A polysaccharide of taxodin ganoderma lucidum with efficacy of relieving colorectal cancer and a preparation method and application thereof

Abstract

This invention discloses a polysaccharide from *Ganoderma lucidum* (also known as *Ganoderma lucidum* or *Ganoderma sinense*) with efficacy in alleviating colorectal cancer, its preparation method, and its applications, belonging to the field of biomedical technology. The polysaccharide is composed of mannose, glucose, galactose, and fucose, with a molar ratio of 3.221:1.414:80.539:14.826 and a molecular weight of 7.056 kDa. It has been confirmed that the polysaccharide can significantly reduce the number and size of colon tumors; alleviate pathological symptoms such as colonic weight index, bleeding, crypt budding, dysplasia, and hyperplasia; and significantly inhibit the levels of IFN-γ, TGF-β, and IL-17A in the colon while increasing the level of IL-4. Therefore, the polysaccharide provided by this invention has a significant effect in alleviating colorectal cancer, and this invention provides theoretical support for the development of effective and biosafe colorectal cancer drugs.

Description

Technical Field

[0001] This invention relates to the field of biomedicine, and in particular to a polysaccharide from Ganoderma lucidum that has the effect of alleviating colorectal cancer, its preparation method, and its application. Background Technology

[0002] Colorectal cancer is the second leading cause of cancer-related deaths worldwide. It is estimated that by 2040, the cancer burden will increase to 3.2 million new cases annually (a 63% increase) and 1.6 million deaths annually (a 73% increase) (World Health Organization, 2022). In addition to population aging and dietary habits, adverse risk factors such as obesity, lack of physical activity, and smoking also increase the risk of cancer. In recent years, with economic development, population aging, and changes in lifestyle, its incidence and mortality rates have been on the rise. According to the latest report from the National Cancer Center, colorectal cancer ranks second in incidence and fifth in mortality among all malignant tumors in my country. Early-stage cancer has a higher survival rate than late-stage cancer; however, colorectal cancer is often diagnosed at an advanced stage, leading to increased mortality. Surgery, radiation therapy, and chemotherapy are currently used to treat colorectal cancer. Patients receiving frequent chemotherapy are prone to developing drug resistance, which may lead to poor treatment outcomes and harmful side effects on normal cells. Therefore, it is necessary to develop more effective and safer methods for treating colorectal cancer.

[0003] In traditional Chinese medicine, Ganoderma lucidum (also known as pine-fir or cypress-leaf ganoderma) is frequently used to improve health, increase vitality, and prolong life. The *Illustrated Compendium of Chinese Medicinal Fungi* records that Ganoderma lucidum strengthens the body and nourishes the spirit; in folk medicine, it is often soaked in alcohol and consumed for its effects of benefiting the heart, calming the mind, tonifying the liver, strengthening bones and muscles, and beautifying the complexion. Polysaccharides are among the most recognized edible and medicinal fungal derivatives, possessing a series of important biological characteristics. Ganoderma lucidum polysaccharides are safe, harmless, and have few toxic side effects; increasingly, researchers are dedicating themselves to developing safe, economical, and effective natural medicines using Ganoderma lucidum polysaccharides. Summary of the Invention

[0004] The purpose of this invention is to provide a Ganoderma lucidum polysaccharide with efficacy in alleviating colorectal cancer, its preparation method, and its application, in order to solve the problems existing in the prior art. This invention extracts and purifies a new Ganoderma lucidum polysaccharide with outstanding effects in alleviating colorectal cancer. This invention provides theoretical support for the development of colorectal cancer drugs with efficacy and biosafety.

[0005] To achieve the above objectives, the present invention provides the following solution:

[0006] This invention provides a Ganoderma lucidum polysaccharide containing pine and fir trees that has the effect of alleviating colorectal cancer. The Ganoderma lucidum polysaccharide contains mannose, glucose, galactose and fucose. The molar ratio of mannose, glucose, galactose and fucose is 3.221:1.414:80.539:14.826.

[0007] The structure of the Ganoderma lucidum polysaccharide from Pinus sylvestris is as follows:

[0008]

[0009] Optionally, the preparation method of the Ganoderma lucidum polysaccharide includes the following steps:

[0010] After drying, ultra-fine pulverization, and sieving, the fruiting bodies of Ganoderma lucidum were added to deionized water, ultrasonically extracted, centrifuged to collect the supernatant, concentrated, and protein removed. The supernatant was then concentrated again, precipitated using an ethanol aqueous solution, the precipitate was collected, and freeze-dried to obtain crude polysaccharide from Ganoderma lucidum.

