Pharmaceutical composition for improving acute alcoholic gastric mucosal lesion and application

Abstract

The invention relates to the technical field of medicines, in particular to a pharmaceutical composition for improving acute alcoholic gastric mucosal lesion and application. The invention discloses the new medical application of the lucid ganoderma, ginseng and kudzuvine root formula for the first time, and experiments and clinical verifications prove that the formula can increase the activity of key enzymes in stomach and liver tissues of a mouse with acute alcoholic gastric mucosal injury, reduce the content of a proinflammatory factor TNF-alpha in the stomach tissues, increase the generation of an anti-inflammatory factor IL-10 in the stomach tissues, relieve the inflammatory injury of ethanol to gastric mucosa, improve the curative effect of alcohol on the gastric mucosa and improve the curative effect of alcohol on the gastric mucosa. The levels of protective factors EGF, SS and PGE-2 in serum are increased, the defense mechanism of gastric mucosa is enhanced, and the repairing capacity of the gastric mucosa is improved. The discovery has great influence on the research and treatment of acute alcoholic gastric mucosal injury diseases, and has clinical practical value.

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical technology, and particularly relates to a pharmaceutical composition for improving acute alcoholic gastric mucosal injury and its application. Background Art

[0002] Acute alcoholic gastric mucosal injury refers to the condition of gastric mucosal damage caused by a large amount of alcohol intake in a short period of time. In severe cases, it can lead to digestive tract perforation and is one of the important causes of gastric bleeding. Epidemiological investigation results show that the incidence of alcoholic gastric mucosal injury is increasing year by year. Drinking alcohol is a relatively common behavior habit in people's lives. However, excessive alcoholism or a large amount of alcohol intake in a short period of time are the key factors causing gastric mucosal damage, seriously affecting people's healthy lives. Currently, the main drugs for treating acute alcoholic gastric mucosal injury clinically include drugs that reduce gastric acid secretion such as H2 receptor antagonists and proton pump inhibitors, gastric mucosal protective drugs such as sucralfate and misoprostol, and gastric mucosal repair agents such as teprenone. Although they can relieve the symptoms in the short term, they are expensive and have large side effects. Therefore, the multi-target and multi-pathway treatment advantages of traditional Chinese medicine are an effective strategy for intervening in acute alcoholic gastric mucosal injury.

[0003] The prior art has also disclosed various pharmaceutical compositions with anti-gastric mucosal injury effects. For example, the Chinese patent with the authorized publication number CN103736054B discloses a pharmaceutical composition for protecting gastric mucosa and its preparation method, which is composed of ginsenoside Rb1, Codonopsis pilosula, Atractylodes lancea, Magnolia officinalis, Fructus Aurantii, Amomum villosum, Amomum kravanh, Drynaria fortunei, roasted licorice root, and Poria cocos. The Chinese patent with the authorized publication number CN107744577B discloses a pharmaceutical composition for treating gastric mucosal injury, which is composed of Periplaneta americana extract, Citrus reticulata Blanco, Acorus tatarinowii Schott, Rheum palmatum L., Poria cocos, Polygonatum odoratum (Mill.) Druce, Atractylodes macrocephala Koidz, and Amomum villosum. However, the formula of these compositions contains many herbs and complex ingredients, making it difficult to control the quality of the products.

[0004] Lingzhi-Shenge formula is a compound preparation with Ganoderma lucidum, Panax ginseng, and Pueraria lobata as the core components. These are all important resources that can be used both as medicine and food, and are commonly found in health care products or traditional Chinese medicine formulas with effects such as liver protection, immune regulation, and metabolism improvement. However, there is no report on using this formula to improve acute alcoholic gastric mucosal injury. Summary of the Invention

[0005] Aiming at the deficiencies of the prior art, the present invention has conducted in-depth research on the pharmacological effects of the Lingzhi-Shenge formula and found that this formula has a significant effect on acute alcoholic gastric mucosal injury, and has fewer herbs, which is convenient for controlling the quality of the product.

[0006] One of the purposes of the present invention is to provide a pharmaceutical composition, which is composed of the following raw materials: 3-10 parts by weight of Ganoderma lucidum, 6-15 parts by weight of Pueraria lobata, 6-15 parts by weight of Panax ginseng, 12-20 parts by weight of Poria cocos, and 110-440 mg / kg of oligomeric walnut peptide.

