Application of a polysaccharide produced by akkermansia muciniphila in guangxi in preparation of anti-aging products

Abstract

This invention provides the application of *Ackermania guangxiensis* extracellular polysaccharide in the preparation of anti-aging products, belonging to the technical field of microbial polysaccharide pharmaceutical applications. This invention extracts extracellular polysaccharides from *Ackermania guangxiensis* using the TCA precipitation method. Animal experiments show that *Ackermania guangxiensis* extracellular polysaccharide can effectively reduce MDA content in aging model mice while increasing SOD activity; it improves the thinning of the dermal layer, reduction and loosening of collagen fibers, and other signs of aging in model mice, and increases the hydroxyproline content in the skin; furthermore, it can activate the expression of BDNF and TrKB genes in the mouse hippocampus and alleviate problems such as slow movement, cognitive impairment, and memory decline in model mice. Therefore, the *Ackermania guangxiensis* extracellular polysaccharide provided by this invention has good anti-aging and antioxidant activity and can be used to prepare various anti-aging products.

Description

Technical Field

[0001] This invention belongs to the field of medical applications of microbial polysaccharides, specifically relating to the application of an extracellular polysaccharide from *Ackermania guangxiensis* in the preparation of anti-aging products. Background Technology

[0002] Aging is a complex process that occurs naturally in organisms over time, involving the gradual decline in the function of cells, tissues, and organs. Oxidative stress is one of the important factors contributing to aging. As the body ages, the capacity of the intracellular antioxidant system weakens, leading to the accumulation of free radicals such as reactive oxygen species (ROS), which in turn trigger lipid peroxidation, protein oxidation, and DNA damage, accelerating cellular aging and functional impairment. Therefore, antioxidant strategies have become an important means of delaying aging.

[0003] Extracellular polysaccharides (EPS) are a class of high-molecular-weight compounds secreted by microorganisms during their growth and metabolism, possessing a variety of biological activities. These polysaccharides not only exhibit good water solubility, stability, and biocompatibility, but also demonstrate significant antioxidant, anti-inflammatory, immunomodulatory, and anti-tumor functions. In probiotics, EPS, as important secondary metabolites, not only facilitate the colonization and competition of probiotics in the gut, but also promote overall health by regulating the host's immune response and improving the balance of the gut microbiota.

[0004] Akkermania ( Akkermansia muciniphila Akkermansia (abbreviated as Akkermansia) is a probiotic that inhabits the mucus layer of the mammalian gut. Its unique ability to degrade mucin makes it one of the key microorganisms for maintaining gut health. However, there are currently no reports on the antioxidant and anti-aging properties of extracellular polysaccharides derived from Akkermansia. Summary of the Invention

[0005] In view of this, the purpose of this invention is to provide an application of Guangxi Akermania extracellular polysaccharide (EPS) in the preparation of anti-aging products.

[0006] This invention provides the application of Guangxi Akermania extracellular polysaccharide in the preparation of anti-aging products.

[0007] Preferably, the aging includes at least one of the following: cognitive impairment, decreased learning and memory, slowed movement and reduced exploratory behavior, increased oxidative capacity, and skin aging.

[0008] Preferably, the product includes pharmaceuticals and / or cosmetics.

[0009] Preferably, the effective concentration of Guangxi Akermania extracellular polysaccharide in the product is not less than 10 mg / ml.

[0010] Preferably, the *Ackermania guangxiensis* extracellular polysaccharide in the *Ackermania guangxiensis* includes *Ackermania guangxiensis* (…). Akkermansia guangxiensis sp. strain N21116;

[0011] The preservation number of the Guangxi Akermania strain N21116 is GDMCC No: 62888.

[0012] Preferably, the method for preparing the Guangxi Akermania extracellular polysaccharide involves separating a liquid phase from the culture broth or fermentation broth of Guangxi Akermania, removing proteins from the liquid phase, obtaining a pretreated liquid phase, and then precipitating the polysaccharide with alcohol to obtain Guangxi Akermania extracellular polysaccharide.

[0013] Preferably, the method for removing protein from the liquid phase is to remove protein using trichloroacetic acid;

[0014] The working volume concentration of the trichloroacetic acid is 3.5% to 4.5%.

[0015] Preferably, the alcohol in the alcohol-precipitated polysaccharide comprises an aqueous ethanol solution with a volume concentration of 93% to 97%;

[0016] The volume ratio of the pretreated liquid phase to the alcohol in the alcohol-precipitated polysaccharide is 1:(2~4).