[0011] The crude polysaccharide of Ganoderma lucidum was dissolved, eluted with diethylaminoethyl agarose gel, and the polysaccharide solution was collected, concentrated, dialyzed, lyophilized, and then purified by chromatography to obtain the polysaccharide of Ganoderma lucidum.

[0012] Optionally, the eluent for the diethylaminoethyl agarose gel is water and sodium chloride solution, and the flow rate is 1 mL / min. Optionally, the purification includes a first purification using a HiPrep 26 / 60 Sephacryl S-400HR preparative grade column, followed by a second purification using a HiLoad 16 / 600 Superdex 200 pg preparative grade column.

[0013] Optionally, the eluent for the HiPrep 26 / 60 Sephacryl S-400HR preparative chromatographic column is a 0.15 mol / L sodium chloride solution; the eluent for the HiLoad 16 / 600 Superdex 200 pg preparative chromatographic column is a 0.15 mol / L sodium chloride solution.

[0014] Optionally, the ultrasound power is 300W and the duration is 1 hour.

[0015] The present invention also provides the application of the aforementioned Ganoderma lucidum polysaccharide in the preparation of a drug for treating colorectal cancer.

[0016] The present invention also provides a drug for treating colorectal cancer, wherein the active ingredient of the drug includes the aforementioned Ganoderma lucidum polysaccharide.

[0017] Optionally, the drug may also include pharmaceutically acceptable carrier substances and / or excipients.

[0018] Optionally, the excipients include at least one of the following: absorbent, diluent, filler, excipient, binder, humectant, disintegrant, surfactant, adsorbent carrier, lubricant, and flavoring agent.

[0019] The present invention discloses the following technical effects:

[0020] This invention extracts and purifies a new Ganoderma lucidum polysaccharide composed of mannose, glucose, galactose and fucose, with a molar ratio of mannose, glucose, galactose and fucose of 3.221:1.414:80.539:14.826. The molecular weight of the Ganoderma lucidum polysaccharide is 7.056 kDa. Cellular and animal experiments have confirmed that Ganoderma lucidum polysaccharide has a significant inhibitory effect on colorectal cancer: it significantly reduces the number and size of colon tumors; alleviates pathological symptoms such as colonic weight index, bleeding, crypt budding, dysplasia, and hyperplasia; and significantly inhibits the levels of IFN-γ (18.46%, p<0.05), TGF-β (20.80%, p<0.01), and IL-17A (24.98%, p<0.01) in the colon, while increasing the level of IL-4. Therefore, the Ganoderma lucidum polysaccharide provided by this invention has a prominent effect in alleviating colorectal cancer, and this invention provides theoretical support for the further development of effective and biosafe colorectal cancer drugs. Attached Figure Description

[0021] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0022] Figure 1 Elution curve of DEAE Sepharose crude polysaccharide from Ganoderma lucidum;

[0023] Figure 2 Statistical graph showing the inhibition of SW480 cell growth by GTP-a and GTP-c;

[0024] Figure 3 Elution curves of water-washed polysaccharides from Ganoderma lucidum (Pine and Fir) HiPrep 26 / 60 Sephacryl S-400HR (A) and HiLoad 16 / 600 Superdex 200pg (B);

[0025] Figure 4 The absolute molecular weight analysis chromatogram of GTP-a2;

[0026] Figure 5A diagram showing the monosaccharide composition analysis of purified polysaccharides from Ganoderma lucidum (Pine and Fir Trees);

[0027] Figure 6 This is the total ion chromatogram of the sample;

[0028] Figure 7 It has a GTP-a2 structure;

[0029] Figure 8 To investigate the role of GTP-α2 in inhibiting the development of AOM / DSS-induced colorectal tumors in mice; A: Colorectal length in each group; B: HE staining; C: Statistical graph of tumor number in each group; D: Colon weight index;

[0030] Figure 9 GTP-α2 regulates AOM / DSS-induced colonic metabolites in mice; A: Colonic metabolites; B: Principal component analysis plot; C: Volcano plot;

[0031] Figure 10 Clustering heatmap of GTP-α2-regulated AOM / DSS-induced mouse colonic metabolites;

[0032] Figure 11 Differential abundance score map for GTP-a2-regulated AOM / DSS-induced KEGG enrichment metabolic pathway in mouse colon;

[0033] Figure 12 This is an immunohistochemical staining image;

[0034] Figure 13 The diagram shows the expression of labeled proteins; A: IFN-γ; B: IL-4; C: TGF-β; D: IL-17A. Detailed Implementation

[0035] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0036] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Any stated value or intermediate value within a stated range, as well as each smaller range between any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0037] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0038] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be readily apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0039] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0040] Example 1

[0041] Using Ganoderma lucidum as raw material, this study provides a method for preparing and purifying polysaccharides with anti-colorectal cancer properties. The method is characterized by tumor number and intestinal tissue metabolites as evaluation indicators, and its activity in alleviating colorectal cancer is investigated.

