Application of plant polysaccharide in preparing product for alcohol detoxification and liver protection

Abstract

The present invention discloses the use of a plant polysaccharide in the preparation of a product for hangover relief and liver protection. The present invention proposes for the first time that frangipani polysaccharide can be effectively used for hangover relief and liver protection, including enhancing the activity of ADH and ALDH, key enzymes in the ethanol metabolic pathway, accelerating the rate of ethanol metabolism in the body, and reducing liver damage caused by drunkenness, having a protective effect on the body. At the same time, it can reduce the drunkenness rate, shorten the drunkenness time, significantly reduce the body's ALT, AST, TC, TG and LDL-C levels, increase the HDL-C level, significantly increase the body's liver CAT, SOD, and GSH activities, reduce the liver's MDA level, and significantly improve the body's liver fat accumulation, providing a new direction for the preparation of hangover relief and liver protection products.

Description

Technical Field

[0001] The invention belongs to the field of biotechnology, and particularly relates to application of frangipani flower polysaccharide in preparing products for sobering up and protecting the liver. Background Art

[0002] Alcoholic liver disease (ALD), caused by chronic alcohol intoxication or acute alcohol consumption, has become one of the most common liver diseases worldwide. Acute alcoholic liver injury (AALI) is an early manifestation of ALD and generally refers to damage to the liver caused by excessive drinking over a short period of time. After alcohol enters the body, 90% is metabolized by the liver, primarily through alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH), which convert it into carbon dioxide and water for excretion. However, excessive alcohol intake over a short period of time can lead to various disorders in the body, including hepatocyte apoptosis, gastric mucosal lesions, immune cell damage, increased oxidative stress, and lipid peroxidation.

[0003] Modern treatments for alcohol-induced acute liver injury primarily rely on interventions such as corticosteroids, S-adenosylmethionine, metadoxine, silymarin, antioxidants, and restoration of intestinal function. However, silymarin, a commonly used clinical medication for liver injury, has significant side effects, such as nausea, vomiting, abdominal pain, loss of appetite, and diarrhea. Therefore, exploring bioactive substances with hepatoprotective properties from natural resources to mitigate alcohol-induced liver injury is of great significance. Summary of the Invention

[0004] The present invention aims to solve at least one of the technical problems existing in the above-mentioned prior art. To this end, the present invention proposes the use of frangipani flower polysaccharide in the preparation of a product for sobering up.

[0005] The present invention also proposes the use of frangipani polysaccharide in preparing a product for preventing and / or treating liver damage caused by drinking.

[0006] The present invention also provides a product.

[0007] According to a first aspect of the present invention, the use of frangipani polysaccharide in preparing a hangover relief product is proposed.

[0008] In some embodiments of the present invention, the method for preparing the frangipani polysaccharide comprises the following steps:

[0009] The frangipani flower sample was subjected to defatting, water extraction, alcohol precipitation, protein removal and dialysis in sequence to obtain frangipani flower polysaccharide.

[0010] In some embodiments of the present invention, the defatting is performed using anhydrous ethanol.

[0011] In some embodiments of the present invention, in the defatting step, the solid-to-liquid ratio of the frangipani flower sample to anhydrous ethanol is 1 g: 8-12 mL.

[0012] In some embodiments of the present invention, the solid-liquid ratio is 1 g:10 mL.

[0013] In some embodiments of the present invention, degreasing is performed by heating under reflux.

[0014] In some embodiments of the present invention, the heating reflux is performed at a temperature of 70 to 90° C. for 1 to 2 hours, and the heating reflux step is repeated 2 to 3 times.

[0015] In some embodiments of the present invention, in the water extraction step, the solid-to-liquid ratio of the defatted frangipani flower sample to water is 1 g: 10-20 mL.

[0016] In some embodiments of the present invention, the solid-liquid ratio is 1 g:15 mL.

[0017] In some embodiments of the present invention, water extraction is performed by combining ultrasonic extraction and water bath extraction.

[0018] In some embodiments of the present invention, the ultrasonic extraction is performed under the condition of ultrasonic power of 90-110w for 15-25min.

[0019] In some embodiments of the present invention, the water bath extraction is carried out at 95-105° C. for 2-3 hours.

[0020] In some embodiments of the present invention, the water extraction further includes a step of solid-liquid separation and collecting the separated liquid phase.

[0021] In some embodiments of the present invention, the alcohol precipitation uses ethanol with a mass fraction of 95% or more.

