Radix fici simplicissimae polysaccharide extract as well as preparation method and application thereof

Abstract

The invention discloses a radix fici simplicissimae polysaccharide extract and a preparation method and application thereof.The radix fici simplicissimae polysaccharide extract is radix fici simplicissimae crude polysaccharide or purified polysaccharide, and the radix fici simplicissimae crude polysaccharide is obtained by conducting primary alcohol precipitation, Sevage method protein removal and secondary alcohol precipitation on a radix fici simplicissimae water extract and then conducting drying; the purified polysaccharide is obtained by performing DEAE-52 fiber column chromatography on RFH, eluting by taking a sodium chloride solution as an eluent, respectively collecting elution parts of 0.3 mol / L sodium chloride solution, 0.4 mol / L sodium chloride solution and 0.5 mol / L sodium chloride solution, and respectively performing dialysis and drying. The radix fici simplicissimae polysaccharide extract prepared through a limited method can effectively activate macrophages to generate immunocompetence and is expected to be used for preparing immunomodulators or related drugs or food for enhancing the immunologic function.

Description

Technical Field

[0001] This invention relates to active ingredients extracted and isolated from plants, specifically to the polysaccharide extract of *Ficus hirta*, its preparation method, and its application. Background Technology

[0002] Polysaccharides are the most abundant biopolymers in nature, present in almost all living organisms, and possess a wide range of biological functions, including energy storage, structural support, defense, and antigenic determinism. Polysaccharides have become a research hotspot due to their potential beneficial effects on human health; for example, polysaccharides from plants and fungi exhibit pharmacological activities such as immunomodulation and antitumor activity. Whether a polysaccharide possesses immunomodulatory activity is usually related to its structure, such as monosaccharide composition, glycosidic bond composition, conformation, molecular weight, functional groups, and branching characteristics. For example, patent application CN117050208A discloses a highly active polysaccharide extract from young kiwifruit. This extract utilizes an optimized microwave-assisted eutectic solvent extraction method, resulting in polysaccharides with lower molecular weights and higher uronic acid content, antioxidant activity, and immunomodulatory activity. Another example is patent application CN102166294A, which discloses an extract of Anemarrhena asphodeloides polysaccharides, its preparation method, and its pharmaceutical uses. In preparation, the raw material was defatted and then extracted with water and precipitated with alcohol to obtain crude polysaccharide. The crude polysaccharide was then subjected to dialysis and ion exchange chromatography to remove proteins, pigments, and small molecule chemical components, yielding a high-purity Anemarrhena asphodeloides polysaccharide extract. The content of Anemarrhena asphodeloides polysaccharide in the obtained extract, calculated as glucose, was over 60 wt%. Experiments showed that the obtained Anemarrhena asphodeloides polysaccharide extract possesses immunosuppressive activity.

[0003] Five-finger fig (Ficus simplicissima Lour.) is the root of a plant belonging to the Ficus genus (Moraceae). It is mainly produced in Guangdong, Guangxi, Yunnan, and Guizhou provinces of my country and is a commonly used traditional Chinese medicine in the Lingnan region. According to ancient texts such as *Sheng Cao Yao Xing Bei Yao*, five-finger fig has the effects of strengthening the spleen and lungs, promoting qi circulation and removing dampness, and relaxing muscles and tendons. In Guangdong, its root is often used in soups and is commonly referred to as "Guangdong ginseng."

[0004] Five-finger peach is rich in various chemical components, including organic acids, flavonoids, sugars, terpenes, alkaloids, and coumarins. Current research on five-finger peach still has certain limitations, especially in the exploration of polysaccharides; to date, there are no reports of five-finger peach polysaccharides possessing immunomodulatory activity. Summary of the Invention

[0005] The technical problem to be solved by the present invention is to provide a polysaccharide extract of *Ficus hirta* that can enhance immune function, its preparation method and application.

[0006] To solve the above-mentioned technical problems, the present invention adopts the following technical solution:

[0007] Five-finger peach polysaccharide extract, which is crude or purified polysaccharide from five-finger peach, including...

