention in any way.

[0037] The experimental methods in the following examples, unless otherwise specified, are all conventional methods, and are performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials, reagents, etc. used in the following examples, unless otherwise specified, can all be obtained from commercial channels.

[0038] Example 1 Preparation of black fungus degradable polysaccharide

[0039] S1. Preparation of crude black fungus polysaccharide

[0040] (1) Using black fungus from the Qinba Mountain area in Northwest China as raw material, first drying it at 60° C., crushing it and setting it aside;

[0041] (2) Weigh 20 g of black fungus powder, add 1 L of distilled water, and extract at 100°C for 2 h. After the extraction, centrifuge at 4000 r / min for 15 min and collect the supernatant; filter through three layers of gauze and take the supernatant for later use, add 600 mL of distilled water to the remaining precipitate and extract again at 100°C for 2 h, centrifuge at 4000 r / min for 15 min, and collect the supernatant.

[0042] (3) Take 1200 mL of the supernatant from the two times and perform rotary evaporation to concentrate it to 800 mL. Take out the concentrated black fungus sugar solution, add 3.2 L of anhydrous ethanol, seal it with plastic wrap, and precipitate it at room temperature for 12 hours.

[0043] (4) After alcohol precipitation, centrifuge at 4000 r / min for 15 min, and dissolve the precipitate in 720 mL of 60°C distilled water until it is free of lumps.

[0044] (5) Add 180 mL of Svage reagent (n-butanol: 36 mL of dichloromethane: 144 mL), stir rapidly for 20 min, centrifuge at 4000 rpm for 15 min, take the supernatant and repeat (5) 3 times.

[0045] (6) The supernatant was collected, dialyzed with running water for 72 h, and then freeze-dried to obtain the crude black fungus polysaccharide, referred to as AAP.

[0046] S2. Preparation of degraded polysaccharides from black fungus

[0047] The black fungus crude polysaccharide prepared in step S1 is controllably degraded by a hydrogen peroxide-ascorbic acid oxidative degradation method to obtain black fungus degraded polysaccharide ADP. The specific steps are as follows:

[0048] (1) Accurately weigh 120 mg of AAP, dissolve it in 3880 mL of deionized water, and prepare a suspension in a stoppered vial.

[0049] (2) Add hydrogen peroxide H to the AAP suspension 2 O 2and ascorbic acid Vc, maintain H 2 O 2 The concentration of and Vc was 5 mM. After sealing, the tube was reacted in a 60°C water bath for 1 h.

[0050] (3) After the reaction, the solution was concentrated and dialyzed at 4°C for three days using a dialysis bag with a molecular cutoff of 3500 Da (to remove H 2 O 2 and Vc), and the degradation samples were collected by freeze drying.

[0051] Determination of rheological properties of degraded polysaccharides from black fungus:

[0052] The apparent viscosity of AAP and ADP solutions varies with shear rate. Figure 1 As shown in the figure, within the shear rate variation range, the viscosity of the two polysaccharides decreased due to the decrease in shear rate. However, ADP was significantly lower than AAP, which may be because the molecular weight of the degraded polysaccharide was lower after treatment, the number of cross-linked regions between polysaccharide chains decreased, and the intermolecular interaction force was weakened, making its molecular structure looser and more irregular, which in turn shortened the length of the molecular chain, reduced the resistance to fluid flow, and thus reduced the apparent viscosity.

[0053] Determination of molecular weight of degraded polysaccharides from black fungus:

[0054] like Figure 2 As shown, the molecular weight was determined. The black fungus polysaccharide was controlled degraded to obtain the black fungus degraded polysaccharide ADP with a molecular weight of 83.56 kDa.

[0055] Monosaccharide composition of degraded polysaccharides from black fungus:

[0056] like Figure 3 As shown in the figure, the black fungus polysaccharide in the Qinba Mountain area of ​​Northwest China before and after degradation is mainly composed of mannose and glucuronic acid, and the proportions of each component are similar. The ratio of mannose to glucuronic acid in the undegraded polysaccharide is 5.38:1; the ratio of mannose to glucuronic acid in the degraded polysaccharide is 9.46:1. After testing, the protein concentration of the black fungus polysaccharide before degradation is (4.18±0.36)%, and the uronic acid concentration is (12.31±3.26)%; the protein concentration of the black fungus polysaccharide after degradation is (7.51±0.21)%, and the uronic acid concentration is (27.07±5.51)%.

