Application of polygonatum odoratum extract or polygonatum odoratum polysaccharide in prevention and treatment of drug-induced liver injury

Abstract

The invention provides application of polygonatum odoratum extract or polygonatum odoratum polysaccharide in prevention and treatment of drug-induced liver injury. Research finds that when the radix polygonati officinalis extract or the radix polygonati officinalis polysaccharide is used for treating an acetaminophen acute liver injury model mouse, the liver function of the model mouse can be effectively improved (the content of ALT, AST and LDH is reduced), the oxidation resistance of liver tissue is improved (the activity of SOD and the content of GSH are improved), the oxidative stress reaction is slowed down (the content of MDA is reduced), and the pathological injury of the liver tissue is improved. Therefore, the polygonatum odoratum extract and the polygonatum odoratum polysaccharide have a good treatment effect on the drug-induced liver injury caused by acetaminophen, can be used for comprehensively conditioning, are mild and safe, and can be used as novel drugs for preventing and treating the drug-induced liver injury and related diseases caused by the drug-induced liver injury.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to the application of Polygonatum odoratum extract or Polygonatum odoratum polysaccharide in the prevention and treatment of drug-induced liver injury. Background Technology

[0002] The liver is the largest organ in the human body and plays a vital role in regulating physiological functions. Drug abuse, alcoholism, and long-term exposure to toxic environments can all damage the liver. Drug-induced liver injury (DILI) refers to abnormal liver function or hypersensitivity of the liver to drugs and their metabolites caused by drugs or their metabolites. DILI is one of the most common and serious adverse drug reactions, and in severe cases, it can lead to cirrhosis, liver cancer, acute liver failure, and even death. Its incidence is increasing year by year, and it is often the result of multiple mechanisms acting sequentially or in combination. To date, there is still a lack of simple, objective, and specific diagnostic indicators and effective treatments.

[0003] Acetaminophen (APAP) is a widely used over-the-counter analgesic and antipyretic drug that is safe and effective at therapeutic doses. However, overdose of APAP can lead to acute liver injury, the severity of which is positively correlated with the dose and can progress to acute liver failure.

[0004] With the rapid development of medical technology, numerous drugs for treating liver injury have been researched and reported, mainly including traditional Chinese medicine, biological drugs, chemical drugs, and trace elements. However, some studies have shown that some anti-liver injury drugs can inevitably produce biotoxicity. For example, N-acetylcysteine, a commonly used drug in clinical practice for treating drug-induced liver injury, can cause systemic reactions, resulting in side effects such as asthma, bleeding, nausea, vomiting, diarrhea, or constipation. Therefore, the prevention and treatment of liver injury remains a serious problem.

[0005] Furthermore, the treatment plans for liver injury caused by different factors are not the same. For example, although drug-induced liver injury and immune-mediated liver injury are both essentially liver injuries, drug-induced liver injury is caused by liver cell death or liver oxidative stress due to the liver's metabolism of drugs, and is mainly related to the liver's drug metabolism function and the function of P450 drug-metabolizing enzymes. In contrast, immune-mediated liver injury is more often caused by abnormal autoimmune function attacking liver cells or the bile duct system, and is related to congenital factors or exposure to immunogenic substances.

[0006] Therefore, there is a need in this field to discover drugs that can safely and effectively prevent and treat drug-induced liver injury. Summary of the Invention

[0007] Based on this, the purpose of the present invention is to provide the application of Polygonatum odoratum extract or Polygonatum odoratum polysaccharide in the prevention and treatment of drug-induced liver injury.

[0008] To achieve the above objectives, the present invention adopts the following technical solution.

[0009] In a first aspect, the invention provides the use of Polygonatum odoratum extract or Polygonatum odoratum polysaccharide as the sole active ingredient in the preparation of a medicament for preventing and treating drug-induced liver injury.

[0010] In a second aspect, the invention provides the use of Polygonatum odoratum extract or Polygonatum odoratum polysaccharide as the sole active ingredient in the preparation of a medicament for the prevention and treatment of diseases caused by drug-induced liver injury.

[0011] In some embodiments, the diseases caused by drug-induced liver injury include acute or chronic liver injury, acute or chronic drug-induced hepatitis, acute or chronic cholestatic liver disease, acute or chronic liver failure, liver fibrosis, liver cancer, and cirrhosis.

[0012] In some embodiments, the drug-induced liver injury is drug-induced liver injury caused by acetaminophen.

[0013] In some embodiments, the application includes reducing the elevation of ALT, AST, and / or LDH levels caused by drug-induced liver injury.

[0014] In some embodiments, the application includes enhancing the reduction of SOD activity caused by drug-induced liver injury, enhancing the reduction of GSH content caused by drug-induced liver injury, and / or enhancing the increase of MDA content caused by drug-induced liver injury.

[0015] In some embodiments, the application includes improving pathological damage to liver tissue caused by drug-induced liver injury.

[0016] In some embodiments, the Solomon's Seal extract is an aqueous extract of the Solomon's Seal tuber.