[0042] 1. Preparation of crude polysaccharides from Ganoderma lucidum and Ganoderma sinense

[0043] The fruiting bodies of *Ganoderma lucidum* were dried to constant weight in an oven at 80℃, then ultra-finely pulverized into powder and passed through a 150-mesh sieve. Deionized water was added at a material-to-liquid ratio of 1g:70mL, and extraction was performed using ultrasound at 300W for 1 hour. After centrifugation, the supernatant was collected and concentrated to 1 / 30 of its volume using rotary evaporation. Protein was removed using the Sevag method. The supernatant was concentrated and precipitated using 75% (v / v) ethanol overnight at 4℃. The precipitate was collected and freeze-dried to obtain crude polysaccharide from *Ganoderma lucidum*.

[0044] 2. Preparation of purified polysaccharides from Ganoderma lucidum and Ganoderma lucidum

[0045] (1) The crude polysaccharide of Ganoderma lucidum was dissolved in deionized water and eluted with a gradient of purified water, 0.1M and 0.3M sodium chloride solution at a flow rate of 1 mL / min using a DEAE Sepharose Fast Flow gel. The solutions were collected separately, and the polysaccharide content was determined by the phenol-sulfuric acid method. Different polysaccharide fractions (GTP-a, GTP-c, and GTP-b was not tested due to its low yield) were collected, concentrated to 1 / 5 volume by rotary evaporation, dialyzed overnight at 4℃, and then lyophilized for collection. Figure 1 ).

[0046] Colorectal cancer cells (SW480) in logarithmic growth phase were seeded into 96-well plates (8 × 10⁶ cells / well). 3 Cells were cultured for 8 hours at 100 cells / well. Then, the cells were cultured for another 24 hours with GTP-a and GTP-c at concentrations of 0.08, 0.16, 0.31, 0.63, 1.25, and 2.50 mg / mL. Cell viability was determined using the MTT assay. Both GTP-a and GTP-c showed inhibitory effects on SW480 cell growth, with half-maximal inhibitory concentrations (IC50) of 100%. 50 The values ​​were 1.0 mg / mL and 1.96 mg / mL, respectively. Figure 2 Among them, GTP-a significantly inhibited the growth of SW480 cells.

[0047] (2) GTP-a obtained by DEAE gel column chromatography was dissolved in deionized water and purified for the first time using a HiPrep 26 / 60 Sephacryl S-400HR preparative grade column. The solution was eluted with 0.15 mol / L sodium chloride solution, collected in separate tubes, and the polysaccharide content was determined using the phenol-sulfuric acid method. The main peak was collected, concentrated to 1 / 5 volume by rotary evaporation, dialyzed overnight at 4℃, and then lyophilized to obtain purified Ganoderma lucidum polysaccharide (GTP-a1). Finally, a second purification was performed using a HiLoad 16 / 600 Superdex 200 pg preparative grade column. The solution was eluted with 0.15 mol / L sodium chloride solution, collected in separate tubes, and the polysaccharide content was determined using the phenol-sulfuric acid method. The main peak was collected, concentrated to 1 / 6 volume by rotary evaporation, dialyzed overnight at 4℃, and then lyophilized to obtain purified Ganoderma lucidum polysaccharide (GTP-a2). Figure 3 ).

[0048] 3. Structural characteristics analysis of purified polysaccharides from Ganoderma lucidum (Pine and Fir)

[0049] (1) Polysaccharide molecular weight analysis: The sample was dissolved in 0.1M NaNO3 aqueous solution (containing 0.02% NaN3, w / w) to a final concentration of 1 mg / mL, and filtered through a 0.45 μm filter before analysis. A gel permeation chromatography (Agilent, 1260 Infinity II MDS) was used. The column was PL aquagel-OH Mixed-H, 8 μm, 7.5 × 300 mm, with a flow rate of 1.0 mL / min and a column temperature of 45℃. The injection volume was 50 μL, with 0.1 mol / L sodium nitrate (0.01% sodium azide) as the mobile phase, and isocratic elution was performed. The molecular weights Mp, Mn, Mw, and Mz of the sample were calculated using GPC software based on the sample viscosity and elution time. The molecular weight of GTP-a2 was 7.056 kDa ( Figure 4 The multi-dispersion index is 1.035 (Table 1).