[0017] Preferably, the temperature for alcohol precipitation of polysaccharides is 3~5℃; and the time for alcohol precipitation of polysaccharides is not less than 10h.

[0018] Preferably, after the alcohol precipitation of polysaccharides, the precipitated polysaccharides are further washed 2-3 times with anhydrous ethanol.

[0019] This invention provides the application of *Ackermania guangxiensis* extracellular polysaccharide in the preparation of anti-aging products. This invention utilizes extracellular polysaccharides extracted from *Ackermania guangxiensis* culture broth to investigate anti-aging and antioxidant effects. The *Ackermania guangxiensis* extracellular polysaccharide effectively reduces MDA content in a D-galactose (D-gal)-induced aging mouse model, while increasing SOD activity. It also improves D-galactose-induced thinning of the dermal layer, reduction and loosening of collagen fibers, and other signs of aging in mouse skin, and increases skin hydroxyproline content. Furthermore, it activates the expression of BDNF and TrKB in the mouse hippocampus and alleviates D-galactose-induced bradykinesia, cognitive impairment, and memory decline in mice. Therefore, the *Ackermania guangxiensis* extracellular polysaccharide provided by this invention has good anti-aging activity and can be used to prepare various anti-aging products. Attached Figure Description

[0020] Figure 1 The results of the water maze experiment in D-gal-induced aging mice were obtained by intervention with extracellular polysaccharide (EPS) from *Ackermania* bacteria in Guangxi.

[0021] Figure 2 Results of an EPS intervention experiment in D-galactose-induced aging mice in a mining facility;

[0022] Figure 3 The results of detecting biochemical indicators such as SOD and MDA in the serum of EPS-intervention D-galactose-induced aging mice;

[0023] Figure 4 The results of detecting biochemical indicators such as SOD and MDA in the liver of EPS-intervention D-galactose-induced aging mice;

[0024] Figure 5 The results of Masson staining experiment on skin tissue of D-galactose-induced aging mice treated with EPS;

[0025] Figure 6 The results of the experiment on the detection of hydroxyproline content in the skin tissue of D-galactose-induced aging mice after EPS intervention;

[0026] Figure 7 The results show the expression levels of the BDNF signaling pathway in the hippocampus of D-galactose-induced aging mice after EPS intervention. Detailed Implementation

[0027] This invention provides the application of Guangxi Akermania extracellular polysaccharide in the preparation of anti-aging products.

[0028] In this invention, the *Ackermania guangxiensis* extracellular polysaccharide preferably includes *Ackermania guangxiensis* (…). Akkermansia guangxiensis sp. The strain is N21116. The preferred accession number for the *Ackermaniasis* strain N21116 is GDMCC No: 62888. The *Ackermaniasis* strain N21116 is a known strain, deposited and published in patent publication number CN116200312A.

[0029] In this invention, the method for preparing the extracellular polysaccharide of *Ackermania guangxiensis* preferably involves separating the liquid phase from the culture broth or fermentation broth of *Ackermania guangxiensis*, removing proteins from the liquid phase, obtaining a pretreated liquid phase, and then precipitating the polysaccharide with alcohol to obtain the *Ackermania guangxiensis* extracellular polysaccharide. The method for preparing the culture broth or fermentation broth of *Ackermania guangxiensis* involves inoculating activated *Ackermania guangxiensis* into a BHI culture medium containing 0.05% (v / v) L-cysteine ​​hydrochloride for anaerobic culture to obtain a seed culture. The seed culture is then inoculated again into a BHI culture medium containing 0.05% (v / v) L-cysteine ​​hydrochloride for expansion culture to obtain a culture medium. The inoculation amount of the seed culture is preferably 5%–10%, but can be 6%–8%, or 7%. The anaerobic culture or expansion culture is preferably conducted under anaerobic conditions. The temperature of the anaerobic culture or expansion culture is preferably 36–38°C, but can be 37°C. The anaerobic culture time is preferably 20–24 hours, but can be 23 hours. The optimal culture time is 40–50 hours, but can be 45–48 hours.

