Ganoderma tsugae polysaccharide with effect of relieving colorectal cancer as well as preparation method and application of ganoderma tsugae polysaccharide

Abstract

The invention discloses ganoderma tsugae polysaccharide with a colorectal cancer relieving effect and a preparation method and application thereof, and belongs to the technical field of biological medicine. The ganoderma tsugae polysaccharide is prepared from mannose, glucose, galactose and fucose, the molar ratio of the mannose to the glucose to the galactose to the fucose is 3.221 to 1.414 to 80.539 to 14.826, and the molecular weight of the ganoderma tsugae polysaccharide is 7.056 kDa. It is proved that the ganoderma tsugae polysaccharide can significantly reduce the number and size of colon tumors; the pathological symptoms such as colon weight index, bleeding, crypt budding, dysplasia and hyperplasia are relieved; compared with the prior art, the ganoderma tsugae polysaccharide provided by the invention has the advantages that the ganoderma tsugae polysaccharide can obviously inhibit the levels of IFN-gamma, TGF-beta and IL-17A in colon and also can obviously inhibit the levels of IFN-gamma, TGF-beta and IL-17A in colon, and meanwhile, the level of IL-4 is improved, so that the ganoderma tsugae polysaccharide provided by the invention has an outstanding action effect in the aspect of relieving colorectal cancer, and a theoretical support is provided for developing colorectal cancer medicines with effectiveness and biological safety.

Description

Technical Field

[0001] The present invention relates to the technical field of biomedicine, and in particular to a pine ganoderma lucidum polysaccharide having the effect of alleviating colorectal cancer, and a preparation method and application thereof. Background Art

[0002] Colorectal cancer is the second leading cause of cancer-related death worldwide. It is estimated that by 2040, the burden of cancer will increase to 3.2 million new cases per year (an increase of 63%) and 1.6 million deaths per year (an increase of 73%) (World Health Organization, 2022). In addition to the aging population and dietary habits, adverse risk factors such as obesity, lack of physical exercise and smoking also increase the risk of cancer. In recent years, with economic development, aging population and changes in lifestyle, its incidence and mortality have shown an upward trend. According to the latest report from the National Cancer Center, the incidence and mortality of colorectal cancer in my country rank second and fifth among all malignant tumors, respectively. The survival rate of early cancer is higher than that of late cancer, however, colorectal cancer is usually diagnosed in the late stage, which leads to an increase in mortality. Surgery, radiotherapy and chemotherapy are currently used to treat colorectal cancer, and patients who receive frequent chemotherapy are prone to drug resistance, which may lead to poor treatment outcomes and have harmful side effects on normal cells. Therefore, it is necessary to develop more effective and safer methods to treat colorectal cancer.

[0003] In traditional Chinese medicine, pine and cedar ganoderma is often used to improve health, increase vitality and prolong life. The "Atlas of Chinese Medicinal Fungi" records that pine and cedar ganoderma can strengthen the body and nourish the body. People soak it in wine and take it. It has the effects of nourishing the heart, calming the spirit, nourishing the liver and qi, strengthening the bones and muscles, and nourishing the skin. Polysaccharides are the most recognized edible and medicinal fungus-derived substances with a series of important biological characteristics. Pine and cedar ganoderma polysaccharides are safe and harmless, with few toxic and side effects. More and more researchers are beginning to devote themselves to using ganoderma polysaccharides to develop safe, economical and effective natural medicines. Summary of the invention

[0004] The purpose of the present invention is to provide a pine and cedar ganoderma lucidum polysaccharide with the effect of alleviating colorectal cancer, and a preparation method and application thereof, so as to solve the problems existing in the above-mentioned prior art. The present invention extracts and purifies a new pine and cedar ganoderma lucidum polysaccharide, and the pine and cedar ganoderma lucidum polysaccharide has an outstanding effect in alleviating colorectal cancer. The present invention provides theoretical support for the development of colorectal cancer drugs with effectiveness and biosafety.