[0022] In some embodiments of the present invention, in the alcohol precipitation step, the volume ratio of the water extract to ethanol is 1:(3-5).

[0023] In some embodiments of the present invention, the alcohol precipitation temperature is 0-4° C., and the alcohol precipitation time is 8-12 h.

[0024] In some embodiments of the present invention, the step of washing the precipitate obtained by alcohol precipitation with anhydrous ethanol, ether and acetone in sequence is further included.

[0025] In some embodiments of the present invention, the protein removal is performed using Sevag reagent.

[0026] In some embodiments of the present invention, the Sevag reagent is a mixed solution of chloroform and n-butanol in a volume ratio of 4:1.

[0027] In some embodiments of the present invention, the volume ratio of the Sevag reagent to the polysaccharide solution obtained by dissolving the precipitate obtained by washing with anhydrous ethanol, ether and acetone in water is 1:4-6.

[0028] In some embodiments of the present invention, the dialysis is performed using a dialysis bag with a molecular weight cut-off of >8000Da.

[0029] In some embodiments of the present invention, the dialysis time is 1 to 3 days.

[0030] In some embodiments of the present invention, the monosaccharides in the frangipani polysaccharide consist of rhamnose, arabinose, galactose, glucose, xylose, mannose and galacturonic acid.

[0031] In some embodiments of the present invention, the molar ratio of rhamnose, arabinose, galactose, glucose, xylose, mannose and galacturonic acid is (14-22): (100-120): (160-185): (200-240): (16-25): (18-27): (400-450).

[0032] In some embodiments of the present invention, the molar ratio of rhamnose, arabinose, galactose, glucose, xylose, mannose and galacturonic acid is (17-21): (105-115): (170-180): (210-230): (18-23): (19-25): (430-440).

[0033] In some embodiments of the present invention, the molar ratio of rhamnose, arabinose, galactose, glucose, xylose, mannose and galacturonic acid is 19:109:174:220:21:22:434.

[0034] In some embodiments of the present invention, the molecular weight of the frangipani polysaccharide is 69445 Da.

[0035] According to a second aspect of the present invention, the use of frangipani polysaccharide in preparing a product for preventing and / or treating liver damage caused by drinking is proposed.

[0036] In some embodiments of the present invention, the alcohol-induced liver damage includes alcohol-induced acute liver damage.

[0037] In some embodiments of the present invention, the product has at least one of the following functions:

[0038] (1) Accelerate the metabolism of ethanol and / or acetaldehyde;

[0039] (2) shorten the duration of intoxication and / or sobriety;

[0040] (3) Reduce the rate of drunkenness;

[0041] (4) Reduce liver index;

[0042] (5) Increase the activity of alcohol dehydrogenase (ADH) and / or acetaldehyde dehydrogenase (ALDH) in the body;

[0043] (6) Reduce serum alanine aminotransferase (ALT) and / or aspartate aminotransferase (AST) levels;

[0044] (7) Increase the level of high-density lipoprotein cholesterol (HDL-C) in serum;

[0045] (8) increasing the activities of catalase (CAT), superoxide dismutase (SOD) and / or glutathione peroxidase (GSH);

[0046] (9) Reduce malondialdehyde (MDA) content;

[0047] (10) Improve ethanol-induced abnormal lipid accumulation;

[0048] (11) Prevent alcohol metabolism disorders caused by excessive alcohol intake.

[0049] In some embodiments of the present invention, increasing the ADH and ALDH enzyme activities in the body includes increasing the ADH and ALDH enzyme activities in the liver.

[0050] In some embodiments of the present invention, the improving ethanol-induced abnormal lipid accumulation includes reducing the levels of triglycerides (TG), cholesterol (TC), and low-density lipoprotein cholesterol (LDL-C) in serum.

[0051] According to a third aspect of the present invention, a product for sobering up or preventing and treating liver damage caused by drinking is provided, wherein the product comprises the above-mentioned frangipani flower polysaccharide.

[0052] In some embodiments of the present invention, the product is at least one of a medicine, a health product, and a functional food.

[0053] In some embodiments of the present invention, the product is a medicine, which further comprises a pharmaceutically acceptable excipient.

[0054] In some embodiments of the present invention, the pharmaceutically acceptable excipients include at least one of a diluent, an excipient, a filler, a binder, a disintegrant, an absorption enhancer, a surfactant, an adsorption carrier, a lubricant, a sweetener, and a flavoring agent.