[0050] Table 1. Molecular characteristics of GTP-a2

[0051]

[0052] (2) Monosaccharide composition analysis: Take a clean chromatographic bottle, accurately weigh 5 mg (±0.05 mg) of polysaccharide sample, add 1 mL of 2M trifluoroacetic acid solution, and heat at 121℃ for 2 hours. Purge with nitrogen and dry. Add 3 mL of methanol to wash, then dry again, repeating the methanol washing 2-3 times. Add 5 mL of sterile water to dissolve, and transfer to a chromatographic bottle for analysis. Take 0.2 mL of monosaccharide standard solution or polysaccharide hydrolysate into a stoppered conical centrifuge tube, add 0.2 mL of 0.5 mol / L sodium hydroxide solution and 0.5 mL of 0.5 mol / L 1-phenyl-3-methyl-5-pyrazolone (PMP) methanol solution, vortex to mix, and react in a 70℃ water bath for 1 h. After the reaction is complete, add 0.2 mL of 0.5 mol / L hydrochloric acid to neutralize the added sodium hydroxide, add 1 mL of chloroform, vortex extract 3 times to remove excess PMP, discard the chloroform layer, and take 0.3 mL and dilute with water to 1 mL. A Thermo U3000 liquid chromatography system was used with a ZORBAX Eclipse XDB-C18 column. The mobile phase was acetonitrile: phosphate buffer (12 g / L potassium dihydrogen phosphate, adjusted to pH 6.8 with 2 M NaOH) with isocratic elution at a volume ratio of 17:83. The flow rate was 0.8 mL / min, the column temperature was 30 °C, the detection wavelength was 250 nm, and the injection volume was 10 μL. Results are as follows: Figure 5 As shown, GTP-a2 is composed of mannose (Man), glucose (Glc), galactose (Gal), and fucose (Fuc), with a molar ratio (mol%) of 3.221:1.414:80.539:14.826.

[0053] (3) Polysaccharide methylation analysis: 2 mg of sample was dissolved in 1 mL of DMSO; 30 mg of NaOH was added and incubated for 30 min; 250 μL of iodomethane solution was added, nitrogen was introduced, and the reaction was carried out in the dark for 1 h; 250 μL of iodomethane solution was added again and the reaction was carried out for 1 h; 1 mL of water and 2 mL of dichloromethane were added, vortexed and mixed, centrifuged, and the aqueous phase was discarded; the lower dichloromethane phase was aspirated and dried under nitrogen; 1 mL of 2M TFA was added and the reaction was carried out at 121 °C for 120 min; the reaction was dried under nitrogen at 30 °C; 1 mL of freshly prepared MNaBD4 (ammonia water) was added. Incubate with magnetic stirring at room temperature for 2.5 hours; terminate the reaction by adding 300 μL of acetic acid and dry under nitrogen; dry twice with 2 mL of 5% (vol / vol) acetic acid in methanol at 40 °C under nitrogen, then dry twice with 2 mL of methanol under nitrogen at 40 °C; add 1.5 mL of acetic anhydride, vortex to mix, and react at 100 °C for 2.5 h; add 2 mL of water and let stand for 10 min; add 1 mL of dichloromethane, vortex to mix, centrifuge, and discard the aqueous phase; take the lower dichloromethane phase and analyze it. The chromatographic system used was an Agilent gas chromatography system (Agilent 7890A; Agilent Technologies, USA), with an HP-5MS capillary column (30m × 0.25mm × 0.25μm, Agilent J&W Scientific, Folsom, CA, USA). High-purity helium (purity not less than 99.999%) was used as the carrier gas. The flow rate was 1.0 mL / min, and the injection port temperature was 260℃. The injection volume was 1 μL, injected as a split at a split ratio of 10:1, with a solvent delay of 2.2 min. The temperature was held at 50℃ for 1.0 min, then increased at 50℃ / min to 130℃, and then increased at 3℃ / min to 230℃, holding for 2 min. Based on retention time and standard data for partially methylated sugar alcohol acetates (PMAA) from the Center for Complex Carbohydrate Research (CCRC) spectral database, GTP-a2 is primarily composed of 11 glycosidic fragments (Table 2): t-Fuc(p) (19.98%), t-Glc(p) (0.43%), t-Man(p) (1.91%), t-Gal(p) (2.10%), 2-Man(p) (0.76%), 4-Glc(p) (0.42%), 2-Gal(p) (2.21%), 6-Glc(p) (1.45%), 6-Gal(p) (50.24%), 4,6-Gal(p) (0.50%), and 2,6-Gal(p) (20.00%). Figure 6 Based on the monosaccharide composition and linkage, the structure of GTP-a2 is inferred to be →6-α-D-Galp-(1→ backbone, α-L-Fucp-(1→ linked at C2) ( ). Figure 7 ).