[0017] In some embodiments, the preparation method of the Solomon's Seal Extract includes the following steps: (1) taking Solomon's Seal tuber and crushing it into coarse powder; (2) taking the coarse powder and adding it to the extraction solvent, refluxing and extracting at least once, and combining the filtrates; (3) concentrating, purifying and drying the filtrate to obtain the Solomon's Seal Extract.

[0018] In some embodiments, the ratio of the coarse powder to the extraction solvent is 1:(10~20)g / mL.

[0019] In some embodiments, the extraction solvent is a mixture of water and ethanol.

[0020] In some embodiments, the volume ratio of water to ethanol in the mixed solution is 7:(2~4).

[0021] In some embodiments, the reflux extraction is performed at a temperature of 75°C to 85°C, and the number of extractions is 1 to 4.

[0022] In some embodiments, the dosage form of the drug includes tablets, capsules, drops, aerosols, pills, powders, solutions, suspensions, emulsions, granules, liposomes, transdermal preparations, lozenges, suppositories, and lyophilized powder injections.

[0023] Compared with the prior art, the present invention has the following beneficial effects.

[0024] This invention has found that treating mice with an acute liver injury model caused by acetaminophen using Polygonatum odoratum extract or Polygonatum odoratum polysaccharide can effectively improve liver function (reduce ALT, AST, and LDH levels), enhance the antioxidant capacity of liver tissue (increase SOD activity and GSH content), alleviate oxidative stress (reduce MDA content), and improve pathological damage to liver tissue. Therefore, Polygonatum odoratum extract and Polygonatum odoratum polysaccharide have a good therapeutic effect on drug-induced liver injury caused by acetaminophen, and can provide comprehensive conditioning with a mild and safe approach. They can be considered as new drugs for the prevention and treatment of drug-induced liver injury and related diseases. Attached Figure Description

[0025] Figure 1 The study investigated the changes in serum alanine aminotransferase (ALT) in male mice from different groups treated with Polygonatum odoratum extract.

[0026] Figure 2 The changes in serum aspartate aminotransferase (AST) in male mice from different groups of Polygonatum odoratum extract were investigated.

[0027] Figure 3 The changes in serum lactate dehydrogenase (LDH) in male mice from different groups of Solomon's seal extract.

[0028] Figure 4 The changes in liver appearance in male mice from different groups of Polygonatum odoratum extract.

[0029] Figure 5 H&E sections of male mouse livers from different groups of Polygonatum odoratum extract.

[0030] Figure 6 The changes in SOD activity in male mice from different groups of Polygonatum odoratum extract.

[0031] Figure 7 The changes in GSH content in male mice from different groups of Polygonatum odoratum extract.

[0032] Figure 8 The variation of MDA content in male mice from different groups of Polygonatum odoratum extract.

[0033] Figure 9 The changes in serum alanine aminotransferase (ALT) in male mice from different groups of Polygonatum odoratum polysaccharide were investigated.

[0034] Figure 10 The changes in serum aspartate aminotransferase (AST) in male mice from different groups of Polygonatum odoratum polysaccharide were investigated.

[0035] Figure 11 The changes in serum lactate dehydrogenase (LDH) in male mice from different groups of Polygonatum odoratum polysaccharide were investigated.

[0036] Figure 12 The changes in liver appearance in male mice from different groups of Polygonatum polysaccharide.

[0037] Figure 13 H&E sections of male mouse livers from different groups of Polygonatum odoratum polysaccharide.

[0038] Figure 14 The changes in SOD activity in male mice from different groups of Polygonatum odoratum polysaccharide were investigated.

[0039] Figure 15 The changes in GSH content in male mice from different groups of Polygonatum odoratum polysaccharide were investigated.

[0040] Figure 16 The variation of MDA content in male mice from different groups of Polygonatum odoratum polysaccharide.

[0041] Figure 17 The changes in serum alanine aminotransferase (ALT) in female mice from different groups of Solomon's seal extract.

[0042] Figure 18 The changes in serum aspartate aminotransferase (AST) in female mice from different groups of Polygonatum odoratum extract.

[0043] Figure 19 The changes in serum lactate dehydrogenase 2 (LDH2) in female mice from different groups of Solomon's seal extract.

[0044] Figure 20 This is the negative ion mode mass spectrum of Polygonatum odoratum extract.

[0045] Figure 21 This is the positive ion mode mass spectrum of Polygonatum odoratum extract. Detailed Implementation

[0046] Unless otherwise specified, the experimental methods described in the following embodiments of the present invention are generally performed under conventional conditions or as recommended by the manufacturer. All commonly used chemical reagents used in the embodiments are commercially available products.

[0047] Unless otherwise defined, all technical and scientific terms used in this invention have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. The terminology used in this specification is for the purpose of describing particular embodiments only and is not intended to limit the invention.

[0048] The terms "comprising" and "having," and any variations thereof, are intended to cover non-exclusive inclusion. For example, a process, method, apparatus, product, or device that includes a series of steps is not limited to the steps or modules listed, but may optionally include steps not listed, or may optionally include other steps inherent to such process, method, product, or device.

[0049] The term "and / or" as used in this invention describes the relationship between related objects, indicating that there can be three relationships. For example, A and / or B can represent three situations: A exists alone, A and B exist simultaneously, and B exists alone.