[0008] The crude polysaccharide of *Ficus hirta* is obtained by first alcohol precipitation, removal of protein by Sevage method and second alcohol precipitation of water extract of *Ficus hirta*, followed by drying. It is denoted as RFH and has a total sugar content of ≥39.0 wt%.

[0009] The purified polysaccharides were obtained by DEAE-52 fiber column chromatography on RFH, eluting with sodium chloride solution. The eluted fractions of 0.3 mol / L, 0.4 mol / L, and 0.5 mol / L sodium chloride solutions were collected, and then dialyzed and dried respectively. The purified polysaccharides were designated as RFH-D1, RFH-D2, and RFH-D3, respectively, with total sugar contents of 90.0–95.0 wt%, 70.0–79.0 wt%, and 80.0–87.0 wt%, respectively.

[0010] In the above technical solution, the total sugar content in the crude polysaccharide of Prunus cerasifera is preferably 39.5-49.0 wt%, and the total sugar content in the purified polysaccharides RFH-D1, RFH-D2 and RFH-D3 is preferably 91.2-93.8 wt%, 72.3-77.9 wt%, and 81.4-86.3 wt%, respectively.

[0011] In the above technical solution, the alcohol used for alcohol precipitation is 95-100 v / v% ethanol; the molecular weight cutoff for dialysis is greater than or equal to 3500 Da, preferably 3500 Da.

[0012] The preparation method of the polysaccharide extract of *Ficus hirta* according to the present invention includes a method for preparing crude polysaccharide from *Ficus hirta* and a method for preparing purified polysaccharide, wherein...

[0013] The preparation method of crude polysaccharide of Prunus pubescens includes: subjecting the water extract of Prunus pubescens to one alcohol precipitation, Sevage method to remove protein and a second alcohol precipitation, and then drying it to obtain the product, denoted as RFH, wherein the total sugar content is ≥39.0wt%.

[0014] The method for preparing purified polysaccharides includes: chromatography on a DEAE-52 fiber column on an RFH column, eluting with sodium chloride solution as the eluent, collecting the eluted fractions from 0.3 mol / L, 0.4 mol / L, and 0.5 mol / L sodium chloride solutions, respectively, and then dialysis and drying them to obtain the purified polysaccharides, which are respectively designated as RFH-D1, RFH-D2, and RFH-D3, with total sugar contents of 90.0–95.0 wt%, 70.0–79.0 wt%, and 80.0–87.0 wt%, respectively.

[0015] The applicant discovered in its experiments that the polysaccharide extract obtained by removing proteins using the Sevage method exhibited superior immunomodulatory activity compared to polysaccharide extracts obtained by other methods (such as the trichloroacetic acid method (TCA method)) with lower polysaccharide content. Furthermore, the applicant's experiments revealed that when the polysaccharides in the obtained polysaccharide extract are basic polysaccharides without acid bonds (such as polysaccharide extracts obtained using an alkaline reagent (1 mol / L NaOH / 0.05% NaBH4)), the resulting extract lacks immunomodulatory activity.

[0016] In the above-mentioned method for preparing crude polysaccharide from *Ficus hirta*, the aqueous extract of *Ficus hirta* is obtained by extracting *Ficus hirta* with water, filtering, collecting the filtrate, and concentrating it. The extraction employs conventional extraction methods, such as reflux extraction or ultrasonic extraction, with reflux extraction being preferred. During extraction, the number of extractions, the material-to-liquid ratio for each extraction, and the extraction temperature are the same as in existing technologies.

[0017] In the above method for preparing crude polysaccharide from *Ficus hirta*, the alcohol used for alcohol precipitation is 95-100 v / v ethanol, preferably 100 v / v ethanol. The amount of alcohol added is preferably such that the concentration of ethanol in the system is greater than or equal to 80 v / v%.

[0018] In the above-mentioned method for preparing purified polysaccharides, the molecular weight cutoff for dialysis is greater than or equal to 3500 Da, preferably 3500 Da.

[0019] The application process has shown that the polysaccharide extract of *Ficus hirta* described in this invention has the effect of activating macrophages, promoting the secretion of cytokines NO, IL-6 and TNF-α, effectively activating macrophages to produce immune activity, and enhancing the body's specific and non-specific immune functions. Therefore, this invention also includes the application of the above-mentioned polysaccharide extract of *Ficus hirta* in the preparation of immune-regulating foods or medicines.