[0057] Example 2 Detection of the immune enhancement and immunomodulatory effects of black fungus degraded polysaccharides

[0058] Mice were fed with the black fungus crude polysaccharide AAP and the black fungus degraded polysaccharide ADP prepared in Example 1, and then the corresponding indicators of the mice were measured to evaluate the effect of ADP on the immune regulation and improvement of intestinal homeostasis in mice. The details are as follows:

[0059] 1. Grouping and treatment of experimental animals

[0060] After one week of adaptive feeding, 54 BALB / C male mice were randomly divided into normal control group (NC), cyclophosphamide-induced immunodeficiency model group (CTX), CTX+AAP (100 mg / kg BW·day), and CTX+ADP (100 mg / kgBW·day). There were 14 mice in each group except the NC group. Among them, NC and CTX were gavaged with 100 mg / kg BW·day of normal saline every day. The other two groups were gavaged with 100 mg / kg BW·day of AAP and ADP respectively. It lasted for 14 days. All experimental mice were kept at a temperature of 20±2℃ and a relative humidity of 60±5%, and were given free food and water.

[0061] 2. Changes in body weight and organ indexes

[0062] like Figure 4 As shown in (B), after 3 days of CTX modeling, the weight of mice in the modeling group decreased significantly (about 1.90±0.03g), the mice were in a depressed mental state, and moved slowly, indicating that the modeling was successful. On the 4th day, with the end of CTX induction and the start of AAP and ADP intervention, the weight of mice dropped to the lowest point. After the 7th day, the weight of mice in each group began to increase, but compared with the CTX group, the weight of mice in the CTX+AAP and CTX+ADP intervention groups recovered faster. On the 17th day, the weight of mice in the CTX group was still significantly lower than that in the NC group, while the weight of mice in the CTX+ADP intervention group was not significantly different from that in the NC group.

[0063] The spleen and thymus are two important immune organs in the human immune system, playing a key role in triggering protective immune responses, promoting the healing process, and removing harmful stimuli. Among them, the thymus provides a place for the differentiation, development, and maturation of T cells, and plays an important role in maintaining the body's immune balance. The spleen contains a large number of lymphocytes and macrophages that can produce immune substances such as immunoglobulins and complement. Figure 4 As shown in (CD), compared with the NC group, the thymus index of mice in the CTX group decreased by 2.38 times, and the spleen index decreased by 1.17 times. The significant decrease in the thymus index and spleen index indicated that CTX caused serious damage to the immune organs. Compared with the CTX group, the thymus index and spleen index of mice in the CTX+AAP and CTX+ADP groups increased significantly. However, the organ index of the CTX+ADP group was closer to the level of the NC group, indicating that CTX+ADP can more effectively promote the enhancement of immune organs, thereby enhancing the host's immune system.

[0064] 3. Changes in spleen oxidative stress response and cytokine content

[0065] CTX-induced hepatotoxicity is a common side effect, mainly due to the formation of cytotoxic metabolites, which leads to increased ROS concentration in the liver. High concentrations of ROS can cause cellular oxidative stress and lead to immunosuppression. Polysaccharides have strong antioxidant activity and can improve CTX-induced immunosuppression by reducing oxidative stress damage. Figure 5 As shown in (AB), compared with the NC group, the CAT and GSH enzyme activities in the spleen of mice in the CTX group decreased by 1.66 and 5.28 times, respectively. The significant decrease in their enzyme activities indicated that CTX had a serious impact on the liver oxidative stress level. Compared with the CTX group, CTX+AAP and CTX+ADP were able to improve the oxidative stress level in the spleen of mice. In particular, in the CTX+ADP group, the GSH and CAT enzyme activities in the liver and spleen of mice increased by 1.65 and 2.96 times, respectively, and all indicators were effectively restored to levels similar to those of the NC group.