[0005] To achieve the above object, the present invention provides the following solutions:

[0006] The present invention provides a pine ganoderma lucidum polysaccharide having the effect of alleviating colorectal cancer. The pine 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;

[0007] The structure of the pine ganoderma polysaccharide is:

[0008]

[0009] Optionally, the method for preparing the pine and ganoderma lucidum polysaccharide comprises the following steps:

[0010] The fruiting bodies of Ganoderma lucidum are dried, ultrafinely crushed, sieved, added with deionized water, ultrasonically extracted, centrifuged to obtain the supernatant, concentrated, protein removed, and the supernatant is concentrated again, precipitated with ethanol aqueous solution, and the precipitate is collected and freeze-dried to obtain the crude polysaccharide of Ganoderma lucidum;

[0011] The crude polysaccharide of Ganoderma lucidum is dissolved, eluted through diethylaminoethyl agarose gel, the polysaccharide solution is collected, concentrated, dialyzed, freeze-dried, and then purified through a chromatographic column to obtain the polysaccharide of Ganoderma lucidum.

[0012] Optionally, the eluent of 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 by a HiPrep 26 / 60 Sephacryl S-400 HR preparative grade chromatographic column, and a second purification by a HiLoad 16 / 600 Superdex 200 pg preparative grade chromatographic column.

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

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

[0015] The present invention also provides the use of the pine and ganoderma lucidum polysaccharide in preparing medicine for treating colorectal cancer.

[0016] The present invention also provides a medicine for treating colorectal cancer, wherein the effective component of the medicine comprises the pine ganoderma lucidum polysaccharide.

[0017] Optionally, the drug further comprises a pharmaceutically acceptable carrier substance and / or adjuvant.

[0018] Optionally, the auxiliary materials include at least one of an absorption promoter, a diluent, a filler, an excipient, a binder, a wetting agent, a disintegrant, a surfactant, an adsorption carrier, a lubricant and a flavoring agent.

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

[0020] The invention extracts and purifies a new pine and spruce ganoderma lucidum polysaccharide, which consists of mannose, glucose, galactose and fucose, wherein the molar ratio of mannose, glucose, galactose and fucose is 3.221:1.414:80.539:14.826, and the molecular weight of the pine and spruce ganoderma lucidum polysaccharide is 7.056 kDa. Cell experiments and animal experiments have confirmed that the pine and 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 colon weight index, bleeding, crypt budding, dysplasia and hyperplasia; it also 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, and at the same time increases the level of IL-4. It can be seen that the pine and ganoderma lucidum polysaccharide provided by the present invention has an outstanding effect in alleviating colorectal cancer. The present invention provides a theoretical support for the further development of colorectal cancer drugs with effectiveness and biosafety. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0022] Figure 1 This is the elution curve of DEAE Sepharose Fast Flow of crude polysaccharide from Ganoderma lucidum;

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

[0024] Figure 3 The elution curves of Ganoderma lucidum water-washed polysaccharide HiPrep 26 / 60Sephacryl S-400HR (A) and HiLoad 16 / 600Superdex200pg (B);

[0025] Figure 4 This is the absolute molecular weight analysis diagram of GTP-a2;

[0026] Figure 5This is a monosaccharide component analysis diagram of the purified polysaccharide from Ganoderma lucidum;

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

[0028] Figure 7 It is the GTP-a2 structure;

[0029] Figure 8 GTP-a2 inhibits the development of colorectal tumors in mice induced by AOM / DSS; A: Colorectal length of each group; B: HE staining; C: Statistical chart of the number of tumors in each group; D: Colon weight index;

[0030] Fig. 9 GTP-a2 regulates AOM / DSS-induced mouse colon metabolites; A: Colon metabolites; B: Principal component analysis; C: Volcano plot;

[0031] Fig.10 Cluster heat map of GTP-a2 regulation of AOM / DSS-induced mouse colon metabolites;

[0032] Fig.11 The differential abundance score diagram of KEGG enriched metabolic pathways of GTP-a2-regulated AOM / DSS-induced mouse colon metabolites;

[0033] Fig.12 For immunohistochemical staining;

[0034] Fig.13 The diagrams are marker protein expressions; A: IFN-γ; B: IL-4; C: TGF-β; D: IL-17A. DETAILED DESCRIPTION

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

[0036] It should be understood that the terms described in the present invention are only for describing a particular embodiment and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper and lower limits of the scope is also specifically disclosed. The intermediate value in any stated value or stated range, and each smaller range between any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0037] Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

[0038] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments of the present invention description without departing from the scope or spirit of the present invention. Other embodiments derived from the present invention description will be apparent to those skilled in the art. The present invention description and examples are exemplary only.