[0020] Furthermore, the present invention also includes a food or pharmaceutical composition containing the above-mentioned Prunus pedunculata polysaccharide extract and a pharmaceutically acceptable carrier.

[0021] Compared with existing technologies, the present invention prepares polysaccharide extract of Prunus pubescens using a defined method. Experiments have shown that the polysaccharide extract obtained by this method has a superior macrophage activation effect compared with extracts obtained by other protein removal methods. It can promote the secretion of cytokines NO, IL-6 and TNF-α, effectively activate macrophages to produce immune activity, and is expected to be used to prepare immunomodulators or related drugs or foods that enhance immune function. Attached Figure Description

[0022] Figure 1This is a gradient elution curve of polysaccharide purified from Prunus pubescens in Example 2 of the present invention. Detailed Implementation

[0023] To better explain the technical solution of the present invention, the present invention will be further described in detail below with reference to the embodiments, but the implementation of the present invention is not limited thereto.

[0024] Example 1: Preparation of crude polysaccharide (RFH) from *Ficus hirta* using the method described in this invention.

[0025] 800g of *Ficus hirta* was crushed and extracted three times by reflux at 100℃ for 1 hour each time, using a material-to-liquid ratio of 1g:8mL. The filtrates were combined and concentrated to obtain the aqueous extract of *Ficus hirta*. 100v / v% ethanol was added to the aqueous extract for one-time alcohol precipitation (the volume of 100v / v% ethanol was 4 times the volume of the aqueous extract). The mixture was placed at 4℃ for 12 hours, the supernatant was decanted, and the ethanol was removed from the precipitate until no alcohol odor remained, yielding residue A. Proteins in residue A were removed using the Sevage method. Sevage reagent (n-butanol:chloroform = 1:4, volume ratio) was added to residue A, with a volume ratio of Sevage reagent to residue A of 1:4. The mixture was shaken for 5 minutes to mix, centrifuged at 3500 rpm for 10 minutes, and the supernatant was decanted. This process was repeated twice. Finally, the Sevage reagent was removed by rotation to obtain residue B. Add 100 v / v% ethanol to residue B for secondary alcohol precipitation (the amount of 100 v / v% ethanol is 4 times the volume of residue B), place at 4°C for 12 h, decant the supernatant, remove ethanol from the precipitate until no alcohol odor remains, freeze dry to obtain RFH.

[0026] The total sugar content in the RFH obtained in this example was determined using the phenol-sulfuric acid method, with glucose as the calibrator for the total sugar content.

[0027] ① Preparation of standard solutions and construction of standard curves: Accurately weigh 5 mg of glucose standard, dissolve it fully in pure water to prepare a 1 mg / mL stock solution, and gradually dilute the series of standard solutions using the glucose stock solution. Pipette 200 μL of each standard solution of different concentrations, add 0.2 mL of 5% phenol solution, then quickly add 1 mL of concentrated sulfuric acid. Boil in water for 15–20 min, allow to cool, and then measure the OD value at 490 nm. Plot a linear regression equation with glucose concentration on the x-axis and absorbance (OD value) on the y-axis. The linear regression equation is: y =

[0028] 0.4542x+0.0287 (R2=0.9916).

[0029] ② Preparation and determination of sample solutions: Accurately weigh 1 mg of crude polysaccharide RFH from *Ficus hirta*, dissolve it thoroughly in pure water to prepare a 1 mg / mL sample stock solution. Pipette 50 μL of each sample stock solution and dilute with 950 μL of pure water to a 0.05 mg / mL sample solution. Pipette 200 μL of each sample solution, 200 μL of 5% phenol solution, and 1 mL of concentrated sulfuric acid solution, respectively, and boil in boiling water for 15–20 min. After cooling, measure the OD value at 490 nm. Substitute these values ​​into the standard curve to calculate the total sugar content of crude polysaccharide RFH from *Ficus hirta*.

[0030] The total sugar content of the RFH obtained in this example was calculated to be 48.4 wt%.