[0007] Among them, Ganoderma lucidum is sweet in taste and neutral in nature. It belongs to the liver, heart, spleen, lung, and kidney meridians, enters the five internal organs, tonifies the liver qi, and is the first herb for nourishing the liver. It mainly nourishes the liver qi to soothe the liver and resolve alcohol. Pueraria lobata is sweet, pungent, and cool in nature. It belongs to the lung and stomach meridians. It dispels pathogenic factors from the exterior to relieve alcoholism, guides various herbs to supplement qi and promote the ascending of clear qi, and generates fluid to quench thirst to relieve symptoms such as dry mouth after drinking. The two herbs together serve as the monarch herbs;

[0008] Ginseng is sweet and slightly bitter in taste and slightly warm in nature. It belongs to the spleen and lung meridians. It tonifies the five internal organs, especially strengthens the qi in the middle jiao, enhances its function of dispelling dampness, and does not cause heat generation due to supplementation. Poria cocos is sweet and light in taste and neutral in nature. It belongs to the heart, lung, spleen, and kidney meridians. It strengthens the spleen and promotes diuresis to eliminate dampness, guides the alcohol pathogen to be dissipated from the lower jiao, restricts excessive supplementation, enables supplementation to have elimination, and supplementation without stagnation. The combination of the two herbs strengthens the spleen and dispels dampness, helps the qi movement in the middle jiao to be smooth, and disperses the property of alcohol, and together they serve as the ministerial herbs;

[0009] Walnut peptide is extracted from walnut kernels, can warm and tonify the liver and kidney, nourish blood and moisten the intestine. It is sweet in taste and belongs to the lung, kidney, and large intestine meridians. It can tonify the liver and kidney, nourish and protect the liver, moisten the intestine to enable the alcohol pathogen to be dissipated from the lower jiao, eliminate pathogenic factors without harming healthy qi, assist Poria cocos to enable the alcohol toxin to be eliminated through the urine and feces, and also harmonize various herbs, and serves as the assistant herb.

[0010] In some specific embodiments, in the pharmaceutical composition of the present invention, the amount of Ganoderma lucidum can be 3 - 10 parts by weight, or 3 - 10 parts by weight, or 5 - 10 parts by weight, or 3 - 6 parts by weight, or 6 - 9 parts by weight, or 6 parts by weight; and,

[0011] the amount of Pueraria lobata can be 6 - 15 parts by weight, or 6 - 12 parts by weight of Pueraria lobata, or 6 - 9 parts by weight of Pueraria lobata, or 9 - 15 parts by weight of Pueraria lobata, or 9 - 12 parts by weight of Pueraria lobata, or 9 parts by weight; and / or

[0012] the amount of ginseng can be 6 - 15 parts by weight, or 6 - 12 parts by weight of ginseng, or 6 - 9 parts by weight of ginseng, or 9 - 15 parts by weight of ginseng, or 9 - 12 parts by weight of ginseng, or 9 parts by weight; and / or

[0013] the amount of Poria cocos can be 12 - 20 parts by weight, or 15 - 20 parts by weight of Poria cocos, or 15 - 18 parts by weight of Poria cocos, or 12 - 18 parts by weight of Poria cocos, or 12 - 15 parts by weight of Poria cocos, or 15 parts by weight; and / or

[0014] the amount of oligomeric walnut peptide can be 110 - 440 mg / kg, or 110 - 380 mg / kg of oligomeric walnut peptide, or 110 - 350 mg / kg of oligomeric walnut peptide, or 150 - 300 mg / kg of oligomeric walnut peptide, or 200 - 250 mg / kg of oligomeric walnut peptide, or 220 mg / kg of oligomeric walnut peptide.

[0015] In some specific embodiments, the Ganoderma lucidum is one or a combination of Ganoderma lucidum powder and Ganoderma lucidum extract; and / or,

[0016] The kudzu root is one or a combination of kudzu root powder and kudzu root extract; and / or,

[0017] The ginseng is one or a combination of ginseng powder and ginseng extract; and / or,

[0018] The poria cocos is one or a combination of poria cocos powder and poria cocos extract.