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

[0040] Example 1

[0041] Using Ganoderma lucidum as raw material, a method for preparing and purifying polysaccharides with anti-colorectal cancer properties was provided. The tumor number and intestinal tissue metabolites were used as evaluation indicators to characterize the polysaccharide, and its activity in alleviating colorectal cancer was explored.

[0042] 1. Preparation of Crude Polysaccharide from Ganoderma Lucidum

[0043] After the fruiting bodies of Ganoderma lucidum were dried in an oven at 80°C to constant weight, they were ultrafinely crushed into powder and passed through a 150-mesh sieve. Deionized water was added at a solid-liquid ratio of 1g:70mL, and the extraction was performed at an ultrasonic power of 300W for 1h. After centrifugation, the supernatant was taken and concentrated to 1 / 30 volume by rotary evaporation, and the protein was removed by the Sevag method. The supernatant was concentrated and precipitated with 75% (v / v) ethanol at 4°C overnight. The precipitate was collected and freeze-dried to obtain the crude polysaccharide of Ganoderma lucidum.

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

[0045] (1) The crude polysaccharide of Ganoderma lucidum was dissolved in deionized water, and gradient eluted with purified water, 0.1 M and 0.3 M sodium chloride solutions through DEAE Sepharose Fast Flow at a flow rate of 1 mL / min. The solutions were collected respectively, and the polysaccharide content was determined by phenol-sulfuric acid method. The polysaccharides of different components (GTP-a, GTP-c, among which the yield of GTP-b was too low to be used in subsequent experiments) were collected, concentrated to 1 / 5 volume by rotary evaporation, dialyzed overnight at 4°C, and freeze-dried and collected ( Figure 1 ).

[0046] Colorectal cancer cells (SW480) in the logarithmic growth phase were seeded into 96-well plates (8×10 3 The cells were then cultured with 0.08, 0.16, 0.31, 0.63, 1.25, and 2.50 mg / mL of GTP-a and GTP-c for another 24 hours. The cell survival rate was determined using the MTT method. Both GTP-a and GTP-c showed an inhibitory effect on the growth of SW480 cells, with a 50% inhibitory concentration (IC 50 ) values ​​were 1.0mg / mL and 1.96mg / mL ( Figure 2 ), among which GTP-a significantly inhibited the growth of SW480 cells.

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

[0048] 3. Analysis of structural characteristics of purified polysaccharides from Ganoderma lucidum

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

[0050] Table 1 Molecular characteristics of GTP-a2

[0051]

[0052] (2) Analysis of monosaccharide composition: 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°C for 2 hours. Pass nitrogen and blow dry. Add 3 mL of methanol to wash, blow dry again, and repeat the methanol washing 2-3 times. Add 5 mL of sterile water to dissolve, and transfer to a chromatographic bottle for testing. Take 0.2 mL of monosaccharide standard solution or polysaccharide hydrolyzate in a stoppered conical bottom centrifuge tube, add 0.2 mL of 0.5 mol / L sodium hydroxide solution, 0.5 mL of 0.5 mol / L 1-phenyl-3-methyl-5-pyrazolone (PMP) methanol solution, vortex mix, and react in a 70°C water bath for 1 hour. After the reaction is completed, add 0.2 mL of 0.5 mol / L hydrochloric acid to neutralize the added sodium hydroxide, add 1 mL of chloroform and vortex extract 3 times to remove excess PMP, discard the chloroform layer, take 0.3 mL and add water to make the volume to 1 mL. Thermo U3000 liquid chromatography system was used, the chromatographic column was ZORBAX Eclipse XDB-C18, the mobile phase was acetonitrile: phosphate buffer (12 g / L potassium dihydrogen phosphate, 2M NaOH adjusted to pH = 6.8) isocratic elution, the volume ratio of acetonitrile to phosphate buffer was 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. The results are shown in Figure 5 As shown, GTP-a2 is composed of mannose (Man), glucose (Glc), galactose (Gal), and fucose (Fuc), and the molar ratio (mol%) is 3.221:1.414:80.539:14.826.