[0030] In this invention, the method for separating the liquid phase is preferably centrifugation. The centrifugal force is preferably 8000-12000g, and can be 10000g. The centrifugation time is preferably 8-12 min, and can be 10 min. The method for removing protein from the liquid phase is preferably using trichloroacetic acid. The working volume concentration of the trichloroacetic acid is preferably 3.5%-4.5%, and can be 4%. The alcohol in the alcohol-precipitated polysaccharide preferably includes an aqueous ethanol solution with a volume concentration of 93%-97%, and can be a 95% aqueous ethanol solution. The volume ratio of the pretreated liquid phase to the alcohol in the alcohol-precipitated polysaccharide is preferably 1:(2-4), and can be 1:3. The temperature for alcohol precipitation of polysaccharide is preferably 3-5℃, and can be 4℃; the time for alcohol precipitation of polysaccharide is preferably not less than 10 h, and can be 12-20 h, and can also be 15 h-18 h. After alcohol precipitation of polysaccharide, it is preferable to further wash the precipitated polysaccharide with anhydrous ethanol 2-3 times.

[0031] In this invention, the aging preferably includes at least one of the following: cognitive impairment, decreased learning and memory ability, slowed movement and reduced exploratory behavior, increased oxidative capacity, and skin aging. In an embodiment of this invention, a D-galactose-induced mouse aging model was used as the experimental subject to verify the function of the Guangxi Akkermansia bacteria extracellular polysaccharide. The daily dose of the D-galactose was 100 mg / kg, administered continuously for 56 days.

[0032] In one embodiment of the present invention, a D-galactose-induced mouse aging model was used as the subject, and the Morris water maze test was used to detect the learning and memory abilities of the mice. The results showed that the escape latency of the model group was significantly prolonged compared with the normal group, while the escape latency of the EPS-treated group was significantly shortened compared with the model group, indicating that its spatial learning and memory abilities were significantly enhanced. The platform movement exploration experiment showed that the number of times the mice in the extracellular polysaccharide-treated group crossed the third quadrant increased, further proving the improvement of their spatial memory abilities.

[0033] In one embodiment of the present invention, a D-galactose-induced mouse aging model was used as the subject. The mice's movement and exploration abilities were tested through a mining experiment. Mice in the model group, after receiving galactose-induced aging treatment, exhibited aging behaviors such as slowed movement and reduced exploration. Mice in the experimental group, after receiving extracellular polysaccharide intervention concurrently with galactose-induced aging treatment, showed more active movement and exploration behaviors. The experimental group mice's movement distance, movement speed, and time spent in the central area were significantly higher than those in the model group, indicating that EPS has a significant improving effect on the behavior of galactose-induced aging mice and may have potential anti-aging value.

[0034] In one embodiment of the present invention, in order to further explore the molecular mechanism by which EPS affects aging model mice, the expression of BDNF gene and its receptor TrkB gene in the hippocampus of mice was also detected. The results showed that the expression of BDNF gene in mice was significantly increased compared with the model group, indicating that extracellular polysaccharides have the effect of upregulating BDNF expression. At the same time, compared with the normal control group, the expression of TrkB gene in the model group mice was reduced, and the expression of TrkB gene in mice after EPS intervention was increased compared with the model group, further supporting the activation effect of extracellular polysaccharides on the BDNF signaling pathway.

[0035] In another embodiment of the present invention, using a D-galactose-induced mouse aging model, the effect of EPS on the antioxidant capacity of galactose-induced aging mice was also verified. Compared with the normal control group, the activity of SOD, an oxidation marker, was significantly decreased and the content of MDA was significantly increased in the serum and liver of the model mice; while EPS could increase SOD activity in the serum and liver of mice, while decreasing MDA content. This indicates that Akkermansia extracellular polysaccharides have the ability to enhance the antioxidant activity of mouse serum and liver.

[0036] In another embodiment of the present invention, the effect of EPS on improving skin aging induced by galactose in mice was verified. Compared with the normal control group, the collagen fiber content in the skin of galactose-induced aging mice was reduced, and the arrangement became looser. EPS intervention improved the content and arrangement of collagen fibers in the skin of aging mice, showing an increase in collagen fiber content and a more compact arrangement. This indicates the protective effect of extracellular polysaccharides on the skin of D-galactose-induced aging mice. Simultaneously, the EPS can also increase the content of hydroxyproline in skin tissue. Hydroxyproline is an imino acid and one of the main components of collagen tissue; therefore, the content of collagen in the skin is often assessed by detecting the content of hydroxyproline in the skin, thereby characterizing the degree of skin aging.