[0050] In this invention, the terms "alanine aminotransferase (ALT)" and "aspartate aminotransferase (AST)" refer to enzymes whose blood levels increase when the liver is damaged, and are used as indicators of liver function by utilizing this property.

[0051] In this invention, the term "lactate dehydrogenase (LDH)" refers to a key enzyme in glycolysis, which is widely distributed throughout the body and can indirectly reflect the degree of cell damage.

[0052] In this invention, the terms "reduced glutathione (GSH), malondialdehyde (MDA), and superoxide dismutase (SOD)" are important indicators of oxidative stress.

[0053] The term "prevention and treatment" refers to the prevention and / or treatment of diseases or symptoms described in this invention, which can be achieved in subjects to treat, prevent, reduce and / or alleviate the disease or symptom described in this invention.

[0054] The term "disease and / or symptom" refers to a physical condition of the subject that is related to the disease and / or symptom described in this invention.

[0055] One embodiment of the present invention relates to the use of Polygonatum odoratum extract or Polygonatum odoratum polysaccharide as the sole active ingredient in the preparation of a medicament for preventing and treating drug-induced liver injury.

[0056] One embodiment of the present invention relates to the use of Polygonatum odoratum extract or Polygonatum odoratum polysaccharide as the sole active ingredient in the preparation of a medicament for the prevention and treatment of diseases caused by drug-induced liver injury.

[0057] This invention has found that treating mice with an acute liver injury model caused by acetaminophen using Polygonatum odoratum extract or Polygonatum odoratum polysaccharide can effectively improve liver function (reduce ALT, AST, and LDH levels), enhance the antioxidant capacity of liver tissue (increase SOD activity and GSH content), alleviate oxidative stress (reduce MDA content), and improve pathological damage to liver tissue. Therefore, Polygonatum odoratum extract and Polygonatum odoratum polysaccharide have a good therapeutic effect on drug-induced liver injury caused by acetaminophen, and can provide comprehensive conditioning with a mild and safe approach. They can be considered as new drugs for the prevention and treatment of drug-induced liver injury and related diseases.

[0058] In some embodiments, the medicament comprises an active ingredient and a pharmaceutically acceptable carrier or excipient; the active ingredient is the Polygonatum odoratum extract and / or Polygonatum odoratum polysaccharide.

[0059] In some embodiments, the Solomon's Seal extract is an extract from the Solomon's Seal tuber.

[0060] In some embodiments, the preparation method of the Solomon's Seal Extract includes the following steps: (1) taking Solomon's Seal tuber and crushing it into coarse powder; (2) taking the coarse powder and adding it to the extraction solvent, refluxing and extracting at least once, and combining the filtrates; (3) concentrating, purifying and drying the filtrate to obtain the Solomon's Seal Extract.

[0061] In some embodiments, the ratio of the coarse powder to the extraction solvent is 1:(10~20)g / mL.

[0062] In some embodiments, the ratio of the coarse powder to the extraction solvent is 1:(13~17)g / mL.

[0063] In some embodiments, the extraction solvent is a mixture of water and ethanol.

[0064] In some embodiments, the volume ratio of water to ethanol in the mixed solution is 7:(2~4).

[0065] In some embodiments, the reflux extraction is performed at a temperature of 75°C to 85°C, and the number of extractions is 1 to 4.

[0066] In some embodiments, the reflux extraction is performed at a temperature of 78°C to 82°C, and the number of extractions is 2 to 3.

[0067] In some embodiments, the filtrate is concentrated to a density of 1.10 to 1.15.

[0068] In some implementations, the concentration method can be any form within the art, such as vacuum concentration.

[0069] In some embodiments, the purification method is macroporous resin purification.

[0070] In some embodiments, the drying method is spray drying.

[0071] In some implementations, the reflux extraction may be performed once, twice, three times, four times, or more.

[0072] In some embodiments, the water can be any form of water in the art, such as purified water, deionized water, recycled water, etc.

[0073] In some embodiments, the pharmaceutically acceptable carrier or excipient includes, but is not limited to, lactose, glucose, sucrose, sorbitol, mannitol, starch, gum arabic, calcium phosphate, alginate, gel, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, hydroxybenzene, methyl formate, propylparaben, talc, magnesium stearate, and mineral oil, but is not limited thereto. In addition to the above-mentioned components, the medicament of the present invention may also contain lubricants, wetting agents, sweeteners, flavoring agents, emulsifiers, suspending agents, and preservatives.

[0074] The medicaments described in this invention can be used in patients or other animals receiving the medicaments of this invention to treat, prevent, alleviate, and / or relieve the diseases or symptoms described in this invention. In some preferred embodiments, the medicaments are used in mammals, including but not limited to humans, cattle, horses, sheep, pigs, goats, rabbits, cats, dogs, rats, and any other mammals with livers that may be damaged.

[0075] In one embodiment, the drug is used on a human.

