Application of phellinus igniarius extract in preparation of medicine for treating or preventing amyloidosis diseases

Abstract

The invention belongs to the technical field of traditional Chinese medicine extracts, and provides an application of a phellinus igniarius extract in preparation of a medicine for treating or preventing amyloidosis disease. Phellinus igniarius medicinal materials from three different regions are extracted by a decoction method. The obtained extract has a treatment effect on amyloidosis cells, and in an amyloidosis hepatocyte model, the phellinus igniarius extract can significantly improve the cell survival rate. The new application lays a foundation for deep research of phellinus igniarius and development of new drugs.

Description

Technical Field

[0001] The present invention belongs to the technical field of traditional Chinese medicine extracts and relates to use of a Phellinus igniarius extract in preparing a medicine for treating or preventing amyloidosis. Background Art

[0002] Amyloidosis (AL) refers to a class of diseases in which abnormally folded proteins accumulate in tissues or organs, forming amyloid fibrils with a β-pleated structure. This leads to tissue damage and organ dysfunction, potentially affecting multiple systems, including the heart, kidneys, and nervous system. These abnormally folded proteins lose their normal physiological functions, and their accumulation in tissues can cause inflammation and tissue damage, leading to various pathological features and ultimately leading to amyloid-related diseases such as Alzheimer's disease (AD), Parkinson's disease (PD), and type 2 diabetes mellitus (T2DM). Currently, treatment options for amyloidosis are relatively limited, with most treatments still in clinical trials and a lack of widely accepted treatment standards.

[0003] Phellinus igniarius, a traditional medicinal fungus, originates from the fruiting body of the Polyporaceae fungus, Phellinus igniarius. Li Shizhen, in his Compendium of Materia Medica, noted its efficacy in stopping bleeding, promoting blood circulation, softening and detoxifying, and soothing the stomach and relieving diarrhea. Modern research indicates that phellinus igniarius is rich in bioactive components, such as proteins, polysaccharides, flavonoids, triterpenes, sterols, and phenolic acids, which possess significant pharmacological activity. Phellinus igniarius demonstrates broad potential in anti-tumor, anti-inflammatory, and immunomodulatory applications, becoming an important natural resource in modern medicine. The Chinese Materia Medica states: Its habitat: It grows on the trunks of broad-leaved trees such as poplar and willow. Its distribution: It is widely distributed in North and Northwest China, as well as in Heilongjiang, Jilin, Taiwan, Guangdong, Sichuan, Yunnan, and Tibet.

[0004] At present, scholars at home and abroad have conducted a series of studies on mulberry igneous extract. For example, the Chinese patent application with publication number CN120241811A discloses the application of mulberry igneous extract in the preparation of lung cancer H1299 cell inhibitors, and the Chinese patent application with publication number CN113244276A discloses the use of mulberry igneous extract as a new coronavirus treatment drug or antiviral preparation. For example, the Chinese patent application with publication number CN103860602A discloses the application of mulberry igneous extract in the preparation of drugs for treating liver cancer, cervical cancer, and breast cancer.

[0005] Since the efficacy of Phellinus igniarius extract needs to be further explored, the present invention has carried out relevant research. Summary of the Invention

[0006] The present invention provides use of a Phellinus igniarius extract in preparing a medicine for treating or preventing amyloidosis.

[0007] The technical solution of the present invention is achieved as follows: The invention discloses a use of a phellinus igniarius extract in preparing a medicine for treating or preventing amyloidosis, wherein the phellinus igniarius extract is extracted by a decoction method.

[0008] As an embodiment of the present invention, the preparation method of the Phellinus igniarius extract comprises the following steps: Take 100g of Phellinus igniarius, crush it, add water at a material-liquid ratio of 1g:18-25mL, soak for 20-28h, use the decoction method, boil over high heat, then switch to low heat and keep it slightly boiling for 25-35min, filter and collect the filtrate; The residue was decocted twice according to the above steps, and the two filtrates were combined and concentrated by rotary evaporation to 80-120 mL.