[0031] Comparative Example 1-1

[0032] Example 1 was repeated, except that protein was removed from residue A using the TCA method. 10% trichloroacetic acid was added to residue A, with a volume ratio of 1:5 between 10% trichloroacetic acid and residue A. The mixture was shaken for 5 minutes to mix, centrifuged at 3500 rpm for 10 minutes, and the supernatant was discarded. This process was repeated twice. Finally, trichloroacetic acid was removed by rotation to obtain residue B.

[0033] Finally, RFH1-1 was obtained.

[0034] The total sugar content in RFH1-1 obtained in this example was determined using the same method as in Example 1. The calculated total sugar content in RFH1-1 obtained in this example was 51.3 wt%.

[0035] Comparative Examples 1-2

[0036] 800g of *Ficus hirta* was crushed and reacted with an alkaline reagent (1 mol / L NaOH / 0.05% NaBH4) at a ratio of 1g:10mL at 25℃ for 16 hours. The mixture was filtered, and the filtrate was concentrated and neutralized to neutral with 50 v / v% acetic acid to obtain material A. 100 v / v% ethanol was added to material A for a first alcohol precipitation (the amount of 100 v / v% ethanol was 4 times the volume of the material). The mixture was placed at 4℃ for 12 hours, the supernatant was decanted, and the precipitate was subjected to rotary evaporation to remove the ethanol until no alcohol odor was detected, yielding material B. 100 v / v% ethanol was added to material B for a second alcohol precipitation (the amount of 100 v / v% ethanol was 4 times the volume of the material). The mixture was placed at 4℃ for 12 hours, the supernatant was decanted, and the precipitate was subjected to rotary evaporation to remove the ethanol until no alcohol odor was detected. The precipitate was freeze-dried to obtain the alkaline polysaccharide of *Ficus hirta*, designated RFH1-2.

[0037] The total sugar content in RFH1-2 obtained in this example was determined using the same method as in Example 1. The calculated total sugar content in RFH1-2 obtained in this example was 29.5 wt%.

[0038] Example 2: Preparation of crude polysaccharide (RFH) from *Ficus hirta* using the method described in this invention.

[0039] Repeat Example 1, except that:

[0040] When extracting five-finger peach with water, the extraction temperature was changed to 90℃;

[0041] In the first and second alcohol precipitation processes, the ethanol used was changed to 95 v / v ethanol.

[0042] The total sugar content in the RFH obtained in this example was determined using the same method as in Example 1. The calculated total sugar content in the RFH obtained in this example was 40.3 wt%.

[0043] Example 3: Preparation of crude polysaccharide (RFH) from *Ficus hirta* using the method described in this invention.

[0044] Repeat Example 1, except that when extracting five-finger peach with water, the ratio of five-finger peach residue to deionized water is 1g:5mL.

[0045] The total sugar content in the RFH obtained in this example was determined using the same method as in Example 1. The calculated total sugar content in the RFH obtained in this example was 39.9 wt%.

[0046] Example 4: Preparation of purified polysaccharides (RFH-D1, RFH-D2, and RFH-D3) from *Ficus hirta* using the method described in this invention.

[0047] The RFH prepared according to the method described in Example 1 was dissolved in water and then subjected to chromatography on a DEAE-52 fiber column (40 mm × 500 mm). After washing with water to remove impurities, elution was performed sequentially using 0.1 mol / L, 0.2 mol / L, 0.3 mol / L, 0.4 mol / L, and 0.5 mol / L sodium chloride solutions as eluents at a flow rate of 3.75 mL / min. The eluent fractions containing polysaccharides were collected from the 0.3 mol / L, 0.4 mol / L, and 0.5 mol / L sodium chloride solutions, respectively (the OD value of the eluent at 490 nm was detected by the phenol-sulfuric acid method). The eluent containing polysaccharides was collected in 15 mL fractions per tube until no polysaccharides were detected. The gradient elution curves are shown in the figure. Figure 1 As shown in the figure, each eluted fraction was dialyzed (with a molecular weight cutoff of 3500 Da) and freeze-dried to obtain 0.3 M fraction purified polysaccharide (denoted as RFH-D1), 0.4 M fraction purified polysaccharide (denoted as RFH-D2), and 0.5 M fraction purified polysaccharide (denoted as RFH-D3), respectively.