[0066] Cytokines are small molecule proteins that stimulate the synthesis of immune cells and certain non-immune cells, and play a role in regulating and transmitting signals in the immune system. Cytokines such as IL-1β and TNF-α bind to specific cell surface receptors, triggering intracellular signal transduction pathways, thereby affecting the activity and function of immune cells. Figure 5 As shown in (CD), compared with the NC group, the levels of IL-1β and TNF-α in the serum of mice in the CTX group decreased significantly, by 1.26 times and 1.25 times, respectively, indicating that CTX had a significant inhibitory effect on the mouse immune system. Compared with the CTX group, CTX+AAP and CTX+ADP increased the levels of IL-1β and TNF-α in the serum. It is gratifying that the IL-1β and TNF-α in the serum of mice in the CTX+ADP group have basically returned to the levels of the NC group, indicating that CTX+ADP has a significant protective effect in regulating the levels of inflammatory factors.

[0067] 4. Histological changes of jejunum

[0068] As a physical barrier, the intestinal epithelium protects the intestine from external damage through various mechanisms and maintains the stability and health of the intestinal internal environment. Goblet cells can participate in mucosal immunity by secreting antimicrobial proteins, chemokines and cytokines, protecting epithelial cells from physical damage and adhesion of harmful bacteria to epithelial cells. Figure 6As shown in the figure, under an optical microscope, the histological morphology of the jejunum of mice in the NC group was normal, with neat and orderly arrangement of villi, clear boundaries between villi, and large number of goblet cells and mucin area. However, the jejunal villi of mice in the CTX group showed serious lesions, which were manifested as incomplete villus structure, villus atrophy, shallow crypts, and a significant decrease in the number of goblet cells and mucin area, indicating that CTX seriously damaged the tissue structure of the mouse jejunum. Compared with the CTX group, CTX+AAP and CTX+ADP were able to improve CTX-induced intestinal mucosal damage to varying degrees, showing longer and thicker intestinal villi, relatively regular arrangement of intestinal villi, complete structure, and increased number of goblet cells and mucin area. This indicates that black fungus polysaccharides may help restore the integrity of the intestinal mucosa by promoting the repair of intestinal epithelial cells and enhancing the function of goblet cells, thereby protecting the intestine from external damage.

[0069] 5. Changes in intestinal flora

[0070] In order to gain a deeper understanding of the effects of different components of black fungus polysaccharides on the intestinal microbiota of normal mice, the present invention first performed 16S rRNA gene sequencing analysis. Figure 7 As shown in (A), the numbers of unique OTU units in the four groups of NC, CTX, CTX+AAP, and CTX+ADP were 17, 3, 3, and 2, respectively. The results of 16S rRNA gene sequencing showed that Figure 7 (B) The numbers of unique OTUs in the NC, CTX, CTX+AAP, and CTX+ADP groups were 412, 212, 331, and 443, respectively. Compared with the NC group, the number of unique OTUs in the CTX group was significantly reduced, while the numbers of unique OTUs in the CTX+AAP and CTX+ADP groups showed different degrees of recovery.

[0071] In addition, PCA results such as Figure 7 As shown in (C), the intestinal microbial communities after CTX+AAP and CTX+ADP intervention were significantly different from those in the NC group, further indicating that the intervention of CTX+AAP and CTX+ADP may have different effects on the intestinal microbial structure of mice.

[0072] Figure 7 (D) It can be intuitively seen that the abundance of the mouse intestinal flora changed significantly after CTX+AAP and CTX+ADP intake. At the phylum level, Bacteroidota and Firmicutes were the dominant groups. After cyclophosphamide treatment, the relative abundance of Bacteroidota and Campilobacterota decreased, while the relative abundance of Firmicutes, Actinobacteriota, and Desulfobacterota increased. Addition of CTX+AAP and CTX+ADP restored the changes in the intestinal microbiota of this phylum.

[0073] like Figure 7 As shown in (E), at the family level, the intervention of CTX+ADP significantly increased the relative abundance of Eggerthellaceae, Coriobacteriales-Incertae-Sedis, and Desulfovibrionaceae compared with the CTX group. With the addition of CTX+ADP, the bacterial communities with reduced abundance in the CTX group have gradually recovered. It can be seen that the addition of CTX+ADP can effectively regulate the intestinal flora disorder caused by cyclophosphamide.