[0055] In some embodiments of the invention, the excipient comprises water.

[0056] In some embodiments of the present invention, the filler comprises at least one of starch and sucrose.

[0057] In some embodiments of the present invention, the binder comprises at least one of a cellulose derivative, alginate, gelatin, and polyvinyl pyrrolidone.

[0058] In some embodiments of the invention, the humectant comprises glycerin.

[0059] In some embodiments of the present invention, the disintegrant comprises at least one of agar, calcium carbonate and sodium bicarbonate.

[0060] In some embodiments of the present invention, the absorption enhancer comprises a quaternary ammonium compound.

[0061] In some embodiments of the invention, the surfactant comprises cetyl alcohol.

[0062] In some embodiments of the present invention, the adsorption carrier includes at least one of kaolin and bentonite.

[0063] In some embodiments of the present invention, the lubricant comprises at least one of talc, calcium stearate, magnesium stearate and polyethylene glycol.

[0064] In some embodiments of the present invention, the dosage form of the drug is in the form of solid, semi-solid or liquid, and can be an aqueous solution, non-aqueous solution or suspension.

[0065] In some embodiments of the present invention, the dosage form of the drug is tablets, capsules, soft capsules, granules, pills, oral liquids, dry suspensions, pellets, dry extracts, injections or infusions, transdermal agents, and transdermal microneedles.

[0066] In some embodiments of the present invention, the drug may be administered by conventional methods in the art, including but not limited to injection or oral administration.

[0067] In some embodiments of the present invention, the injection administration can be intravenous injection, intramuscular injection, intraperitoneal injection, intradermal injection or subcutaneous injection.

[0068] In some embodiments of the present invention, the mass fraction of the frangipani polysaccharide in the product is 0.01% to 100%.

[0069] According to some preferred embodiments of the present invention, the mass fraction of the frangipani polysaccharide in the product is 0.05% to 95%.

[0070] According to some preferred embodiments of the present invention, the mass fraction of the frangipani polysaccharide in the product is 0.05% to 50%.

[0071] According to some embodiments of the present invention, at least the following beneficial effects are achieved: the present invention proposes for the first time that jasmine flower polysaccharide can be effectively used for hangover and liver protection, including enhancing the activity of key enzymes ADH and ALDH in the ethanol metabolic pathway, accelerating the rate of ethanol metabolism in the body, and reducing liver damage caused by drunkenness, having a protective effect on the body. At the same time, it can reduce the drunkenness rate, shorten the drunkenness time, significantly reduce the body's ALT, AST, TC, TG and LDL-C levels, increase the HDL-C content, significantly increase the body's liver CAT, SOD, GSH activity, reduce the liver's MDA content, and significantly improve the body's liver fat accumulation, providing a new direction for the preparation of alcohol-relieving and liver-protecting products. BRIEF DESCRIPTION OF THE DRAWINGS

[0072] The present invention will be further described below with reference to the accompanying drawings and embodiments, in which:

[0073] Figure 1 This is a glucose standard curve diagram in Example 1 of the present invention;

[0074] Figure 2 This is the ultraviolet spectrum analysis diagram in Example 1 of the present invention;

[0075] Figure 3 This is an infrared spectrum analysis diagram in Example 1 of the present invention;

[0076] Figure 4 This is a high performance gel permeation chromatography analysis chart in Example 1 of the present invention;

[0077] Figure 5 This is a diagram showing the monosaccharide composition of the frangipani polysaccharide in Example 1 of the present invention;

[0078] Figure 6 Figure 2 shows the test results of mouse weight and drunkenness rate in Example 2 of the present invention, wherein A shows the test results of mouse weight, and B shows the test results of drunkenness rate. ###p<0.001 compared with the normal group; *p<0.05, **p<0.01 compared with the model group (n=10);

[0079] Figure 7 Figures 2 show the liver phenotype and liver index test results in Example 2 of the present invention, where A shows the liver morphology and B shows the liver index test results. ####p<0.0001 compared with the normal group; *p<0.05 compared with the model group (n=10);

[0080] Figure 8Graphs showing the results of alcohol detoxification-related enzyme tests in Example 2 of the present invention, wherein A shows the results of ADH enzyme tests, and B shows the results of ALDH enzyme tests. ##p<0.01, ###p<0.001 compared with the normal group; *p<0.05, **p<0.01 compared with the model group (n=10);