[0009] As an embodiment of the present invention, the preparation method of the Phellinus igniarius extract comprises the following steps: Take 100g of Phellinus igniarius, crush it, add water at a material-liquid ratio of 1g:20mL, soak for 20-28h, use the decoction method, boil over high heat, then switch to low heat and keep it slightly boiling for 25-35min, filter and collect the filtrate; The residue was decocted twice according to the above steps, and the two filtrates were combined and concentrated to 100 mL by rotary evaporation.

[0010] As an embodiment of the present invention, the preparation method of the Phellinus igniarius extract comprises the following steps: Take 100g of Phellinus igniarius, crush it, add water at a material-liquid ratio of 1g:20mL, soak for 24h, use the decoction method, boil over high heat until boiling, then switch to low heat and keep it slightly boiling for 30min, filter and collect the filtrate; The residue was decocted twice according to the above steps, and the two filtrates were combined and concentrated to 100 mL by rotary evaporation.

[0011] As an embodiment of the present invention, the rotary evaporation conditions include: temperature 40-60° C., vacuum pressure 10-30 mbar, and rotation speed 100-150 rpm. The temperature is the water bath temperature.

[0012] As an embodiment of the present invention, the rotary evaporation conditions include: temperature 50° C., vacuum pressure 20 mbar, and rotation speed 120 rpm.

[0013] As an embodiment of the present invention, the polysaccharide content of the Phellinus igniarius extract is 1-8%, and the protein content is 9-20%. Preferably, the polysaccharide content of the Phellinus igniarius extract is 1.98-7.5%, and the protein content is 9.8-18.36%. Preferably, the polysaccharide content of the Phellinus igniarius extract is 1.98-3.53%, and the protein content is 11.28-18.36%; further preferably, the polysaccharide content of the Phellinus igniarius extract is 1.98%, and the protein content is 18.36%.

[0014] As an embodiment of the present invention, the use of the above-mentioned Phellinus igniarius extract in the preparation of a medicament for treating or preventing systemic amyloidosis.

[0015] As an embodiment of the present invention, the use of the above-mentioned Phellinus igniarius extract in the preparation of a medicament for treating or preventing hepatic amyloidosis.

[0016] The beneficial effects of the above technical solution of the present invention are: 1. The present invention extracts Phellinus igniarius medicinal materials from three different regions by decoction. It has been verified that the obtained extract has a therapeutic effect on amyloid cells. In the amyloid liver cell model, the Phellinus igniarius extract can significantly improve the cell survival rate.

[0017] 2. The new use provided by the present invention lays a foundation for in-depth research on Phellinus igniarius and development of new drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] The present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments.

[0019] Figure 1 This is a graph showing the cytotoxicity test results of the Phellinus igniarius extract of the present invention, wherein: n refers to the number of independent repeated experiments, in this experiment n=6, *** indicates P<0.001 compared with the blank group; ns indicates no significant difference compared with the blank group, P>0.05.

[0020] Figure 2 This is a diagram showing the results of the dosage screening of Phellinus igniarius extract. In the figure, n refers to the number of independent repeated experiments. In this experiment, n=6. ns: There is no significant difference compared with the model group (P>0.05); *: Compared with the model group (P<0.05); **: Compared with the model group (P<0.01); ***: Compared with the model group (P<0.001);

[0021] Figure 3 The results of the protective effect of phellinus igniarius extracts from different origins on amyloidosis hepatocytes are shown in the figure. In the figure, n refers to the number of independent repeated experiments. In this experiment, n=6. *Compared with the model group, P<0.05; **Compared with the model group, P<0.01; ***Compared with the model group, P<0.001; K represents the blank group, and M represents the model group.

[0022] Figure 4 This is the standard curve of protein content using the BCA method.

[0023] Figure 5 This is the standard spectrum of 17 amino acids in the protein of Phellinus igniarius extract.

[0024] Figure 6 This is the spectrum of 17 amino acids in the protein of Phellinus igniarius extract.

[0025] Figure 7 This is the standard curve for polysaccharide content determination. DETAILED DESCRIPTION

[0026] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the embodiments described are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts are within the scope of protection of the present invention.