[0037] In this invention, the product preferably includes pharmaceuticals and / or cosmetics. The effective concentration of *Ackermania guangxiensis* extracellular polysaccharide in the product is preferably not less than 10 mg / ml, and can be 10-15 mg / ml, or even 112-13 mg / ml. This invention does not impose any particular limitation on the preparation method of the product; any product preparation method well known in the art can be used.

[0038] The following detailed description, in conjunction with embodiments, illustrates the application of an extracellular polysaccharide from *Akermansia spp.* provided by this invention in the preparation of anti-aging products. However, these descriptions should not be construed as limiting the scope of protection of this invention.

[0039] Example 1

[0040] A method for extracting extracellular polysaccharides (EPS) from *Ackermania guangxiensis*.

[0041] Activated *Akermansia guangxiensis* N21116 was inoculated into BHI medium containing L-cysteine ​​hydrochloride (0.05% v / v) and cultured anaerobically to obtain a seed culture. This seed culture was then inoculated at a rate of 10% (v / v) into BHI medium containing L-cysteine ​​hydrochloride (0.05% v / v) and cultured anaerobically at 37°C for 48 h to obtain the fermentation broth. The fermentation broth was centrifuged at 10000×g for 10 min, and the supernatant was collected for the extraction of extracellular polysaccharides.

[0042] The fermentation supernatant was mixed with 4% trichloroacetic acid (TCA) and allowed to precipitate proteins overnight. After centrifugation at 10000×g for 10 min, the supernatant was collected. Three volumes of 95% ethanol solution were added to the supernatant while stirring, and the mixture was incubated at 4°C overnight. The precipitate was then collected after centrifugation at 10000×g for 10 min, reconstituted with water, and dialyzed using an 8–14 kDa dialysis bag. The resulting product was then freeze-dried under vacuum to obtain *Ackermania guangxiensis* extracellular polysaccharide (EPS).

[0043] The crude polysaccharide prepared above was determined by the sulfuric acid-phenol method, and the polysaccharide yield of Guangxi Akermania extracellular polysaccharide (EPS) was 80%–85%.

[0044] Example 2

[0045] Efficacy verification of *Ackermania* extracellular polysaccharide (EPS) in Guangxi

[0046] 1. Construction of a D-galactose (D-gal)-induced aging mouse model

[0047] Thirty 6-8 week old C57BL / 6J mice were randomly divided into a control group, a model group, and an experimental group, with 10 mice in each group. After one week of acclimatization, the control group mice were subcutaneously injected with 200 μl of physiological saline in their backs daily, while the model group and experimental group mice were subcutaneously injected with the same volume of D-galactose solution (100 mg / kg) as the control group mice in their backs daily. At the same time, the experimental group mice were administered 200 μl of EPS (150 mg / kg) by gavage daily for 56 days.

[0048] 2. Water Maze Experiment

[0049] One week before the end of the experiment, the learning and memory abilities of mice were assessed using the Morris water maze test. The test pool had a diameter of 1.2m and a height of 40-50cm, and water was poured into the pool to a depth of 30cm. The pool was divided into four quadrants, with a circular platform of 9cm in diameter and 28cm in height fixed in the third quadrant. Food-grade white pigment was added to the water to conceal the platform. A camera was positioned vertically above the center of the water maze pool, ensuring the camera could cover the entire test area. The water temperature was maintained at 22±1℃. The experimental site was kept well-lit and free of significant noise during the test. Day 1: Adaptation training. Mice were placed in the water from four directions (east, south, west, and north) facing the pool wall to familiarize them with the environment. Days 2-5: Formal testing. The time it took for the mouse to find the underwater platform was recorded (i.e., the escape latency). If the mouse did not find the platform within 60 seconds, it was recorded as 60 seconds. Day 6: Conduct the exploration experiment. Remove the platform and place the mice into the water from the first quadrant facing the pool wall. Record the number of times the mice cross the third quadrant.

[0050] See results Figure 1 Compared to the aging model group, mice in the D-galactose + EPS group showed a significantly shorter escape latency over several days of testing, indicating a marked improvement in their spatial learning and memory abilities. Figure 1 (A). Exploration experiment results showed that, in the exploration experiment after platform removal, mice in the extracellular polysaccharide-treated group traversed the third quadrant more times, further demonstrating their improved spatial memory ability. Figure 1 (B)

[0051] 3. Mine Field Experiment

[0052] One week before the end of the experiment, the mice's mobility and exploration abilities were assessed using a mine-based experiment. The experiment was conducted in a quiet environment to minimize external interference with the mice's behavior. The test chamber was 50cm long, 50cm wide, and 40cm high, with a white interior to ensure clear imaging and prevent the mice's behavior from being affected by color.