[0076] The dosage of the drug of the present invention depends on many factors, such as the nature and severity of the disease to be prevented or treated, the sex, age, weight and individual response of the patient or animal, the route of administration and the frequency of administration. The dosage can be administered in a single dose form or in several dose forms, such as two, three or four dose forms. The dosage level must be selected based on the specific route of administration, the severity of the condition being treated, and the patient's condition and medical history.

[0077] However, it should be recognized that the total daily dosage of the medicine of the present invention must be determined by the attending physician within the scope of reliable medical judgment. For any specific patient, the specific therapeutically effective dosage level must be determined based on a variety of factors, including the disorder being treated and its severity; the specific composition used; the patient's age, weight, general health condition, sex, and diet; the timing, route of administration, and excretion rate; the duration of treatment; any other medicines used in combination or concurrently; and similar factors known in the medical field. For example, it is practiced in the art to start with a dose below the level required to achieve the desired therapeutic effect and gradually increase the dose until the desired effect is achieved.

[0078] In some embodiments, the drug is administered in a single dose, i.e., a single-dose form. In a further preferred embodiment, the person's weight is assumed to be 60 kg, and the single dose is equivalent to 5 g to 200 g of the drug, for example, 5 g, 8 g, 10 g, 15 g, 20 g, 30 g, 40 g, 50 g, 60 g, 70 g, 80 g, 90 g, 100 g, 110 g, 120 g, 130 g, 140 g, 150 g, 160 g, 170 g, 180 g, or 190 g, and more preferably 15 g to 100 g of Polygonatum odoratum. Those skilled in the art should understand that, generally, a person's weight is assumed to be 60 kg; if the person's weight is not 60 kg, this standard can be used for conversion.

[0079] The drug of this invention can be administered in unit dose form via enteral or non-enteric routes, such as oral, intramuscular, subcutaneous, nasal, oral mucosa, skin, peritoneum, or rectum. Dosage forms include tablets, capsules, drops, aerosols, pills, powders, solutions, suspensions, emulsions, granules, liposomes, transdermal preparations, lozenges, suppositories, lyophilized powder injections, etc. It can be a conventional formulation, a sustained-release formulation, a controlled-release formulation, and various microparticle delivery systems.

[0080] The following description is based on specific implementation methods.

[0081] Experimental materials Animals: SPF-grade male C57BL / 6 mice, weighing 20 (±2) g, 6 weeks old, provided by the Animal Experiment Center of Guilin Medical University; APAP reagent with a purity (AR) of 99% was purchased from MCE (MedChemExpress), lot number 114541; Liver function test kits (AST, ALT, LDH) were purchased from Shenzhen Mindray Bio-Medical Electronics Co., Ltd. The oxidative stress reagent kit MDA, and the GSH and SOD reagent kits were purchased from Nanjing Jiancheng Company.

[0082] The Polygonatum odoratum polysaccharide was provided by Fufeng Ciyuan Biotechnology Co., Ltd., with a purity of over 95%.

[0083] Example 1: Preparation of Polygonatum odoratum extract Morphological identification and HPLC fingerprint comparison ensured that the raw material was the dried rhizome of Polygonatum odoratum, a plant of the Liliaceae family. After testing for moisture (≤12%), ash (≤5%), and exogenous contaminants (heavy metals, pesticide residues, etc.) according to the standards of the Chinese Pharmacopoeia, the raw material was screened through a 40-mesh sieve and then pulverized at low temperature (≤40℃) to ensure that the raw material met the standards of the Chinese Pharmacopoeia (moisture ≤12%, ash ≤5%).

[0084] Dynamic countercurrent extraction: The extraction solvent is a mixed solution of water and ethanol, with a water:ethanol ratio of 70:30 (volume ratio); extraction temperature: 80℃; solid-liquid ratio: 1:15 g / mL; extraction is performed 3 times; the filtrates are combined and concentrated under reduced pressure to a density of 1.10~1.15; after purification with D101 macroporous resin (elution with 30% ethanol), spray drying is performed (air inlet at 180℃, air outlet at 85℃) to obtain powder, which is the Polygonatum odoratum extract.

[0085] Example 2: Therapeutic effect of acetaminophen-induced acute liver injury in mice This embodiment studies the therapeutic effects of Polygonatum odoratum extract and Polygonatum odoratum polysaccharide prepared in Example 1 on acetaminophen-induced acute liver injury in mice. Acetaminophen (APAP), also known as paracetamol, is a widely used antipyretic analgesic. Its chemical name is N-(4-hydroxyphenyl)acetamide, and its CAS number is 103-90-2. APAP is commonly used as an over-the-counter drug to treat symptoms such as colds, fever, joint pain, neuralgia, and migraines.

[0086] I. Experimental Methods 1. Grouping and administration (1) Treatment with Polygonatum odoratum extract: 42 male C57BL / 6J mice weighing between 18 and 22g at 6 weeks of age were randomly divided into 6 groups: control group, model group (APAP), high-dose Polygonatum odoratum extract group (3g / kgPO), medium-dose Polygonatum odoratum extract group (2g / kgPO), low-dose Polygonatum odoratum extract group (1g / kgPO), and N-acetylcysteine ​​control group (NAC, 100mg / kg), with 7 mice in each group. The Polygonatum odoratum extract group was administered the corresponding dose of the drug by gavage (high dose was 3g / kg; medium dose was 2g / kg; low dose was 1g / kg), the N-acetylcysteine ​​control group was administered the corresponding dose of the drug by gavage, and the control group and model group were administered distilled water by gavage once a day for 3 consecutive days.