[0048] The total sugar content in RFH-D1, RFH-D2, and RFH-D3 obtained in this example was determined using the same method as in Example 1. The calculated total sugar contents in RFH-D1, RFH-D2, and RFH-D3 were 93.8 wt%, 77.9 wt%, and 86.3 wt%, respectively.

[0049] Experimental Example: Determination of the immunomodulatory activity of the polysaccharide extract of *Ficus hirta* (including RFH, RFH-D1, RFH-D2, and RFH-D3) described in this invention.

[0050] 1. The CCK-8 assay was used to detect the proliferative activity of polysaccharide extract from *Ficus hirta* on RAW264.7 cells (macrophages).

[0051] 1.1 Proliferative activity of RFH prepared according to the method described in Example 1 and RFH-D1, RFH-D2 and RFH-D3 prepared according to the method described in Example 4.

[0052] RAW264.7 cells were cultured in DMEM medium containing 10% FBS, 100 U / mL penicillin, and 100 μg / mL streptomycin, and incubated at 37°C in a 5% CO2 humidified incubator. Cells were then sputtered at a rate of 1 × 10⁻⁶ cells / mL. 5 Cells / mL were seeded in 96-well plates and cultured. After culturing, the cells were treated with 100 μL of culture medium (blank control group) or different concentrations of crude polysaccharide RFH and purified polysaccharides RFH-D1, RFH-D2, and RFH-D3 (20, 40, 60, 80, and 100 μg / mL) for 24 h. CCK-8 reagent was added along the wall of each well at a concentration of 10 μL, and the plates were shaken up and down and left and right. The absorbance was measured at 450 nm after 1 h. The results are shown in Table 1.

[0053] Table 1. Influence of different parts of *Ficus hirta* polysaccharide on the proliferation rate of RAW264.7 cells.

[0054]

[0055]

[0056] As shown in Table 1, within the concentration range of 20–100 μg / mL, the RFH, RFH-D1, RFH-D2, and RFH-D3 treatment groups were all non-toxic to macrophages and could promote cell proliferation.

[0057] 1.2 Proliferative activity of RFH1-1 and RFH1-2 prepared according to the methods described in Comparative Examples 1-1 and 1-2

[0058] The activity test method is the same as described in 1.1 above, and the results are shown in Tables 2 and 3, respectively.

[0059] Table 2. Influence of RFH1-1 polysaccharide on the proliferation rate of RAW264.7 cells.

[0060]

[0061] As shown in Table 2, the polysaccharide RFH1-1 of *Ficus hirta* in the concentration range of 20–100 μg / mL promoted the proliferation of RAW264.7 cells without toxicity.

[0062] Table 3. Influence of the alkaline polysaccharide RFH1-2 from *Ficus hirta* on the proliferation rate of RAW264.7 cells.

[0063]

[0064] As shown in Table 3, the RFH1-2 treatment group was not toxic to macrophages in the concentration range of 20–100 μg / mL, and promoted cell proliferation at a concentration of 100 μg / mL.

[0065] 2. The effect of *Ficus hirta* polysaccharide extract on NO release from RAW264.7 cells was determined using a NO kit.

[0066] 2.1 Effects of RFH prepared according to the method described in Example 1 and RFH-D1, RFH-D2 and RFH-D3 prepared according to the method described in Example 4 on NO release from RAW264.7 cells.

[0067] RAW264.7 cells were added to a 24-well plate (1×10⁻⁶ cells per well). 5 In a 96-well plate, cells were treated with different concentrations of crude and purified polysaccharides of *Ficus hirta* for 24 hours after reaching 60% confluence. A blank control group (NC) and a lipopolysaccharide (LPS) positive control group were also included. The blank control group received 100 μL of culture medium, while the positive control group received 1 μg / mL LPS. 50 μL of supernatant was collected from each well and transferred to a 96-well plate according to the kit manufacturer's instructions. 50 μL each of Griess A and Griess B reagents were added to each well, and the mixture was shaken to mix. The OD value was measured at 570 nm. The NO secretion level in the culture medium was calculated based on the prepared NaNO2 standard curve. The results are shown in Table 4 (RFH was tested alone; RFH-D1, RFH-D2, and RFH-D3 were tested on the same plate).