[0053] Position the camera equipment directly above and center of the test chamber, adjusting the shooting angle and focus to ensure the field of view covers the entire experimental area. Allow the mice to adapt to the test chamber environment and reduce stress. After the adaptation test, place the mice into the test chamber from a designated corner. Record the mice's movement trajectory for 10 minutes after entering the chamber using the camera equipment, including movement distance, speed, and exploration area. Clean the test chamber before each mouse's test to avoid interference from the previous mouse's excrement and odor. Analyze the recorded mouse movement trajectories to compare the behavioral differences between the experimental and control groups in the mine experiment.

[0054] See results Figure 2 Mice in the model group, after receiving D-galactose-induced aging treatment, exhibited signs of aging such as slowed movement and reduced exploratory behavior. Mice in the experimental group, after receiving D-galactose-induced aging treatment concurrently with extracellular polysaccharide intervention, showed more active movement and exploratory behavior. The distance traveled by the experimental group mice (…) Figure 2 (A) Movement speed ( Figure 2 (B) and the time spent in the central area ( Figure 2 The C value in the middle group was significantly higher than that in the model group, indicating that extracellular polysaccharides have a significant effect on improving the behavior of D-galactose-induced aging mice and may have potential anti-aging value.

[0055] 4. Effects of EPS on antioxidant capacity in D-gal-induced aging mice

[0056] ① Detection of oxidation indicators in serum: At the end of the experiment, serum samples were collected by drawing blood from the hearts of mice. The activity of SOD and the content of MDA in the serum were detected using the corresponding kits.

[0057] See results Figure 3 Compared with the normal control group, the serum SOD activity and MDA content of mice in the aging model group were significantly decreased, while those in the extracellular polysaccharide intervention group were significantly increased compared with those in the aging model group. Figure 3 In the middle A group, the MDA content was significantly lower than that in the aging model group. Figure 3 (B) indicates that Akkermania extracellular polysaccharide has the ability to enhance the antioxidant activity of mice.

[0058] ② Detection of oxidation indicators in the liver: After slaughtering mice, the abdominal cavity was quickly opened and the liver was removed. The liver surface was washed with physiological saline to remove blood and impurities, and then the moisture was blotted with filter paper. An appropriate amount of tissue was taken for subsequent detection of antioxidant indicators. An appropriate amount of liver tissue was taken, homogenized, centrifuged, and the supernatant was collected according to the SOD / MDA kit instructions. Then, the reaction solution was added to carry out the colorimetric reaction, the absorbance was measured, and the SOD activity and MDA content were calculated.

[0059] See results Figure 4 Compared with the normal control group, the SOD activity and MDA content in the liver of mice in the aging model group were significantly decreased, indicating that D-gal-induced aging leads to a decline in the liver's antioxidant capacity. The SOD activity in the liver of mice in the extracellular polysaccharide intervention group was significantly higher than that in the aging model group. Figure 4 In the middle A group, the MDA content was significantly lower than that in the aging model group. Figure 4 (B) indicates that extracellular polysaccharides can enhance the liver's antioxidant capacity.

[0060] 5. EPS improves skin aging induced by D-gal in mice.

[0061] ① Masson staining experiment on skin tissue: After slaughtering mice, the hair on the back of the mice was removed and the skin was harvested. The skin tissue was fixed, dehydrated, impregnated with paraffin, and embedded to prepare paraffin sections. The skin was stained using a Masson staining kit, and finally the sections were mounted with neutral resin. The sections were observed under a microscope, and images were acquired and analyzed.

[0062] See results Figure 5 Collagen fibers were stained blue (counterstained with aniline blue), while muscle fibers and cellulose were stained red. The skin tissue structure was clear, with collagen fibers and muscle fibers clearly distinguishable. Compared to the control group ( Figure 5 In D-gal-induced aging mice, the content of collagen fibers in the skin decreased and their arrangement became looser. Figure 5 (B) After extracellular polysaccharide intervention, the content and arrangement of collagen fibers in the skin of aging mice were improved, manifested as increased collagen fiber content and more compact arrangement. Figure 5 (C) This indicates the protective effect of extracellular polysaccharides on the skin of D-gal-induced aging mice. Akkermania extracellular polysaccharides have significant anti-aging effects.