[0087] (2) Treatment with Polygonatum odoratum polysaccharide: 25 male C57BL / 6J mice weighing between 18 and 22g at 6 weeks of age were randomly divided into 5 groups: control group, model group (APAP), high-dose Polygonatum odoratum polysaccharide group (0.8g / kg POPL), medium-dose Polygonatum odoratum polysaccharide group (0.4g / kg POPL), and low-dose Polygonatum odoratum polysaccharide group (0.2g / kg POPL), with 5 mice in each group. The Polygonatum odoratum polysaccharide group was administered the corresponding dose (0.8g / kg for high dose, 0.4g / kg for medium dose, and 0.2g / kg for low dose) of the drug by gavage, while the normal and model groups were administered distilled water by gavage. The treatment was administered once a day for 3 consecutive days.

[0088] (3) Treatment with Polygonatum odoratum extract: Fifteen female C57BL / 6 mice were randomly divided into five groups of three each: blank control group (NC), model group (MD), high-dose Polygonatum odoratum extract group (3 g / kg PO (PH)), medium-dose Polygonatum odoratum extract group (2 g / kg PO (PM)), and low-dose Polygonatum odoratum extract group (1 g / kg PO (PL)). Each treatment group was administered the corresponding dose of the test sample by gavage daily, while the blank control group and model group were administered an equal volume of physiological saline by gavage for 7 consecutive days.

[0089] 2. Establishment of an APAP acute liver injury model In the above (1) and (2), fasting for 16 hours began on the evening of the second day of the dosing cycle (3 days in total). After fasting, the last dose of Polygonatum odoratum was administered. One hour after the last dose, the blank group was injected with physiological saline intraperitoneally, while the other groups were injected with 300 mg / kg APAP (dissolved in physiological saline) intraperitoneally to induce drug-induced acute liver injury in mice.

[0090] The above (3) began fasting for 16 hours on the evening of the sixth day of the administration cycle (7 days in total). After fasting, the last dose of Polygonatum odoratum was administered. One hour after the last dose, the blank group was injected with physiological saline intraperitoneally, while the other groups were injected with 300 mg / kg APAP (dissolved in physiological saline) intraperitoneally to induce drug-induced acute liver injury in mice.

[0091] 3. Sample collection Mice in each group were injected intraperitoneally with APAP or physiological saline for 24 hours. Body weight was measured, and blood was collected from the eyeballs. After standing at room temperature for 1 hour, the blood was centrifuged at 3000 rpm for 15 minutes at 4°C. The supernatant, or serum, was collected and stored at -80°C. Mice were euthanized by cervical dislocation, and the liver was harvested. The liver was washed with physiological saline, and the surface liquid was blotted off with filter paper before body weight was measured. The largest lobe of the liver tissue was divided into three parts, fixed in 4% paraformaldehyde solution for subsequent pathological analysis, and the remaining liver lobes were chopped, flash-frozen in liquid nitrogen, and then stored at -80°C.

[0092] 4. Determination of biochemical indicators Alanine aminotransferase (ALT), purchased from Shenzhen Mindray Bio-Medical Electronics Co., Ltd., batch number 140124009; aspartate aminotransferase (AST), purchased from Shenzhen Mindray Bio-Medical Electronics Co., Ltd., batch number 140224004; lactate dehydrogenase (LDH), purchased from Shenzhen Mindray Bio-Medical Electronics Co., Ltd., batch number 142723011, was detected using a fully automated biochemical analyzer.

[0093] 5. Processing of pathological sections Liver tissues from the largest right lobe of different mice (with the largest cross-section, including the portal vein area and central vein area) were fixed in 4% paraformaldehyde, dehydrated, cleared, embedded, and stored at room temperature. When needed, sections (4µm) were prepared. Before staining, the sections were baked in an oven at 70℃ for 2 hours, followed by dewaxing with xylene, hematoxylin-eosin (H&E) staining, dehydration with gradients of different concentrations of ethanol, clearing with xylene, mounting with neutral resin, and observation under an optical microscope.

[0094] 6. Detection of oxidative stress indicators Accurately weigh 1 g of liver tissue, add 9 times the volume of physiological saline, and prepare a 10% liver tissue homogenate under ice bath conditions. Centrifuge at 2500 r / min for 15 min at 4℃, and collect the supernatant. Determine SOD activity and GSH and MDA content strictly according to the instructions of the assay kit.

[0095] 7. Statistical processing methods Data were summarized using Excel spreadsheets, and data analysis and graphing were performed using GraphPad 8.0.2 statistical software. Paired t-tests were used for pairwise comparisons. A p-value < 0.05 was considered statistically significant.