[0068] Table 4. Effects of different parts of *Ficus hirta* polysaccharide on NO release from RAW264.7 cells.

[0069]

[0070] Note: Compared with NC1 group, ***P<0.001; compared with NC2 group, RFH-D1, RFH-D2, and RFH-D3, **P<0.01, ***P<0.001.

[0071] NO is a messenger molecule involved in immune inflammation, vasodilation, and nerve conduction, and is an important immunomodulatory factor. Table 4 shows that, compared with the blank control group (NC), NO levels significantly increased with increasing polysaccharide concentrations in different parts of *Ficus hirta*. The high-concentration groups of RFH, RFH-D1, and RFH-D3 showed stronger NO secretion than the 1 μg / mL LPS group. Appropriate amounts of NO contribute to the fine regulation of immune responses, enhance non-specific immune function, and inhibit specific immune responses.

[0072] 2.2 Effects of RFH1-1 and RFH1-2 prepared according to the methods described in Comparative Examples 1-1 and 1-2 on NO release from RAW264.7 cells

[0073] The activity test method is the same as described in section 2.1 above, and the results are shown in Tables 5 and 6, respectively.

[0074] Table 5. Effects of RFH1-1 polysaccharide from *Ficus hirta* on NO release from RAW264.7 cells.

[0075]

[0076] Note: Compared with the NC group, *P<0.05; **P<0.01; ***P<0.001;

[0077] As shown in Table 5, although the polysaccharide RFH1-1 of Prunus pedatus at concentrations of 20 and 40 μg / mL could significantly promote the release of NO factor from RAW264.7 cells, the promoting effect was weaker compared to the LPS group.

[0078] Table 6. Effects of the alkaline polysaccharide RFH1-2 from *Ficus hirta* on NO release from RAW264.7 cells.

[0079]

[0080] Note: Compared with the NC group, ***P<0.001;

[0081] As shown in Table 6, compared with the blank control group (NC), the alkaline polysaccharide RFH1-2 of Prunus pedunculata did not promote the release of NO from cells, and had no or very weak activation effect on macrophages.

[0082] 3. Effects of the polysaccharide extracts of *Ficus hirta* prepared according to the method described in Example 1, and RFH-D1, RFH-D2, and RFH-D3 prepared according to the method described in Example 4, on cytokines.

[0083] Cytokines, known as biological response modulators, include interleukin (IL-6) and tumor necrosis factor (TNF-α), which regulate inflammation, immunity, and defense against external pathogens. When the body is infected or subjected to other external threats, IL-6 can promote the proliferation and differentiation of white blood cells, activate immune cells, and accelerate the initiation of the immune response.

[0084] Cells at 1×10 5 Cells were seeded into well plates. When the cells reached 60% confluence, they were pre-incubated with drugs for 24 hours according to different groups. The supernatant was collected, and the contents of IL-6 and TNF-α in the cell culture medium were detected using an ELISA kit. The results are shown in Tables 7 and 8 (RFH, RFH-D1, and RFH-D2 were tested on one plate, and RFH-D3 was tested separately).

[0085] Table 7. Effects of different parts of *Ficus hirta* polysaccharide on the secretion of IL-6 by RAW264.7 cells.

[0086]

[0087] Note: When comparing RFH, RFH-D1, and RFH-D2 with NC1, **P < 0.01, ***P < 0.001; when comparing RFH-D3 with NC2, **P < 0.01, ***P < 0.001.

[0088] Table 8. Effects of different parts of *Ficus hirta* polysaccharide on the content of TNF-α secreted by RAW264.7 cells.

[0089]

[0090]

[0091] Note: Compared with group NC1, *P<0.05, **P<0.01, ***P<0.001 for RFH, RFH-D1, and RFH-D2; compared with group NC2, **P<0.01, ***P<0.001 for RFH-D3.