[0027] Unless otherwise defined, all technical terms and scientific terms used herein have the same meaning as those generally understood by those skilled in the art of the technical field of the present invention. The terms used herein in the specification of the present invention are only for the purpose of describing specific embodiments and are not intended to limit the present invention. The experimental techniques or test methods involved in the embodiments of the present invention, unless otherwise specified, are conventional methods in the prior art, and their names and / or abbreviations are conventional names in this area, and are very clear and definite in the relevant application fields. Those skilled in the art can understand conventional process steps and apply corresponding equipment according to the names, and implement them according to conventional conditions or conditions recommended by the manufacturer. The various instruments, equipment, raw materials or reagents used in the embodiments of the present invention have no special restrictions on source, are conventional products that can be purchased through regular commercial channels, and can also be prepared according to conventional methods well known to those skilled in the art.

[0028] Preparation Example 1 Three batches of samples of Phellinus igniarius fruiting bodies were collected from Shijiazhuang, Hebei, Nanyang, Henan and Chengde, Hebei.

[0029] The preparation method of the Phellinus igniarius extract comprises the following steps: Take 100g of Phellinus igniarius, crush it, add water at a material-liquid ratio of 1g:20mL, soak for 24h, use the decoction method, boil over high heat until boiling, then switch to low heat and keep it slightly boiling for 30min, filter and collect the filtrate; The filter residue was decocted twice according to the above steps, and the combined filtrates were rotary evaporated (water bath temperature 50°C, vacuum pressure 20 mbar, rotation speed 120 rpm) and concentrated to 100 mL. The resulting Phellinus igniarius extracts were designated SH1 (Shijiazhuang, Hebei), SH2 (Nanyang, Henan), and SH3 (Chengde, Hebei). The two heating methods are commonly used in Traditional Chinese Medicine (TCM) and range from 200-300°C for high heating to 80-100°C for low heating.

[0030] Preparation Example 2 Take 100g of Phellinus igniarius, crush it, add water at a material-liquid ratio of 1g:18mL, soak for 28h, use the decoction method, boil over high heat until boiling, then switch to low heat and keep it slightly boiling for 25min, filter and collect the filtrate; The filter residue was decocted twice according to the above steps, and the two filtrates were combined and rotary evaporated (water bath temperature 55°C, vacuum pressure 20 mbar, speed 100 rpm) and concentrated to 120 mL.

[0031] Preparation Example 3 Take 100g of Phellinus igniarius, crush it, add water at a material-liquid ratio of 1g:25mL, soak for 20h, use the decoction method, boil over high heat until boiling, then switch to low heat and keep it slightly boiling for 35min, filter and collect the filtrate; The filter residue was decocted twice according to the above steps, and the two filtrates were combined and rotary evaporated (water bath temperature 60°C, vacuum pressure 20 mbar, speed 150 rpm) and concentrated to 80 mL.

[0032] Example 1 1. Cytotoxicity test of Phellinus igniarius extract 1.1 Cytotoxicity Cytotoxicity assay ① Cell preparation: Select a cell line (NCTC1469 mouse hepatocyte line, purchased from Shanghai Biotechnology Co., Ltd.) and pre-incubate the cells in a CO2 incubator at 37°C and 5% CO2 for 24 hours until adherent. Add 2 ml of trypsin (0.25% trypsin digestion solution, New Saimei Biotechnology Co., Ltd.) and seed the hepatocytes into 96-well plates (1 × 104 cells / well).

[0033] ②Test substance treatment: The concentration of the mulberry linterus extract prepared in Preparation Example 1 is defined as equivalent to 1 g / mL of the native drug (here it is the concentration converted according to the feed amount, not the actual solute concentration), hereinafter referred to as the mulberry linterus extract concentration.

[0034] 100 μL of cell suspension was inoculated into each well of a 96-well plate, and different concentrations of phellinus linteus extract were added to make the final mixed solution volume the same. The gradient concentrations of phellinus linteus extract SH3 in the final mixed solution were 50 μg / mL, 100 μg / mL, 500 μg / mL, and 1000 μg / mL, respectively. After adding different concentrations of phellinus linteus extract, the cells were incubated in a cell culture incubator at 37 degrees Celsius for 24 hours.

[0035] ③ Microplate reader detection: 10 μL CCK-8 reagent (Biyuntian Biotechnology Research Institute) was added to each well, incubated in the dark for 3 h, and the OD value at a wavelength of 450 nm was detected by a microplate reader.