[0096] II. Experimental Results 1. The therapeutic effect of Polygonatum odoratum extract administered for 3 days (1) Changes in serum ALT, AST, and LDH activities in male mice in each group like Figure 1 As shown, serum ALT activity in the Model was significantly higher than that in the Control group, with a statistically significant difference (P<0.001), indicating successful model establishment. Compared with the Model, 3 g / kg PO and 1 g / kg PO significantly reduced ALT activity (P<0.001), indicating that Polygonatum odoratum extract can effectively prevent and treat APAP-induced acute liver injury.

[0097] and Figure 1 The relevant data is shown in Table 1.

[0098] Table 1. Changes in serum ALT activity (U / L) in mice of different groups like Figure 2 As shown, AST activity in the serum of the Model group was significantly higher than that in the Control group (P<0.001), indicating successful modeling. Compared with the model group, 3 g / kg PO and 1 g / kg PO significantly reduced AST activity (P<0.001), indicating that Polygonatum odoratum extract can effectively prevent and treat APAP-induced acute liver injury.

[0099] and Figure 2 The relevant data is shown in Table 2.

[0100] Table 2. Changes in serum AST activity (U / L) in mice of different groups like Figure 3 As shown, LDH activity in the Model serum was significantly higher than that in the Control group, with a statistically significant difference (P<0.001), indicating successful model establishment. Compared with the model group, 3 g / kg PO significantly reduced LDH activity (P<0.001), indicating that the active ingredients of Polygonatum odoratum can effectively prevent and treat APAP-induced acute liver injury.

[0101] and Figure 3 The relevant data is shown in Table 3.

[0102] Table 3. Changes in serum LDH activity (U / L) in mice of different groups (2) Changes in liver morphology of male mice in each group treated with Polygonatum odoratum extract like Figure 4 As shown, compared to the control group, the Model showed significant liver enlargement, a light red and uneven color, and visible punctate hemorrhages and necrosis on the surface. The dorsal membrane exhibited numerous millet-like white spots, resembling acute liver injury. This indicated the rapid onset of drug-induced acute liver injury. 3 g / kg PO significantly improved this condition; the liver color approached that of the Control group, the swelling decreased, and no obvious hemorrhages or necrosis were observed. The results indicate that Polygonatum odoratum extract can protect the appearance of the mouse liver.

[0103] (3) Changes in pathological sections of male mice in each group of Polygonatum odoratum extract groups like Figure 5As shown, the control hepatocytes are arranged radially around the central vein, the hepatic cords are clearly visible, and there is no dilation of the hepatic sinusoids. No obvious hepatocyte necrosis or degeneration was observed, and there was no inflammatory cell infiltration or fibrosis in the hepatic lobules or portal area. Compared with the control group, the model showed significantly enlarged and congested hepatic sinusoids, with extensive inflammatory cell infiltration and increased eosinophilicity in the cytoplasm of multiple hepatocytes. Bridging necrosis was observed in the hepatic lobules, mainly concentrated around the central vein. Congestion bands were visible within some areas of the necrotic region. Chromatin condensation, apoptotic bodies and vacuoles with cytoplasmic condensation, and large bilirubin-like granules were observed at the edges of the necrotic areas. The central vein disrupted the endothelium of the cell nucleus, and nuclear dissolution and pyknosis were observed. 3 g / kg PO significantly reversed these changes. The hepatic cords were neatly arranged, and there was no congestion or enlargement within the hepatic sinusoids, only a small amount of necrotic areas, inflammatory cell infiltration, and eosinophilic changes. Although there were scattered lesions in the 1 g / kg PO group, the lesions were more characterized by ballooning hepatocytes and edema, with necrotic areas, but significantly fewer than in the model. These results indicate that Polygonatum odoratum extract can improve the pathological damage of liver tissue in mice with APAP-induced acute liver injury.

[0104] (4) Changes in oxidative stress-related indicators in male mice in each group treated with Polygonatum odoratum extract Depend on Figure 6 , Figure 7 and Figure 8 It was found that, compared with the control group, the model group mice showed significantly decreased SOD activity and GSH content in the liver (P<0.01) (P<0.05), and significantly increased MDA (P<0.001). The high-dose PO group restored SOD activity and significantly increased GSH content while decreasing MDA content (P<0.001) (P<0.05) (P<0.001). This indicates that Polygonatum odoratum extract can enhance the antioxidant capacity of liver tissue and alleviate oxidative stress.

[0105] and Figures 6-8 The relevant data is shown in Table 4.

[0106] Table 4. Changes in oxidative stress-related indicators in male mice of each group. The results above show that Polygonatum odoratum extract can significantly and rapidly reduce transaminase and lactate dehydrogenase, improve liver function indicators, protect liver appearance and tissue morphology, and provide multi-faceted conditioning. It is gentle and safe and can be used to prevent and / or treat liver damage, thereby preventing further development of liver damage and related diseases.

[0107] 2. The therapeutic effect of Polygonatum odoratum polysaccharide administration for 3 days (1) Changes in serum ALT, AST, and LDH activities in male mice in different groups treated with Polygonatum odoratum polysaccharide like Figure 9As shown, serum ALT activity in the Model was significantly higher than that in the Control group (P<0.001), indicating successful model establishment. However, compared to the Model, 0.8 g / kg POPL, 0.4 g / kg POPL, and 0.2 g / kg POPL significantly reduced ALT activity (P<0.05), indicating that Polygonatum odoratum polysaccharide can effectively prevent and treat APAP-induced acute liver injury.