[0092] As shown in Tables 7 and 8, compared with the blank control group, the polysaccharide extract of *Ficus hirta* significantly increased the concentration-dependent IL-6 secretion by macrophages. Compared with the positive control group, the high-concentration RFH and RFH-D1 groups showed stronger IL-6 secretion levels. On the other hand, TNF-α can activate and enhance macrophages through stimulation, thereby improving the body's immune function. As shown in Tables 7 and 8, while the polysaccharide concentration in various parts of *Ficus hirta* continuously increased, the TNF-α content also showed a significant upward trend. The high-concentration RFH group secreted stronger TNF-α than the positive control group. Therefore, the polysaccharide extract of *Ficus hirta* can effectively activate macrophages to produce immune activity.

Claims

1. Five-finger peach polysaccharide extract, which is crude or purified polysaccharide from five-finger peach, wherein, The crude polysaccharide of *Ficus hirta* is obtained by first alcohol precipitation, removal of protein by Sevage method and second alcohol precipitation of water extract of *Ficus hirta*, followed by drying. It is denoted as RFH and has a total sugar content of ≥39.0 wt%. The purified polysaccharides were obtained by DEAE-52 fiber column chromatography on RFH, eluting with sodium chloride solution. The eluted fractions of 0.3 mol / L, 0.4 mol / L, and 0.5 mol / L sodium chloride solutions were collected, and then dialyzed and dried respectively. The purified polysaccharides were designated as RFH-D1, RFH-D2, and RFH-D3, respectively, with total sugar contents of 90.0–95.0 wt%, 70.0–79.0 wt%, and 80.0–87.0 wt%, respectively.

2. The polysaccharide extract of *Ficus hirta* according to claim 1, characterized in that, The total sugar content of the crude polysaccharide from *Ficus hirta* is 39.5–49.0 wt%. The total sugar contents of the purified polysaccharides RGH-D1, RGH-D2, and RGH-D3 were 91.2–93.8 wt%, 72.3–77.9 wt%, and 81.4–86.3 wt%, respectively.

3. The method for preparing the polysaccharide extract of *Ficus hirta* according to claim 1, comprising a method for preparing crude polysaccharide from *Ficus hirta* and a method for preparing purified polysaccharide, wherein, The preparation method of crude polysaccharide of Prunus pubescens includes: subjecting the water extract of Prunus pubescens to one alcohol precipitation, Sevage method to remove protein and a second alcohol precipitation, and then drying it to obtain the product, denoted as RFH, wherein the total sugar content is ≥39.0wt%. The method for preparing purified polysaccharides includes: chromatography on a DEAE-52 fiber column on an RFH column, eluting with sodium chloride solution as the eluent, collecting the eluted fractions from 0.3 mol / L, 0.4 mol / L, and 0.5 mol / L sodium chloride solutions, respectively, and then dialysis and drying them to obtain the purified polysaccharides, which are respectively designated as RFH-D1, RFH-D2, and RFH-D3, with total sugar contents of 90.0–95.0 wt%, 70.0–79.0 wt%, and 80.0–87.0 wt%, respectively.

4. The preparation method according to claim 3, characterized in that, In the preparation method of the crude polysaccharide of Prunus pubescens, the alcohol used for alcohol precipitation is 95-100 v / v ethanol.

5. The preparation method according to claim 3, characterized in that, In the preparation method of crude polysaccharide from *Ficus hirta*, the water extract of *Ficus hirta* is obtained by extracting *Ficus hirta* with water, filtering, collecting the filtrate, and concentrating it.

6. The preparation method according to claim 5, characterized in that, The extraction method is either reflux extraction or ultrasonic extraction.

7. The preparation method according to claim 3, characterized in that, In the method for preparing the purified polysaccharide, the molecular weight cutoff for dialysis is greater than or equal to 3500 Da.

8. The preparation method according to any one of claims 3 to 7, characterized in that, The total sugar content of the crude polysaccharide from *Ficus hirta* is 39.5–49.0 wt%. The total sugar contents of the purified polysaccharides RGH-D1, RGH-D2, and RGH-D3 were 91.2–93.8 wt%, 72.3–77.9 wt%, and 81.4–86.3 wt%, respectively.

9. The use of the polysaccharide extract of *Ficus hirta* according to claim 1 in the preparation of immune-modulating foods or medicines.

10. A food or pharmaceutical composition comprising the polysaccharide extract of *Ficus hirta* as described in claim 1 and a pharmaceutically acceptable carrier.

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