[0036] 2. Determination of the effect of Phellinus igniarius extract on amyloid cells 2.1 Amyloidosis Hepatocyte Model Model establishment: Normal liver cells from NCTC1469 mice were used to establish the model. DMEM medium (Gibco) supplemented with 3% fetal bovine serum and 1% double-antibody was used for culture in a 37°C, 5% CO2 incubator. Lysine-6 (1×10 -5 mol / L) were used to induce amyloidosis in hepatocytes.

[0037] Preparation of amyloidosis lysozyme LYSO-6: LYSO incubation solution: Weigh an appropriate amount of LYSO powder and add it to a hydrochloric acid solution with a pH of 2. The final concentration is 1×10 -3 M, let it stand at 4℃ overnight to allow it to fully dissolve. After it is completely dissolved, filter it once with a 0.22μm filter membrane.

[0038] Preparation of amyloidosis lysozyme LYSO-6: LYSO incubation solution was placed in a constant temperature shaker with the shaker setting conditions of 65°C and 50 rpm. The incubation was continued for 6 days and the samples were stored at 4°C.

[0039] 2.2 Effects of different therapeutic concentrations Phellinus igniarius extract SH3 dosage screening: set up blank group, model group, low-dose administration group (final concentration 50μg / mL), medium-dose administration group (final concentration 100μg / mL), and high-dose administration group (final concentration 200μg / mL). The specific steps include: inoculating 100 μL per well of a 6-well plate Different concentrations of Phellinus linteus extract were added to the cell suspension to make the final mixed solution volume the same. The gradient concentrations of Phellinus linteus extract SH3 in the final mixed solution were 0 μg / mL (NCTC1469 without any treatment), 0 μg / mL (NCTC1469+LYSO-6 induction), 50 μg / mL (NCTC1469+LYSO-6 induction+50 μg / mL Phellinus linteus extract SH3), 100 μg / mL (NCTC1469+LYSO-6 induction+100 μg / mL Phellinus linteus extract SH3), and 200 μg / mL (NCTC1469+LYSO-6 induction+200 μg / mL Phellinus linteus extract SH3). After adding different concentrations of Phellinus linteus extract, the cells were incubated for 24 hours, and the cell viability was detected by CCK-8 method.

[0040] 2.3 Experimental study on the protection of three batches of phellinus igniarius extracts on amyloidosis hepatocytes (1) Blank group: NCTC1469, i.e., no treatment; (2) Model group: NCTC1469+[LYSO-6 (1×10 -5 mol / L)], that is, LYSO-6 (1×10-5mol / L) was used to induce the establishment of an amyloidosis hepatocyte model; (3) Phellinus igniarius extract SH1 group: NCTC1469 + [LYSO-6 (1×10 -5 mol / L)]+[SH1 Phellinus linteus extract (100 μg / mL)]; (4) Phellinus igniarius extract SH2 group: NCTC1469 + [LYSO-6 (1×10 -5 mol / L)] + [SH2 Phellinus linteus extract (100 μg / mL)]; (5) Phellinus igniarius extract SH3 administration group: NCTC1469 + [LYSO-6 (1×10 -5 mol / L)]+[SH3 Phellinus linteus extract (100 μg / mL)].

[0041] Among them, the Phellinus igniarius extract SH1-SH3 administration groups used LYSO-6 (1×10 -5 After inducing the amyloidosis hepatocyte model with 1% ethanol (100 μmol / L), the phellinus linteus extract was added to make the final mixed solution have the same volume and the concentration of the phellinus linteus extract in the final mixed solution was 100 μg / mL.

[0042] CCK-8 assay for cell viability: After the five groups of cells were incubated in a cell culture incubator at 37°C for 24 hours, the cell viability was assayed using the CCK-8 assay.

[0043] Data analysis: Cell viability (%) = (OD value of experimental group - OD value of blank group) / (OD value of control group - OD value of blank group) × 100%. Graphs were generated using GraphPad Prism, and statistical significance was determined (p < 0.05).