[0108] and Figure 9 The relevant data is shown in Table 5.

[0109] Table 5. Changes in serum ALT activity (U / L) in mice from different groups treated with Polygonatum odoratum polysaccharide. like Figure 10 As shown, serum AST activity in the Model was significantly higher than that in the Control group (P<0.001), indicating successful model establishment. However, compared to the Model, 0.8 g / kg POPL, 0.4 g / kg POPL, and 0.2 g / kg POPL significantly reduced AST activity (P<0.01), suggesting that Polygonatum odoratum polysaccharide can effectively prevent and treat APAP-induced acute liver injury.

[0110] and Figure 10 The relevant data is shown in Table 6.

[0111] Table 6. Changes in serum AST activity (U / L) in mice from different groups treated with Polygonatum odoratum polysaccharide. like Figure 11 As shown, LDH activity in the serum of the Model was significantly higher than that in the Control group, with a statistically significant difference (P<0.001), indicating successful model establishment. Compared with the Model, 0.8 g / kg POPL, 0.4 g / kg POPL, and 0.2 g / kg POPL significantly reduced LDH activity (P<0.05), indicating that Polygonatum odoratum polysaccharide can effectively prevent and treat APAP-induced acute liver injury.

[0112] and Figure 11 The relevant data is shown in Table 7.

[0113] Table 7. Changes in serum LDH activity (U / L) in mice from different groups treated with Polygonatum odoratum polysaccharide. (2) Changes in the morphology of the livers of male mice in each group treated with Polygonatum odoratum polysaccharide like Figure 12As shown, compared to the control group, the Model also exhibited significant liver enlargement, with a light red and uneven color, and visible punctate hemorrhages and necrosis on the surface. The dorsal membrane showed numerous millet-like white spots, resembling acute liver injury. This indicated that drug-induced acute liver injury was rapidly progressive. 0.2 g / kg POPL significantly improved this condition; the liver color approached that of the normal group, the swelling was reduced, and no obvious hemorrhages or necrosis were observed. The results indicate that Polygonatum odoratum polysaccharide can protect the appearance of the mouse liver.

[0114] (3) Pathological changes in male mice in each group of Polygonatum polysaccharide groups like Figure 13 As shown, in the Control group, hepatocytes were arranged radially around the central vein, with clear hepatic cord structures and no dilation of hepatic sinusoids. No obvious hepatocyte necrosis or degeneration was observed, and there was no inflammatory cell infiltration or fibrosis in the hepatic lobules or portal area. Compared with the Control group, the Model group showed significantly enlarged hepatic sinusoids with congestion, extensive inflammatory cell infiltration, and increased eosinophilicity in the cytoplasm of multiple hepatocytes. Bridging necrosis was observed in the hepatic lobules, mainly concentrated around the central vein. Congestion bands were visible in some areas within the necrotic region. Chromatin condensation, apoptotic bodies and vacuoles with cytoplasmic condensation, and large bilirubin-like granules were observed at the edges of the necrotic areas. The central vein disrupted the endothelium of the cell nucleus, and nuclear dissolution and pyknosis were observed. Although scattered lesions were observed in the 0.4 g / kg POPL and 0.2 g / kg POPL groups, the lesions were more characterized by hepatocyte ballooning degeneration and edema, with visible necrotic areas, but significantly fewer than in the Model group. The results indicate that Polygonatum odoratum polysaccharide can improve the pathological damage of liver tissue in mice with APAP-induced acute liver injury.

[0115] (4) Changes in oxidative stress-related indicators in male mice in each group treated with Polygonatum odoratum polysaccharide Depend on Figure 14 , Figure 15 and Figure 16 It was found that, compared with the control group, the model group mice showed significantly decreased SOD activity and GSH content in the liver (P<0.01), and significantly increased MDA (P<0.01). 0.2 g / kg POPL restored SOD activity and significantly increased GSH content while decreasing MDA content (P<0.01). This indicates that Polygonatum odoratum polysaccharide can enhance the antioxidant capacity of liver tissue and alleviate oxidative stress.

[0116] and Figures 14-16 The relevant data is shown in Table 8.

[0117] Table 8. Changes in oxidative stress-related indicators in male mice of each group. The results above show that Polygonatum odoratum polysaccharide can significantly and rapidly reduce transaminase and lactate dehydrogenase, improve liver function indicators, protect liver appearance and tissue morphology, and has a multi-faceted conditioning effect. It is mild and safe and can be used to prevent and / or treat liver damage, thereby preventing further development of liver damage and related diseases.