[0081] Figure 9 Graphs showing the test results of serum biochemical indicators of mice in Example 2 of the present invention, wherein A is a graph showing the test results of AST, B is a graph showing the test results of ALT, C is a graph showing the test results of TC, D is a graph showing the test results of TG, and E is a graph showing the test results of LDL-C; F is a graph showing the test results of HDL-C, ##p<0.01, ###p<0.001, ####p<0.0001 compared with the normal group; *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001 compared with the model group, (n=10);

[0082] Figure 10 Graphs showing the test results of biochemical indicators in Example 2 of the present invention, wherein A is a graph showing the test results of CAT, B is a graph showing the test results of GSH, C is a graph showing the test results of MDA, and D is a graph showing the test results of SOD. ##p<0.01, ####p<0.0001 are compared with the normal group; *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001 are compared with the model group (n=10). DETAILED DESCRIPTION

[0083] The following will clearly and completely describe the concept and technical effects of the present invention in conjunction with the embodiments to fully understand the purpose, features and effects of the present invention. Obviously, the embodiments described are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, other embodiments obtained by those skilled in the art without creative work are all within the scope of protection of the present invention.

[0084] Example 1 Preparation of Frangipani Polysaccharide

[0085] This example prepared a frangipani polysaccharide, and the specific preparation and verification methods are as follows:

[0086] 1. Extraction of Frangipani Polysaccharide

[0087] (1) crushing: crushing the fully dried frangipani flowers, sieving, and drying to obtain frangipani pollen;

[0088] (2) Degreasing: Jasmine pollen was degreased with 10 times the amount of anhydrous ethanol (i.e., the ratio of Jasmine pollen to anhydrous ethanol was 1 g: 10 mL) at 85°C and refluxed for 90 min. The reflux was repeated once, and the mixture was filtered, and the anhydrous ethanol was evaporated to obtain the medicinal residue, which was then dried.

[0089] (3) Ultrasonic-assisted extraction: The defatted dry powder was mixed with distilled water at a material-liquid ratio of 1 g:15 mL, ultrasonic power 100 W, ultrasonication for 20 min, then placed in a 100 °C water bath for 2.5 h, centrifuged and filtered to obtain the aqueous extract;

[0090] (4) Alcohol precipitation: The filtrate was concentrated to 20% of the original volume using a rotary evaporator. Then, 4 times the amount of anhydrous ethanol was added to the concentrate. The mixture was refrigerated overnight and filtered to collect the precipitate. The ethanol on the surface was evaporated in a fume hood to obtain a precipitate.

[0091] (5) Washing: The precipitate was washed twice with anhydrous ethanol, ether, and acetone in sequence, and dried to obtain a crude polysaccharide extract I;

[0092] (6) Protein removal: Dissolve the above-mentioned crude polysaccharide extract I in an appropriate amount of water, add Sevag reagent according to the ratio of crude polysaccharide to Sevag reagent 5:1, shake thoroughly for 30 minutes, and then centrifuge at 6000 rpm for 15 minutes; carefully aspirate the upper polysaccharide solution, remove the organic phase and precipitate, and then add Sevag solvent equivalent to 1 / 5 of its volume, repeat several times until no obvious precipitation is produced, and store at 4°C to prevent deterioration for subsequent impurity removal;

[0093] (7) Impurity removal: Use a ready-to-use dialysis bag with a diameter of >8000D to remove impurities. First, cut the dialysis bag into a suitable length, soak the preservation solution with deionized water, place the deproteinized frangipani polysaccharide solution into the bag, seal it with a dialysis bag clip, and place a weight at the bottom to make it vertical to the bottom. Then, immerse the dialysis bag in a beaker filled with deionized water and a magnet, and then place the entire beaker on a magnetic stirrer to accelerate the dialysis rate, thereby assembling a complete dialysis device. After dialysis for 48 hours, remove the liquid in the dialysis bag and centrifuge to obtain a precipitate. After freeze-drying the precipitate, obtain the crude frangipani polysaccharide extract II (i.e., purified frangipani polysaccharide).