[0044] 3. Measurement results 3.1 Cytotoxicity like Figure 1 The results showed that within the concentration range of 0-500 μg / mL, there was no significant difference in the survival rate of mouse liver cells in the treatment group compared with the blank group, and the extract of Phellinus igniarius had no obvious toxicity to mouse liver cells. When the concentration reached 1000 μg / mL, the cell survival rate in the treatment group was significantly decreased compared with the blank group.

[0045] 3.2 Effects of different therapeutic concentrations like Figure 2 The experimental results of different dosages showed that mulberry ignia extract had a protective effect on amyloidosis hepatocytes. At the same time, as the concentration of mulberry ignia extract increased, the survival of amyloidosis hepatocytes first increased and then decreased. Therefore, the final concentration of mulberry ignia extract was 100 μg / mL as the optimal therapeutic concentration.

[0046] 3.3 Therapeutic effects of phellinus igniarius extracts from different origins on amyloidosis hepatocytes In the amyloidosis hepatocyte model, the SH3 group of Phellinus igniarius extract significantly improved the cell survival rate and the effect was better than that of the SH2 and SH1 groups. Figure 3 The experimental results showed that Phellinus igniarius extract has a protective effect on amyloidosis hepatocytes.

[0047] Example 2 Protein content determination 1. Experimental materials: Phellinus linteus extracts SH1, SH2, and SH3 obtained in Preparation Example 1; 96-well plates and PBS (Wuhan Sewell Biotechnology Co., Ltd.); BCA protein quantification kit (RW0201 Report); 17 amino acid standard mix (Shanghai Anpu); MCI buffer L-8500-PH set (Mitsubishi Chemical); amino acid-specific ninhydrin buffer solution (Japan Photosynthesis).

[0048] 2. Protein content determination 2.1 Standard curve According to the instructions of the BCA protein quantification kit, appropriate amounts of BCA working solution and standard protein solution were prepared, and distilled water was used as a blank control. The protein content standard curve was established by microplate determination.

[0049] 2.2 Protein content determination The concentrations of SH1, SH2, and SH3 extracts were diluted from 1 mg / mL to 200 μg / mL and determined using the microplate method according to the following steps: ① 20 μL of each of the three batches of samples of Phellinus igniarius extract SH1-SH3 and the distilled water blank control were added to a 96-well plate, and duplicate wells were set to ensure data accuracy; ② 200 μL of BCA working solution was injected into each well with a pipette and gently shaken to mix to avoid cross-contamination caused by vigorous operation; ③ Incubate in a 37°C oven for 30 min, detect with a microplate reader at a wavelength of 562 nm, and repeat the determination three times; ④ Substitute the OD value into the standard curve equation to calculate the protein content.

[0050] 2.3 Protein content determination results Prepare standard protein solution and use distilled water as blank control to determine the standard curve using BCA method, such as Figure 4 Formula: y=0.0013×c+0.1101 (y: absorbance, c: concentration μg / mL). The absorbance measured by the Phellinus igniarius extract was substituted into the curve and further converted to obtain the protein content of the Phellinus igniarius extract.

[0051] The results of protein content determination are as follows: The protein contents of the three batches of medicinal materials were determined by BCA protein kit, which were SH1 9.47%, SH2 11.28%, and SH3 18.37%, respectively. The protein content in SH3 was the highest.

[0052] 3. Analysis of amino acid composition in Phellinus igniarius protein 3.1 Chromatographic conditions Chromatographic columns: Standard protein hydrolysis analysis column 4.6 mm*60 mm#2622SC-PH (Hitachi, Japan); derivatization column 4.6 mm*40 mm#2650L (Hitachi, Japan) 3.2 Pre-processing Amino acid hydrolysis: Slowly add 2.5 volumes of 95% ethanol to Phellinus linteus extract SH1 (i.e., a volume ratio of 1:2.5 for Phellinus linteus extract SH1 to 95% ethanol). Stir thoroughly with a glass rod and allow to precipitate at 4°C for 24 hours. Filter the precipitate, discard the supernatant, and freeze-dry the precipitate (pre-freeze at -80°C for 12 hours; then, dry in a freeze dryer at 20 Pa vacuum and -60°C for 48 hours). This yields 8.5 g of lyophilized Phellinus linteus powder. Weigh 0.2 g of lyophilized Phellinus linteus powder, add 15 mL of 6 M hydrochloric acid solution, add 3-4 drops of phenol, flush with nitrogen for 1 minute, and seal the cap. Place the container in a 110°C oven for hydrolysis for 24 hours. Remove the container, cool, filter, and dilute to 50 mL. Take 1 mL of the filtrate, blow dry with nitrogen, reconstitute with 1 mL of sodium citrate solution (pH 2.2), filter through a 0.22 μm filter, and filter into a filtration apparatus.