[0118] 3. The therapeutic effect of Polygonatum odoratum extract administered for 7 days (1) Changes in serum biochemical indicators Depend on Figure 17 , Figure 18 and Figure 19 The results showed that, compared with the blank control group, the serum ALT, AST, and LDH activities of female mice in the model group were significantly increased, indicating that the APAP-induced drug-induced liver injury model in female mice was successfully established. Compared with the model group, the ALT, AST, and LDH activities of the high-dose group (3 g / kg) and medium-dose group (1 g / kg) of Polygonatum odoratum extract were significantly decreased, indicating that Polygonatum odoratum extract can effectively improve liver function after drug-induced liver injury in female mice.

[0119] and Figures 17-19 The relevant data are shown in Tables 9, 10 and 11.

[0120] Table 9. Changes in serum ALT activity (U / L) in female mice treated with Polygonatum odoratum extract. Table 10. Changes in serum AST activity (U / L) of female mice treated with Polygonatum odoratum extract in each group. Table 11 Changes in serum LDH2 activity (U / L) of female mice treated with Polygonatum odoratum extract in each group Example 3 Mass spectrometry analysis was performed on the components of the Polygonatum odoratum extract prepared in Example 1. The mass spectrometry conditions were as follows: electrospray ionization source (ESI source), simultaneous monitoring mode for positive and negative ions (ESI+, ESI-), scanning mode was full scan / data-dependent secondary mass spectrometry (Full MS / ddMS2), and the main source parameters were set as follows: sheath gas flow rate was 9 arb, spray voltage was 3 kV, capillary (ion transport tube) temperature was 320℃, S-lens voltage was 55 kV, and auxiliary gas heating temperature was 30℃. The primary mass spectrometry mode was Full MS, with a scan range (m / z) of 100–1200, a primary resolution of 70,000, a C-Trap ion count of 1e6, and a maximum ion implantation time of 100 ms. The secondary scan mode was ddMS2, with a secondary resolution of 35,000, a vertex excitation time of 3–9 s, and step collision energies of 20 kV, 40 kV, and 60 kV, and a dynamic exclusion time of 8 s.

[0121] Figure 20 This is the negative ion mode mass spectrum of the Polygonatum odoratum extract. Figure 21 This is the positive ion mode mass spectrum of the Polygonatum odoratum extract.

[0122] Mass spectrometry analysis, using SwissADME, identified 96 components that met five "yes" criteria in the Druglikeness category, had a Bioavailability Score > 0.3, and a high GI absorption. These components were considered the main active ingredients of Polygonatum odoratum extract. Table 12 below shows the top 14 most studied active ingredients.

[0123] Table 12 The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0124] The embodiments described above are merely illustrative of several implementations of the present invention, and while the descriptions are specific and detailed, they should not be construed as limiting the scope of the present invention. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these modifications and improvements all fall within the scope of protection of the present invention. Therefore, the scope of protection of this patent should be determined by the appended claims.

Claims

1. Application of Polygonatum odoratum extract or Polygonatum odoratum polysaccharide as the sole active ingredient in the preparation of drugs for the prevention and treatment of drug-induced liver injury.

2. Application of Polygonatum odoratum extract or Polygonatum odoratum polysaccharide as the sole active ingredient in the preparation of drugs for the prevention and treatment of diseases caused by drug-induced liver injury.

3. The application as described in claim 2, characterized in that, The diseases caused by drug-induced liver injury include acute or chronic liver injury, acute or chronic drug-induced hepatitis, acute or chronic cholestatic liver disease, acute or chronic liver failure, liver fibrosis, liver cancer, and cirrhosis.

4. The application as described in any one of claims 1 to 3, characterized in that, The drug-induced liver injury mentioned refers to drug-induced liver injury caused by acetaminophen.

5. The application as described in any one of claims 1 to 3, characterized in that, The applications include reducing elevated ALT, AST, and / or LDH levels caused by drug-induced liver injury.

6. The application as described in any one of claims 1 to 3, characterized in that, The applications include reducing SOD activity caused by drug-induced liver injury, reducing GSH levels caused by drug-induced liver injury, and / or reducing MDA levels caused by drug-induced liver injury.

7. The application as described in any one of claims 1 to 3, characterized in that, The applications include improving pathological damage to liver tissue caused by drug-induced liver injury.

8. The application as described in any one of claims 1 to 3, characterized in that, The Solomon's Seal Extract is an extract from the tuber of Solomon's Seal. Preferably, the preparation method of the Solomon's Seal Extract includes the following steps: (1) taking Solomon's Seal tuber and crushing it into coarse powder; (2) taking the coarse powder and adding it to the extraction solvent, refluxing and extracting at least once, and combining the filtrates; (3) concentrating, purifying and drying the filtrate to obtain the Solomon's Seal Extract.

9. The application as described in claim 8, characterized in that, The ratio of the coarse powder to the extraction solvent is 1:(10~20) g / mL; and / or, The extraction solvent is a mixture of water and ethanol; preferably, the volume ratio of water to ethanol in the mixture is 7:(2~4); and / or, The reflux extraction temperature is 75℃~85℃, and the number of extractions is 1~4.

10. The application as described in any one of claims 1 to 3, characterized in that, The dosage forms of the drugs include tablets, capsules, drops, aerosols, pills, powders, solutions, suspensions, emulsions, granules, liposomes, transdermal preparations, lozenges, suppositories, and lyophilized powder injections.

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