[0063] ② Hydroxyproline content determination experiment in skin tissue: After slaughtering mice, the hair on the back of the mice was removed and the skin tissue from the back was collected as a sample. The sample freshness was ensured, and it was immediately frozen to prevent degradation of hydroxyproline. Hydroxyproline was extracted from the skin tissue and its content was determined according to the instructions of the hydroxyproline content determination kit.

[0064] See results Figure 6 Compared with the control group, the hydroxyproline content in the skin of D-gal-induced aging mice was significantly reduced. After intervention with extracellular polysaccharides, the hydroxyproline content in the skin of aging mice increased. This indicates that extracellular polysaccharides have a significant anti-aging effect and can improve the hydroxyproline content in the skin.

[0065] 6. EPS activates the BDNF signaling pathway in the hippocampus.

[0066] Hippocampal tissue was collected from mice after slaughter, immediately flash-frozen in liquid nitrogen, and then stored at -80℃. 0.02g of tissue was weighed, and total RNA was extracted from the brain and hippocampus using the Trizol method according to the instructions of the R001 BetterZol Reagent reagent (Beijing Jinbaite Biotechnology Co., Ltd.). RNA was converted to cDNA using a reverse transcription kit (Tiangen Biotech Co., Ltd.) as a template for qPCR. Specific primers were designed and synthesized based on the sequence information of BDNF pathway-related genes (BDNF, receptor TrkB) (Table 1). qPCR was performed on a QuantStudio™ System with the following quantitative fluorescence program: amplification curve: 94℃ 30s, 94℃ 15s, 58℃ 30s, 40 cycles; melting curve: 95℃ 15s, 60℃ 1min, 95℃ 15s. The housekeeping gene GAPDH was used as an internal control. ﹣△△Ct The method allows for quantitative analysis of the target gene.

[0067] Table 1 Primers for Real-Time PCR

[0068]

[0069] See results Figure 7 Compared with the normal control group, the expression of the BDNF gene in the hippocampus of mice in the D-galactose-induced aging model group was significantly reduced. The expression of the BDNF gene in mice in the EPS-treated group was significantly increased compared with the model group. Figure 7 (A) indicates that EPS upregulates BDNF expression. TrkB is the main receptor for BDNF, and changes in its expression level reflect the activity of the BDNF signaling pathway. Compared with the normal control group, TrkB gene expression was decreased in the model group mice, while TrkB gene expression was increased in the extracellular polysaccharide treatment group mice compared with the model group (A). Figure 7 (B) This further supports the activation effect of EPS on the BDNF signaling pathway.

[0070] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. The application of an extracellular polysaccharide from *Akermansia amurensis* in the preparation of an anti-aging product, wherein the aging includes at least one of the following: cognitive impairment, decreased learning and memory ability, slowed movement and reduced exploratory behavior, increased oxidative capacity, and skin aging; wherein the product is a pharmaceutical or cosmetic product; The Guangxi Akermania is *Ackermania guangxiensis* (…). Akkermansia guangxiensis sp. The strain N21116; the preservation number of the Guangxi Akermania strain N21116 is GDMCC No: 62888; The method for preparing Guangxi Akermania extracellular polysaccharide involves separating a liquid phase from the culture broth or fermentation broth of Guangxi Akermania, removing proteins from the liquid phase, obtaining a pretreated liquid phase, and then precipitating the polysaccharide with alcohol to obtain Guangxi Akermania extracellular polysaccharide.

2. The application according to claim 1, characterized in that, The effective concentration of Guangxi Akermania extracellular polysaccharide in the product is not less than 10 mg / ml.

3. The application according to claim 1, characterized in that, The method for removing protein from the liquid phase is to remove protein using trichloroacetic acid; The working volume concentration of the trichloroacetic acid is 3.5% to 4.5%.

4. The application according to claim 1, characterized in that, The alcohol in the precipitated polysaccharide includes an aqueous solution of ethanol with a volume concentration of 93% to 97%. The volume ratio of the pretreated liquid phase to the alcohol in the alcohol-precipitated polysaccharide is 1:(2-4).

5. The application according to claim 1, characterized in that, The temperature for alcohol precipitation of polysaccharides is 3-5°C; the time for alcohol precipitation of polysaccharides is not less than 10 hours.

6. The application according to claim 1, characterized in that, The process of precipitating polysaccharides with alcohol also includes washing the precipitated polysaccharides 2 to 3 times with anhydrous ethanol.

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