[0053] 3.3 Instrument conditions Flow rate: Pump 1: 0.40 mL / min, Pump 2: 0.35 mL / min; Injection volume: 20 μL; Column temperature: Separation column: 57°C, Derivatization column: 135°C; Detection wavelengths: 570 nm and 440 nm.

[0054] 3.4 Analysis results of protein and amino acid composition in lyophilized powder of Phellinus igniarius Table 4 Amino acid determination results of Phellinus linteus freeze-dried powder As shown in Table 4, the five amino acids with the highest content are Glu (glutamic acid), Asp (aspartic acid), Arg (arginine), Lys (lysine), Thr (threonine), and Pro (proline). These five amino acids all have a protective effect on the liver. Glutamic acid and aspartic acid work together to eliminate toxins and participate in the urea cycle detoxification; arginine improves liver blood supply and promotes regeneration; lysine regulates fat metabolism and prevents fatty liver; threonine strengthens liver cell membrane structure; and proline strikes a delicate balance between repair and fibrosis. Together, they form a metabolic network that not only provides energy for liver cells, but also protects against oxidative damage and regulates proliferation and apoptosis. Their synergistic effect protects the liver from energy supply, detoxification, and structural maintenance.

[0055] Example 3 Determination of polysaccharide content 1 Experimental materials and instruments The phellinus igniarius extracts SH1, SH2, and SH3 obtained in Preparation Example 1, concentrated sulfuric acid (Hebei Xulong Chemical Co., Ltd.), phenol (Shandong Desenlai Chemical Co., Ltd.), UV-visible spectrometer (T6 Puxi General Instrument Co., Ltd.), and glucose standard (Shanghai Yuanye Biotechnology Co., Ltd.) were used.

[0056] 2. Standard curve establishment Take 10 mg of glucose standard and dilute to 100 mL to obtain a stock solution (0.1 mg / mL). Precisely pipette 0.2, 0.4, 0.6, 0.8, and 1.0 mL of this stock solution into five 10 mL stoppered test tubes and add water to 1 mL. Add 1 mL of freshly prepared 5% phenol and immediately add 5 mL of concentrated sulfuric acid to shake well. Incubate in a boiling water bath for 30 minutes, followed by an ice bath for 5 minutes. Measure the absorbance of each group at 488 nm using UV-Vis spectrophotometry (General Method 0401). Draw a standard curve with glucose concentration (μg / mL) on the horizontal axis and absorbance (Abs) on the vertical axis.

[0057] 3 Polysaccharide content determination method Use a pipette to measure 1 mL each of the SH1, SH2, and SH3 extracts and dilute them to a concentration of 1 mg / mL as test solutions. Transfer 1 mL of each of the three groups of Phellinus igniarius test solutions to a stoppered test tube, add 1 mL of freshly prepared 5% phenol solution, and then add 5 mL of concentrated sulfuric acid to mix thoroughly. Heat in a water bath for 30 minutes and then in an ice bath for 5 minutes. Using the corresponding reagent as a blank control, measure absorbance at a wavelength of 488 nm according to UV-Vis spectrophotometry (General Rule 0401). Measure three times for each group and calculate the mean absorbance. Substitute the mean absorbance value into the standard curve to calculate the polysaccharide content in the Phellinus igniarius extract.

[0058] 4 Methodological validation Using sample SH1 as an example, a spike-in recovery experiment was conducted. Three concentration gradients (low, medium, and high) were designed, with known amounts of glucose standard added (80%, 100%, and 120%). Polysaccharide content was measured using the phenol-sulfuric acid method described above. The recovery (reference value: 80–120%) and RSD (reference value: <5%) were calculated to verify the accuracy of the method.