[0094] 2. Determination of Frangipani Polysaccharide Content

[0095] (1) Glucose standard curve drawing and polysaccharide determination method

[0096] Standard curve preparation: The purified frangipani polysaccharide was determined using the phenol-sulfuric acid method. Accurately weigh 100 mg of glucose standard into a 100 mL volumetric flask and dilute to 100 mL with distilled water to prepare a 0.1 mg / mL glucose standard solution. Transfer the prepared glucose standard solution and dilute to 5 mL with distilled water to prepare concentrations of 0.02 mg / mL, 0.04 mg / mL, 0.06 mg / mL, 0.08 mg / mL, 0.10 mg / mL, 0.12 mg / mL, and 0.14 mg / mL, respectively. Take 200 μL of each dilution and place them in 2 mL test tubes. Add 100 μL of 6% phenol and 500 μL of concentrated sulfuric acid to each test tube, let it stand for 10 minutes, shake it well, and let it stand at room temperature for 20 minutes. Punch 200 μL into each well of a 96-well plate, and punch 3 wells into each test tube. Use a microplate reader to measure the absorbance at a wavelength of 490 nm, using distilled water as a blank control, and obtain a standard curve (such as Figure 1 shown).

[0097] Determination of polysaccharide content: Prepare a 1 mg / mL solution of total polysaccharides from Jasmine flower with distilled water. Dilute 10-fold with distilled water. Pipette 200 μL of the solution into three wells of a 96-well plate. Add phenol and sulfuric acid in the appropriate proportions. Let stand for 10 minutes, shake well, and let stand at room temperature for 20 minutes. Measure the absorbance at 490 nm using a microplate reader. Calculate the polysaccharide content using the following formula: Polysaccharide content (%) = (C × V2 × D) / (W × V1 × 10) × 100% (Formula 2); where C is the sugar content calculated using the standard curve (mg); V2 is the total volume of the extract (mL); V1 is the volume used for measurement (mL); D is the dilution factor; and W is the sample weight (g).

[0098] The glucose standard curve was obtained by phenol-sulfuric acid method. Figure 1 As shown, the regression equation is y = 6.8588x-0.0177; the goodness of fit of the equation is R 2 =0.999. In the range of 0-0.14 mg / mL, there is a good linear relationship between glucose concentration and absorbance.

[0099] The absorbance of total polysaccharides was measured and substituted into the regression equation to determine the polysaccharide concentration. Substituting this into (Equation 2) yielded a total polysaccharide content of 66.03%. Many factors affect polysaccharide concentration and purity, primarily including: higher extraction temperatures can cause polysaccharide decomposition, and more Sevage deproteinization cycles also result in polysaccharide loss.

[0100] 3. Ultraviolet spectrum analysis of frangipani polysaccharide

[0101] The purified frangipani polysaccharide was dissolved in ultrapure water to prepare a polysaccharide solution with a concentration of 1 mg / mL, and the ultraviolet absorption spectrum of the polysaccharide solution was scanned (200-600 nm) using an ultraviolet-visible spectrophotometer.

[0102] The results are as follows Figure 2 As shown in the figure, it can be seen that aromatic amino acids in proteins (such as tryptophan, tyrosine, and phenylalanine) have characteristic absorption peaks near 280nm, and frangipani polysaccharide has a small absorption peak near 280nm, indicating that it contains a small amount of protein; chlorophyll, carotenoids, etc. usually have characteristic absorption peaks near 380nm, and frangipani polysaccharide has no absorption peak at 380nm, indicating that it does not contain these pigments or the pigment content is extremely low.

[0103] 4. Infrared spectroscopy analysis of frangipani polysaccharides

[0104] Take a certain amount of the dried frangipani polysaccharide sample after purification, mix it with dry KBr powder and press it into tablets. -1 Perform infrared spectrum scanning within the range and record the infrared spectrum.

[0105] The results are as follows Figure 3 As shown in the figure, it can be seen that the frangipani polysaccharide is at 3414.351cm -1 The strong absorption peak is caused by the stretching vibration of OH, which is a characteristic peak of sugars; it proves that the substance is sugar. -1 The absorption peak at wave number 1748.156 cm is the characteristic peak of CH stretching vibration; -1 It is the characteristic absorption peak of carboxyl group, indicating that frangipani polysaccharide contains carboxyl group. -1 The observed absorption peaks are associated with the stretching of COC and COH, indicating the possible presence of a pyranose ring.

[0106] 5. Determination of relative molecular weight of frangipani polysaccharide

[0107] The molecular weight of the purified frangipani polysaccharide was determined by high-performance gel permeation chromatography (HPGPC). The chromatographic conditions were as follows: mobile phase: 0.05 M NaCl solution; column: BRT105-103-101 tandem gel column (8 × 300 mm); flow rate: 0.7 mL / min; column temperature: 40°C; injection volume: 25 μL; detector: differential refractive index detector (RID-20A); and analysis time: 70 min.