[0053] (3) Polysaccharide methylation analysis: Add 1 ml μL DMSO to 2 mg sample to dissolve; add 30 mg NaOH and incubate for 30 min; add 250 μL iodomethane solution, fill with nitrogen, react for 1 h in the dark, add 250 μL iodomethane solution again to react for 1 h; add 1 mL water and 2 mL dichloromethane, vortex to mix, centrifuge, and discard the aqueous phase; aspirate the lower dichloromethane phase and blow dry with nitrogen; add 1 mL 2 M TFA and react at 121 °C for 120 min; blow dry with nitrogen at 30 °C; add 1 mL freshly prepared MNaBD4 (in ammonia water). Incubate at room temperature with magnetic stirring for 2.5 hours; add 300 μL of acetic acid to terminate the reaction and blow dry with nitrogen; dry twice with 2 mL of 5% (vol / vol) acetic acid in methanol at 40°C with nitrogen, and then dry twice with 2 mL of methanol and nitrogen at 40°C; add 1.5 mL of acetic anhydride, vortex mix, and react at 100°C for 2.5 hours; add 2 mL of water and let stand for 10 minutes; add 1 mL of dichloromethane, vortex mix, centrifuge, and discard the aqueous phase; take the lower dichloromethane phase and detect on the machine. The chromatographic system used was an Agilent gas chromatography system (Agilent 7890A; Agilent Technologies, USA), an HP-5MS capillary column (30m×0.25mm×0.25μm, Agilent J&W Scientific, Folsom, CA, USA), a carrier gas of high-purity helium (purity not less than 99.999%), a flow rate of 1.0mL / min, and an injection port temperature of 260°C. The injection volume was 1μL, split injection, the split ratio was 10:1, and the solvent delay was 2.2min. 50°C was maintained for 1.0min, the temperature was increased to 130°C at 50°C / min, and the temperature was increased to 230°C at 3°C / min and maintained for 2min. According to the retention time and standard data of partially methylated alditol acetate (PMAA) in the spectral database of the Complex Carbohydrate Research Center (CCRC), GTP-a2 was mainly composed of 11 glycoside 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 connection mode, the structure of GTP-a2 is speculated to be →6-α-D-Galp-(1→ skeleton, α-L-Fucp-(1→ connected at C2 ( Figure 7 ).

[0054] Table 2 GTP-a2 methylation analysis results

[0055]

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

[0057] (1) Grouping and administration of mice: 30 C57BL / 6 mice (male; 6-8 weeks; 18-20 g) were kept at 23±1°C and 55±5% relative humidity. After a one-week adaptation period, 20 mice were intraperitoneally injected with 10 mg / kg AOM (azomethane), and their drinking water was changed to 2% DSS at weeks 1, 4, and 7. After 4 weeks, 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 (100 mg / 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 every day for 6 weeks.

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

[0059] (3) Histopathological observation: Intestinal tissues were fixed in 4% paraformaldehyde and stored at 4°C. They were routinely paraffin-embedded, sectioned, stained with HE and immunohistochemistry, and observed under an optical microscope for pathological analysis.

[0060] (4) Intestinal samples (n = 5 / group) were randomly selected from the normal group, AOM / DSS group, and AOM / DSS+GTP-2 group. Nontargeted metabolomics analysis was performed using ultra-high performance liquid chromatography (1290 Infinity LC; Agilent Technologies) with time-of-flight and Orbitrap mass spectrometry (AB Triple TOF 6600Q; AB SCIEX, MA, USA). The structures of metabolites in biological samples were determined by matching the retention time, molecular weight (molecular weight error <10 ppm), secondary fragmentation spectrum, collision energy, and other information 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 (p < 0.05) and projected variable importance (VIP) values ​​> 2.0 of metabolites were considered as standard values ​​for differential metabolites.

[0061] (5) Immunohistochemical analysis: After the sections in (3) were sealed, they were incubated with primary antibodies (IL-17A, TGF-β, IL-4, IFN-γ) at 4°C for 16 h. Then, the sections were incubated with goat anti-rabbit secondary antibodies at 4°C for 4 h. Finally, all slides were observed by optical microscopy, and the images obtained were quantified using Image J software.

[0062] (6) Experimental results

[0063] 6.1 GTP-a2 inhibits the development of AOM / DSS-induced colorectal tumors in mice

[0064] After GTP-a2 administration, a significant reduction in the number and size of colon tumors was observed ( Figure 8 In addition, colon weight index and bleeding were alleviated ( Figure 8 Middle A, Figure 8 Middle D). Histological examination revealed complex crypt budding, dysplasia, and hyperplasia in the colon of AOM / DSS-induced mice, and these histopathological changes were abrogated after GTP-a2 treatment ( Figure 8 (middle B).