[0059] 5. Polysaccharide content determination results 5.1 Standard curve The prepared glucose solution was used as the standard, and the absorbance was measured at a wavelength of 488 nm using the phenol-sulfuric acid method. The standard curve was established by fitting the data. Figure 7 shown.

[0060] The standard curve equation was: Abs=0.00874×C+0.02019 (R=0.9980), with a linear range of 20-100 μg / mL. The curve had a good linear relationship.

[0061] 5.2 Content determination The test solution was measured three times at a wavelength of 488 nm using the phenol-sulfuric acid method, and the absorbance was recorded. The polysaccharide content was calculated based on the average absorbance. The results are as follows: Table 5 Polysaccharide content determination results 5.3 Methodological Validation Sample recovery experiments were conducted using sample SH1 as an example. Three concentration gradients (low, medium, and high) were established, and known amounts of glucose standards were added. The recovery and RSD were calculated. The results are shown in Table 6. This method has been verified to be accurate and reliable, meeting pharmacopoeial requirements.

[0062] Table 6 SH1 sample recovery experiment Comprehensive analysis of Examples 1-3 shows that: At the same dosage concentration, SH3 mulberry ignia extract with the highest protein content and low polysaccharide content has the strongest hepatoprotective effect, while SH1 mulberry ignia extract with the lowest protein content and high polysaccharide content has the weakest hepatoprotective effect. This shows that the high protein content in mulberry ignia extract has a good therapeutic effect on amyloidosis hepatocytes.

[0063] The above are only preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A use of a Phellinus linteus extract in the preparation of a medicament for treating or preventing amyloidosis, characterized in that: The Phellinus igniarius extract is extracted by a decoction method.

2. Use of a Phellinus linteus extract according to claim 1 in preparing a medicament for treating or preventing amyloidosis, characterized in that: The preparation method of the Phellinus linteus extract comprises the following steps: Take 100g of Phellinus igniarius, crush it, add water at a material-liquid ratio of 1g:18-25mL, soak for 20-28h, use the decoction method, boil over high heat, then switch to low heat and keep it slightly boiling for 25-35min, filter and collect the filtrate; The residue was decocted twice according to the above steps, and the two filtrates were combined and concentrated by rotary evaporation to 80-120 mL.

3. Use of a Phellinus linteus extract according to claim 1 in preparing a medicament for treating or preventing amyloidosis, characterized in that: The preparation method of the Phellinus linteus extract comprises the following steps: Take 100g of Phellinus igniarius, crush it, add water at a material-liquid ratio of 1g:20mL, soak for 20-28h, use the decoction method, boil over high heat, then switch to low heat and keep it slightly boiling for 25-35min, filter and collect the filtrate; The residue was decocted twice according to the above steps, and the two filtrates were combined and concentrated to 100 mL by rotary evaporation.

4. Use of a Phellinus linteus extract according to claim 1 in preparing a medicament for treating or preventing amyloidosis, characterized in that: The preparation method of the Phellinus linteus extract comprises the following steps: Take 100g of Phellinus igniarius, crush it, add water at a material-liquid ratio of 1g:20mL, soak for 24h, use the decoction method, boil over high heat until boiling, then switch to low heat and keep it slightly boiling for 30min, filter and collect the filtrate; The residue was decocted twice according to the above steps, and the two filtrates were combined and concentrated to 100 mL by rotary evaporation.

5. Use of the Phellinus linteus extract according to claim 1 in preparing a medicament for treating or preventing amyloidosis, characterized in that: The rotary evaporation conditions include: temperature 40-60° C., vacuum pressure 10-30 mbar, and rotation speed 100-150 rpm.

6. Use of the Phellinus linteus extract according to claim 1 in preparing a medicament for treating or preventing amyloidosis, characterized in that: The rotary evaporation conditions include: temperature 50° C., vacuum pressure 20 mbar, and rotation speed 120 rpm.

7. The use according to any one of claims 1 to 6, characterized in that The amyloidosis disease is a systemic amyloidosis disease.

8. The use according to any one of claims 1 to 6, characterized in that The amyloidosis disease is hepatic amyloidosis disease.

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