[0054] Table 2. Results of GTP-α2 methylation analysis

[0055]

[0056] 4. Study on the activity of GTP-a2 in alleviating colorectal cancer

[0057] (1) Mouse grouping and administration: Thirty male C57BL / 6 mice (6-8 weeks old; 18-20g) were housed at a temperature of 23±1℃ and a relative humidity of 55±5%. After a one-week acclimatization period, 20 mice were intraperitoneally injected with 10mg / kg AOM (azomethane), and their drinking water was changed to 2% DSS at weeks 1, 4, and 7. Four weeks later, the mice were randomly divided into two groups (n=10): the AOM / DSS group, which was given double-distilled water by gavage, and the GTP-a2-treated AOM / CSS group, which was given GTP-a2 (100mg / kg) daily. After receiving intraperitoneal saline injection on the first day of the experiment, the 10 mice in the normal group received double-distilled water daily for 6 weeks.

[0058] (2) Sample preparation: After the last administration, all mice were fasted overnight and euthanized by inhaling carbon dioxide after blood was drawn from the tail vein. Colon tissue samples were collected from each mouse to examine the size and number of tumors. A portion of these tissues were fixed in 4% paraformaldehyde solution for future studies, while the remainder was stored at -80°C.

[0059] (3) Histopathological observation: Intestinal tissue was fixed in 4% paraformaldehyde and stored at 4°C. It was routinely embedded in paraffin, sectioned, stained with hematoxylin and eosin (HE) and immunohistochemically, and observed under an optical microscope for pathological analysis.

[0060] (4) Intestinal samples were randomly selected from the normal group, AOM / DSS group, and AOM / DSS+GTP-2 group (n=5 / group). Untargeted metabolomics analysis was performed using ultra-high performance liquid chromatography (1290 Infinity LC; Agilent Technologies) combined with time-of-flight and Orbitrap mass spectrometry (AB Triple TOF 6600Q; AB SCIEX, MA, USA). The structures of metabolites in the biological samples were determined by matching the retention time, molecular weight (molecular weight error <10 ppm), secondary fragmentation spectra, and collision energies of metabolites in the local database. The processed data were subjected to orthogonal partial least squares discriminant analysis (OPLS-DA) for metabolite variation analysis. Statistically significant differences in metabolites (p<0.05) and a projected variable importance (VIP) value >2.0 were considered as the standard values ​​for differentially expressed metabolites.

[0061] (5) Immunohistochemical analysis: After sealing the slides from (3), they were incubated with an antibody (IL-17A, TGF-β, IL-4, IFN-γ) at 4°C for 16 hours. Then, the slides were incubated with goat anti-rabbit secondary antibody at 4°C for 4 hours. Finally, all slides were observed under an optical microscope, and the obtained images were quantified using ImageJ software.

[0062] (6) Experimental Results

[0063] 6.1 GTP-α2 inhibits the development of AOM / DSS-induced colorectal tumors in mice

[0064] Following GTP-a2 administration, a significant reduction in both the number and size of colon tumors was observed. Figure 8 (C). Furthermore, colonic weight index and bleeding were relieved ( Figure 8 China A Figure 8 (D). Histological examination revealed complex crypt budding, dysplasia, and hyperplasia in the colon of AOM / DSS-induced mice, while these histopathological changes were observed after GTP-a2 treatment. Figure 8 (B)

[0065] 6.2 GTP-α2 regulates metabolite levels in the colon of mice induced by AOM / DSS.