[0108] The molecular weight spectrum of frangipani polysaccharide is as follows Figure 4As shown in the figure, it can be seen that the frangipani polysaccharide peaks at 34.879 min, its relative peak area is 100%, and its molecular weight is 69445 Da.

[0109] 6. Determination of monosaccharide composition of frangipani polysaccharide

[0110] Monosaccharide composition was determined using ion chromatography. Accurately weigh 5 mg of the purified frangipani polysaccharide sample into an ampoule, add 2 mL of 3M TFA, and hydrolyze at 120°C for 3 h. Accurately transfer the acid hydrolysis solution to a tube, blow dry with nitrogen, add 5 mL of water, and vortex to mix. Aspirate 100 μL of the solution and add 900 μL of deionized water. Centrifuge at 12,000 rpm for 5 min. The supernatant was subjected to IC analysis.

[0111] The results are shown in Table 1 and Figure 5 As shown, it can be seen from the peak time of standard polysaccharides and polysaccharide samples that the main monosaccharide composition and molar percentage of frangipani polysaccharide are rhamnose (Rha): arabinose (Ara): galactose (Gal): glucose (Glc): xylose (Xyl): mannose (Man): galacturonic acid (GalA) = 0.019: 0.109: 0.174: 0.220: 0.021: 0.022: 0.434; among which the monosaccharides with the highest content are galacturonic acid (GalA), glucose (Glc), galactose (Gal) and arabinose (Ara).

[0112] Table 1

[0113]

[0114] Example 2 Application of Frangipani Polysaccharide in the Preparation of Products for Alcohol Relief and Liver Protection

[0115] In this example, an ethanol-induced acute alcoholic liver injury model was established in mice to verify whether frangipani polysaccharide has the effect of sobering up and protecting the liver. The specific verification method is as follows:

[0116] 1. Animals and groups

[0117] Animal model: 70 commercially available SPF-grade C57BL / 6 male mice, aged 6-8 weeks and weighing 20-24 g were used. After one week of adaptive feeding, the mice were randomly divided into seven groups, with 10 mice in each group. The average weight of each group was approximately 23 g. The groups were as follows: (1) normal group (NC); (2) model group (Alcohol); (3) silymarin group (SM, 50 mg / kg·BW); (4) King Drink group (King Drink, 150 mg / kg·BW); (5) low-dose frangipani polysaccharide group (SP-L, 50 mg / kg·BW); (6) medium-dose frangipani polysaccharide group (SP-M, 100 mg / kg·BW); and (7) high-dose frangipani polysaccharide group (SP-H, 300 mg / kg BW).

[0118] 2. Experimental methods

[0119] (1) Construction of a 50% ethanol-induced acute alcoholic liver injury model in C57BL / 6 mice

[0120] The mice were gavaged with 0.1 ml / 10 g of ethanol for 14 consecutive days. Body weights were recorded every two days. The normal and model groups were gavaged with pure water, while the remaining treatment groups were gavaged with the same dosage as above. Two hours after the last dose, the mice were gavaged with 0.14 ml / 10 g of 50% ethanol to induce an acute alcoholic liver injury model, except for the normal group, which was gavaged with pure water.

[0121] After 30 minutes, the drunkenness time and soberness time of the mice were recorded, and the drunkenness rate of the mice after modeling was recorded (drunkenness rate = number of drunk mice (mouse) / total number of mice (mouse) × 100%). The drunkenness of the mice was determined by whether the righting reflex disappeared. After gavage, the mice were placed with their backs down and gently placed in a cage. If the mice maintained the back-down posture for more than 30 seconds, the righting reflex disappeared, indicating drunkenness. If the righting reflex of the drunk mice recovered and they could move freely, they were considered sober.

[0122] After excluding mice that died immediately after gavage and mice that did not show drunkenness for more than 3 hours, the number of drunk mice, drunkenness time and soberness time of each group of mice were observed and recorded.

[0123] After gavage with 50% ethanol, the mice were fasted but not watered for 16 hours before being weighed. The eyeballs were then removed to collect blood, placed in a 1.5 ml ordinary centrifuge tube at room temperature for more than 2 hours, centrifuged at 3000 rpm and 4°C for 10 minutes, and the supernatant was placed in another new centrifuge tube as serum, which was stored at -80°C for later use. After blood collection, the mice were killed, the liver and spleen were removed, washed in physiological saline and the excess blood was absorbed, the liver morphology was photographed, and the liver and spleen were weighed for the calculation of the organ coefficient: a portion of the liver tissue was fixed in a 4% paraformaldehyde solution, and the rest was placed in a cryovial and quickly frozen with liquid nitrogen and stored at -80°C. This is used to calculate the liver index (liver index = liver (mg) / mouse body weight (mg) × 100%).