[0074] Then, through the inter-group T-test, we found the species with significant differences at the genus level, such as Figure 7 As shown in (F), the CTX group significantly increased the abundance of UCG-010, Bacillus, and Rikenella, and the addition of ADP significantly reduced them. Among them, UCG-010 was negatively correlated with the concentration of immune markers; the abundance of Bacillus was positively correlated with the severity of immune diseases; and the abundance of Rikenella was significantly negatively correlated with the level of anti-inflammatory cytokines (IL-10).

[0075] like Figure 7 As shown in (G), Spearman correlation analysis was used to further study the relationship between bacterial species with significant differences at the genus level and immune response parameters after ADP intervention. The relative abundance of these differential bacteria was significantly different from the immune organ index.

[0076] The embodiments described above are only descriptions of the preferred modes of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary technicians in this field should all fall within the protection scope determined by the claims of the present invention.

Claims

1. A black fungus degradable polysaccharide, characterized in that: The monosaccharide composition of the black fungus degraded polysaccharide includes mannose and glucuronic acid, and the molar ratio of mannose to glucuronic acid is 9.46:

1.

2. The black fungus degradable polysaccharide according to claim 1, characterized in that: The molecular weight of the black fungus degraded polysaccharide is 83.56 kDa.

3. The black fungus degradable polysaccharide according to claim 1, characterized in that: The protein concentration of the black fungus degraded polysaccharide is 7.51±0.21%, and the uronic acid concentration is 27.07±5.51%.

4. A method for preparing black fungus degraded polysaccharide, characterized in that: The preparation method comprises the following steps: S1, drying and crushing black fungus, extracting with hot water to obtain a polysaccharide extract, concentrating the polysaccharide extract, and sequentially subjecting the extract to alcohol precipitation, deproteinization, dialysis and drying to obtain black fungus crude polysaccharide; S2. Dissolving the black fungus crude polysaccharide in water to form a black fungus crude polysaccharide suspension, adding hydrogen peroxide and ascorbic acid to the black fungus crude polysaccharide suspension to form a reaction system, reacting the reaction system at 40° C. to 60° C. for 1 h to 3 h, and then concentrating, dialyzing and drying in sequence to obtain the black fungus degraded polysaccharide.

5. The preparation method according to claim 4, characterized in that: In step S1, the extraction temperature of hot water extraction is 95°C to 100°C; and / or, In step S1, the hot water extraction time is 1 h to 2.5 h; and / or, In step S1, the deproteinization uses a sevage reagent, wherein the sevage reagent is a mixed solution of n-butanol and chloroform in a volume ratio of 1:4; and / or, In step S1, the dialysis time is 72±2h.

6. The preparation method according to claim 4, characterized in that: In step S2, the solid-liquid ratio of the black fungus crude polysaccharide suspension is 1:(30-35) mg / mL; and / or, In step S2, the concentration of hydrogen peroxide is 3-100 mM, and the concentration of ascorbic acid is 3-100 mM; and / or, In step S2, the molar concentration ratio of the hydrogen peroxide to the ascorbic acid is 1:1; and / or, In step S2, the dialysis uses a 3500Da dialysis bag and the dialysis time is 72±2h.

7. Use of the degraded black fungus polysaccharide according to any one of claims 1 to 3 or the degraded black fungus polysaccharide prepared by the preparation method according to any one of claims 4 to 6 in preparing products for enhancing immune function and / or immunomodulation.

8. The use according to claim 7, characterized in that: The immunity enhancement or immune regulation includes: restoring body weight, increasing thymus index, increasing spleen index, alleviating oxidative stress damage, increasing cytokine content, reducing intestinal tissue damage, and regulating intestinal flora.

9. The use according to claim 8, characterized in that: The cytokines include: IL-6, IL-10, IL-17, IL-1β, TNF-α; and / or, The regulating of the intestinal flora includes: restoring the number of OTU units and regulating the abundance of the intestinal flora.

10. The use according to claim 9, characterized in that: The regulation of the abundance of intestinal flora includes: inhibited the decrease in the relative abundance of Bacteroidota and Campilobacterota caused by cyclophosphamide; inhibited the increase in the relative abundance of Firmicutes, Actinobacteriota, and Desulfobacterota caused by cyclophosphamide; increased relative abundance of Eggerthellaceae, Coriobacteriales-Incertae-Sedis, and Desulfovibrionaceae; Inhibits the increase in the relative abundance of UCG-010, Bacillus, and Rikenella caused by cyclophosphamide.

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