[0066] A total of 576 metabolites were identified in the colon, including lipids and lipid molecules (31.0%), organic acids and derivatives (30.9%), and organic heterocyclic compounds (12.90%). Figure 9 (A). Principal component analysis (PCA) plots showed that GTP-a2 significantly altered the levels of metabolites in the colon. Figure 9 (B) Compared with the normal group, 348 metabolites changed in the AOM / DSS group, and 304 metabolites changed in the GTP-a2-treated AOM / CSS group. Furthermore, compared with the AOM / DSS group, mice treated with GTP-a2 showed increased levels of 17 metabolites and decreased levels of 32 metabolites. Figure 9 (C). Following GTP-a2 treatment, 20 metabolic pathways related to protein digestion and absorption, cancer-centric carbon metabolism, alanine, aspartate, and glutamate metabolism, as well as mineral absorption, were also altered. Figure 10 Based on VIP scores >1.5 and p<0.05, there were significant differences in the levels of 30 metabolites among the three groups. Figure 11 ).

[0067] 6.3 GTP-α2 regulates immune function and alleviates colorectal cancer

[0068] Immunohistochemical analysis showed that, compared with the normal group, the expression of IFN-γ (p<0.01), TGF-β (p<0.01), and IL-17A (p<0.01) in the colon was enhanced in AOM / DSS-induced mice. Compared with AOM / DSS-induced mice, GTP-a2 treatment significantly inhibited the levels of IFN-γ (18.46%, p<0.05), TGF-β (20.80%, p<0.01), and IL-17A (24.98%, p<0.01) in the colon, while increasing the level of IL-4 (19.42%, p<0.01). These findings indicate a decrease in Th1 and Th17 cells and an increase in Th2 cells. Figure 12 , Figure 13 ).

[0069] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A polysaccharide from Ganoderma lucidum (including pine and fir trees) with efficacy in alleviating colorectal cancer, characterized in that... The Ganoderma lucidum polysaccharide is composed of mannose, glucose, galactose and fucose; the molar ratio of mannose, glucose, galactose and fucose is 3.221:1.414:80.539:14.

826. The structure of the Ganoderma lucidum polysaccharide from Pinus sylvestris is as follows:

2. The Ganoderma lucidum polysaccharide as described in claim 1, characterized in that, The preparation method of the pine and fir Ganoderma lucidum polysaccharide includes the following steps: After drying, ultra-fine pulverization, and sieving, the fruiting bodies of Ganoderma lucidum were added to deionized water, ultrasonically extracted, centrifuged to collect the supernatant, concentrated, and protein removed. The supernatant was then concentrated again, precipitated using an ethanol aqueous solution, the precipitate was collected, and freeze-dried to obtain crude polysaccharide from Ganoderma lucidum. The crude polysaccharide of Ganoderma lucidum was dissolved, eluted with diethylaminoethyl agarose gel, and the polysaccharide solution was collected, concentrated, dialyzed, freeze-dried, and then purified by chromatography to obtain the polysaccharide of Ganoderma lucidum. The elution buffer for the diethylaminoethyl agarose gel is water and sodium chloride solution, with a flow rate of 1 mL / min.

3. The Ganoderma lucidum polysaccharide as described in claim 2, characterized in that, The purification process includes a first purification using a HiPrep 26 / 60 Sephacryl S-400 HR preparative grade column, followed by a second purification using a HiLoad 16 / 600 Superdex 200pg preparative grade column.

4. The Ganoderma lucidum polysaccharide as described in claim 3, characterized in that, The eluent for the HiPrep 26 / 60 Sephacryl S-400 HR preparative chromatographic column is 0.15 mol / L sodium chloride solution; the eluent for the HiLoad 16 / 600 Superdex 200 pg preparative chromatographic column is 0.15 mol / L sodium chloride solution.

5. The Ganoderma lucidum polysaccharide as described in claim 2, characterized in that, The ultrasound power was 300W, and the duration was 1 hour.

6. The use of the Ganoderma lucidum polysaccharide as described in any one of claims 1 to 5 in the preparation of a medicament for treating colorectal cancer.

7. A drug for treating colorectal cancer, characterized in that, The active ingredient of the drug includes the Ganoderma lucidum polysaccharide as described in any one of claims 1 to 5.

8. The drug as described in claim 7, characterized in that, The drug also includes pharmaceutically acceptable excipients.

9. The medicament as described in claim 8, characterized in that, The excipients include at least one of the following: absorption promoter, diluent, filler, excipient, binder, humectant, disintegrant, surfactant, adsorbent carrier, lubricant, and flavoring agent.

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