[0124] (2) Test results

[0125] 1) Mouse weight and drunkenness rate test results after modeling

[0126] The results are as follows Figure 6 As shown, from Figure 6 As shown in Figure A, weight gain in all groups of mice gradually increased over the 14 days following dosing prior to modeling, indicating that frangipani polysaccharide has no significant toxicity to mice. Following gavage with 50% ethanol to establish the model, all groups of mice were fasted for 16 hours. Compared to the NC group, weight loss in all dosing groups decreased to varying degrees, with the alcohol group experiencing the greatest weight loss.

[0127] from Figure 6 As shown in Figure B, the drunkenness rate test results show a significant increase in the model group compared to the normal group, indicating successful modeling. The drunkenness rates of mice treated with frangipani polysaccharide decreased to varying degrees, with the high-dose frangipani polysaccharide group showing the fewest mice experiencing drunkenness, indicating that frangipani polysaccharide supplementation can reduce drunkenness in mice.

[0128] 2) Liver phenotype and liver index

[0129] The results are as follows Figure 7 As shown, from Figure 7 As shown in Figure A, the liver morphology of the normal group mice was rosy and shiny, while the liver of the alcohol group mice was enlarged, pale, and had a grainy surface. The paleness of the liver surface of the mice pretreated with frangipani polysaccharide was alleviated, and the graininess of the liver was reduced.

[0130] from Figure 7 As can be seen from Figure B, compared with the normal group, the liver index of the model group was significantly increased (p<0.0001), while the liver index of the jasmine flower polysaccharide-treated group decreased in a dose-dependent manner.

[0131] 3) Detection of alcohol-detoxifying enzymes.

[0132] Experimental method: Cut 20-50 mg of liver tissue and place it in a 1.5 ml centrifuge tube. Add the homogenization medium extract at a ratio of 1:9 between tissue and normal saline. Use a manual tissue homogenizer to prepare a 10% liver homogenate. Measure the ADH and ALDH enzyme activities in sequence according to the kit instructions.

[0133] The results are as follows Figure 8 As shown in the figure, it can be seen that compared with the normal group, the activities of ADH and ALDH enzymes in the model group were significantly reduced (p<0.01, p<0.0001). After treatment with frangipani polysaccharide, the activities of ADH and ALDH enzymes were significantly restored, and the effect was dose-dependent, indicating that frangipani polysaccharide can increase the activities of ADH and ALDH enzymes and accelerate the metabolism of ethanol in mice.

[0134] 4) Detection of mouse serum biochemical indicators

[0135] Experimental method: Mouse serum was obtained and diluted with normal saline for a certain multiple before direct testing; AST, ALT, TC, TG, LDL-C and HDL-C were determined according to the operating instructions of the Nanjing Jiancheng test kit.

[0136] The results are as follows Figure 9 As shown in the figure, compared with the normal group, the model group mice significantly increased the levels of AST, ALT, TC, TG and LDL-C in the serum and decreased the level of HDL-C; while the mice in the frangipani polysaccharide supplementation group had significantly decreased levels of ALT, AST, TC, TG and LDL-C in the serum, and increased HDL-C. This indicates that frangipani polysaccharide can improve ethanol-induced acute alcoholic liver damage and abnormal lipid accumulation in mice.

[0137] 5) Detection of biochemical indicators in mouse liver

[0138] Experimental method: 20-50 mg of liver tissue was cut and placed in a 1.5 ml centrifuge tube. Normal saline was added as a homogenization medium at a ratio of 1:9 between tissue and normal saline. A 10% liver homogenate was prepared using a manual tissue homogenizer. The liver homogenate was diluted a certain number of times with normal saline to obtain the optimal detection concentration, and then used for the detection of biochemical indicators. The enzyme activities of CAT, SOD, GSH and the content of MDA were measured in sequence according to the operating instructions of the kit.