[0065] 6.2 GTP-a2 regulates metabolite levels in AOM / DSS-induced mouse colon

[0066] A total of 576 metabolites were identified in the colon, including lipids and lipid-like molecules (31.0%), organic acids and derivatives (30.9%), and organic heterocyclic compounds (12.90%) ( Fig. 9 The principal component analysis (PCA) plot showed that GTP-a2 significantly altered metabolite levels in the colon ( Fig. 9 (B). Compared with the normal group, 348 metabolites were changed in the AOM / DSS group and 304 metabolites were changed in the GTP-a2-treated AOM / CSS group. In addition, compared with the AOM / DSS group, mice treated with GTP-a2 showed increased levels of 17 metabolites and decreased levels of 32 metabolites ( Fig. 9 C). Twenty metabolic pathways related to protein digestion and absorption, cancer central carbon metabolism, alanine, aspartate, and glutamate metabolism, and mineral absorption were also altered after GTP-a2 treatment ( Fig.10 ). Based on VIP score >1.5 and p<0.05, the levels of 30 metabolites were significantly different among the three groups ( Fig.11 ).

[0067] 6.3GTP-a2 regulates immune function and alleviates colorectal cancer

[0068] Immunohistochemical analysis showed that the expression of IFN-γ (p<0.01), TGF-β (p<0.01), and IL-17A (p<0.01) in the AOM / DSS-induced colon was enhanced compared with the normal group. GTP-a2 treatment significantly suppressed 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 compared with AOM / DSS-induced mice, 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 ( Fig.12 , Fig.13 ).

[0069] 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 pine ganoderma lucidum polysaccharide having the effect of alleviating colorectal cancer, characterized in that: The pine 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 pine ganoderma polysaccharide is:

2. The pine ganoderma lucidum polysaccharide according to claim 1, characterized in that The preparation method of the pine and ganoderma lucidum polysaccharide comprises the following steps: The fruiting bodies of Ganoderma lucidum are dried, ultrafinely crushed, sieved, added with deionized water, ultrasonically extracted, centrifuged to obtain the supernatant, concentrated, protein removed, and the supernatant is concentrated again, precipitated with ethanol aqueous solution, and the precipitate is collected and freeze-dried to obtain the crude polysaccharide of Ganoderma lucidum; The crude polysaccharide of Ganoderma lucidum is dissolved, eluted through diethylaminoethyl agarose gel, the polysaccharide solution is collected, concentrated, dialyzed, freeze-dried, and then purified through a chromatographic column to obtain the polysaccharide of Ganoderma lucidum.

3. The pine ganoderma lucidum polysaccharide according to claim 2, characterized in that The eluent of the diethylaminoethyl agarose gel is water and sodium chloride solution, and the flow rate is 1 mL / min.

4. The pine ganoderma lucidum polysaccharide according to claim 2, characterized in that The purification includes a first purification using a HiPrep 26 / 60 Sephacryl S-400HR preparative chromatography column, and a second purification using a HiLoad 16 / 600 Superdex 200 pg preparative chromatography column.

5. The pine ganoderma lucidum polysaccharide according to claim 4, characterized in that The eluent of the HiPrep 26 / 60 Sephacryl S-400 HR preparative chromatographic column is a 0.15 mol / L sodium chloride solution; the eluent of the HiLoad 16 / 600 Superdex 200 pg preparative chromatographic column is a 0.15 mol / L sodium chloride solution.

6. The pine ganoderma lucidum polysaccharide according to claim 2, characterized in that The power of the ultrasound is 300W and the time is 1 hour.

7. Use of the Ganoderma lucidum polysaccharide according to any one of claims 1 to 6 in the preparation of a drug for treating colorectal cancer.

8. A drug for treating colorectal cancer, characterized in that: The active ingredient of the medicine comprises the Ganoderma lucidum polysaccharide according to any one of claims 1 to 6.

9. The drug according to claim 8, characterized in that The drug also includes pharmaceutically acceptable excipients.

10. The drug according to claim 9, characterized in that The auxiliary materials include at least one of an absorption promoter, a diluent, a filler, an excipient, a binder, a wetting agent, a disintegrant, a surfactant, an adsorption carrier, a lubricant and a flavoring agent.

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