[0139] The results are as follows Figure 10As shown in the figure, compared with the normal group, the levels of CAT, SOD, and GSH in the liver of the model group were significantly reduced (p<0.01), and the level of MDA was significantly increased (p<0.0001). Treatment with frangipani polysaccharide significantly increased the activity of CAT, SOD, and GSH, and reduced the level of MDA. These results suggest that frangipani polysaccharide can protect the liver from damage by enhancing antioxidant activity.

[0140] Jasmine is a winding, vine-like evergreen ornamental shrub of the genus Jasmine (Oleaceae). Jasmine is bitter in taste, neutral in nature and non-toxic, and enters the liver meridian; it has the traditional efficacy of "soothing the liver and relieving depression, promoting qi circulation and relieving pain." Modern medicinal chemistry studies have found that Jasmine contains iridoids, triterpenes, flavonoids, lignans and other ingredients. Among them, there are monomeric compounds such as oleuropein, hydroxytyrosol, hydroquinone, kaempferol, ethyl gallate, salidroside and quercetin. It is a medicinal and edible resource rich in active ingredients. However, Jasmine polysaccharides and related activities have not been reported so far. The present invention is the first to discover that Jasmine polysaccharides have the effect of sobering up, and can also improve ethanol-induced acute alcoholic liver damage.

[0141] While the embodiments of the present invention have been described in detail above with reference to the accompanying drawings, the present invention is not limited to the embodiments described above. Various modifications may be made within the scope of knowledge possessed by a person skilled in the art without departing from the spirit of the present invention. Furthermore, the embodiments of the present invention and the features thereof may be combined with one another unless there is a conflict.

Claims

1. The application of frangipani flower polysaccharide in the preparation of alcohol-relieving medicines, characterized in that: The preparation method of the frangipani flower polysaccharide comprises the following steps: subjecting the frangipani flower sample to defatting, water extraction, alcohol precipitation, protein removal and dialysis in sequence to obtain the frangipani flower polysaccharide.

2. Use of frangipani polysaccharide in the preparation of a drug for preventing and / or treating liver damage caused by drinking, characterized in that: The preparation method of the frangipani flower polysaccharide comprises the following steps: subjecting the frangipani flower sample to defatting, water extraction, alcohol precipitation, protein removal and dialysis in sequence to obtain the frangipani flower polysaccharide.

3. The use of frangipani polysaccharide in the preparation of a functional food for alleviating liver damage caused by drunkenness, characterized in that: The preparation method of the frangipani flower polysaccharide comprises the following steps: subjecting the frangipani flower sample to defatting, water extraction, alcohol precipitation, protein removal and dialysis in sequence to obtain the frangipani flower polysaccharide.

4. The use according to claim 1 or 2, characterized in that The drug has at least one of the following functions: (1) Accelerate the metabolism of ethanol and / or acetaldehyde; (2) shorten the duration of intoxication and / or sobriety; (3) Reduce the rate of drunkenness; (4) Reduce liver index; (5) Increase the activity of ADH and / or ALDH enzymes in the body; (6) Reduce serum ALT and / or AST levels; (7) Increase serum HDL-C levels; (8) Increase the activity of CAT, SOD and / or GSH; (9) Reduce MDA content; (10) Improving ethanol-induced abnormal lipid accumulation; the improvement of ethanol-induced abnormal lipid accumulation includes reducing the levels of triglycerides and low-density lipoprotein cholesterol in serum; (11) Prevent alcohol metabolism disorders caused by excessive alcohol intake.

5. The use according to any one of claims 1 to 3, characterized in that The degreasing is carried out using anhydrous ethanol.

6. The use according to claim 5, characterized in that In the defatting step, the solid-liquid ratio of the frangipani flower sample to anhydrous ethanol is 1 g: 8-12 mL.

7. The use according to any one of claims 1 to 3, characterized in that The dialysis was performed using a dialysis bag with a molecular weight cut-off of >8000Da.

8. The use according to any one of claims 1 to 3, characterized in that The dialysis time is 1-3 days.

9. The use according to any one of claims 1 to 3, characterized in that The monosaccharides in the frangipani polysaccharide consist of rhamnose, arabinose, galactose, glucose, xylose, mannose and galacturonic acid.

10. The use according to claim 9, characterized in that The molar ratio of rhamnose, arabinose, galactose, glucose, xylose, mannose and galacturonic acid is (14-22): (100-120): (160-185): (200-240): (16-25): (18-27): (400-450).

11. The use according to any one of claims 1 to 3, characterized in that: The molecular weight of the frangipani polysaccharide is 69445 Da.

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