[0019] The second object of the present invention is to provide the use of the above drug composition in the preparation of a drug for preventing, alleviating or improving acute alcoholic gastric mucosal injury.

[0020] Furthermore, the following uses are also within the protection scope of the present invention:

[0021] The drug composition as an active ingredient can be used in combination in the preparation of the drug, or can be compounded with other components having anti-inflammatory activity; the composition of the active components is: 3-10 parts by weight of ganoderma lucidum, 6-15 parts by weight of kudzu root, 6-15 parts by weight of ginseng, 12-20 parts by weight of poria cocos, 110-440 mg / kg of oligomeric walnut peptide

[0022] The drug does not contain other effective anti-inflammatory active ingredients except the drug composition; the composition of the active components is: 3-10 parts by weight of ganoderma lucidum, 6-15 parts by weight of kudzu root, 6-15 parts by weight of ginseng, 12-20 parts by weight of poria cocos, 110-440 mg / kg of oligomeric walnut peptide

[0023] The third object of the present invention is to provide a pharmaceutical preparation, which is prepared from the above drug composition with pharmaceutically acceptable excipients into a pharmaceutically acceptable dosage form.

[0024] Furthermore, the dosage form of the pharmaceutical preparation is an oral dosage form or an injection dosage form, and a controlled release or sustained release dosage form well known in the modern pharmaceutical industry can also be adopted. Preferably, the drug is a dosage form for gastrointestinal administration.

[0025] The dosage form for gastrointestinal administration refers to a dosage form in which a pharmaceutical preparation enters the gastrointestinal tract after oral administration and exerts a local or systemic effect through absorption, such as powder, tablet, granule, capsule, solution, emulsion, and suspension, etc.

[0026] Furthermore, the amount of the active ingredient is 0.001-90 wt% relative to the total weight of the drug; or, 0.02-50 wt%; or 0.1-20%; or 0.005-5%.

[0027] In some specific embodiments, the amount of the active ingredient can be selected as 0.01%, 0.05%, 0.1%, 0.5%, 2%, 5%, 10%, 30%, 40%, 50%, 80% relative to the total weight of the drug;

[0028] It is well understood by those skilled in the art that the pharmaceutically acceptable carriers are generally recognized for this purpose and serve as inactive ingredients of the medicament.

[0029] The adjuvants include solvents, propellants, solubilizers, cosolvents, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, glidants, flavoring agents, preservatives, suspending agents, coating materials, fragrances, antiadhesives, chelating agents, penetration enhancers, pH regulators, buffers, plasticizers, surfactants, foaming agents, defoaming agents, thickeners, clathrates, humectants, absorbents, diluents, flocculants and deflocculants, filter aids, and release retardants.

[0030] Among them, the diluent can be one or more of mannitol, sucrose, lactose, sorbitol, xylitol, polyethylene glycol, propylene glycol, vegetable oil, and mineral oil; the disintegrant can be one or more of croscarmellose sodium, colloidal silicon dioxide, and citric acid; the binder can be one or more of starch paste, ethanol, water, and povidone alcohol solution; the preservative can be one or more of ethyl p-hydroxybenzoate, propyl hydroxybenzoate, sorbic acid, potassium sorbate, calcium propionate, sodium dehydroacetate, sodium diacetate, and sodium lactate; the antioxidant can be one or more of ethylenediaminetetraacetic acid, disodium ethylenediaminetetraacetate, dibutylhydroxytoluene, glycine, inositol, ascorbic acid, sodium ascorbate, lecithin, malic acid, hydroquinone, citric acid, succinic acid, and sodium metabisulfite; the flavoring agent can be one or more of aspartame, sucrose, xylitol, stevioside, sodium cyclamate, sorbitol, cocoa, pure vanilla, vanillin, ethyl vanillin, chocolate, malt, and mint; the suspending agent can be one or more of xanthan gum, polyvinylpyrrolidone, sodium alginate, aluminum stearate, and hydrogenated vegetable oil; the emulsifier can be one or more of alkyl sulfates, soaps, dodecylbenzenesulfonates, lactate esters, sulfosuccinates, monoglyceride sulfonates, phosphate esters, siloxanes, and taurates.

[0031] The pharmaceutical composition can be prepared according to methods known in the art. For this purpose, if necessary, the active ingredient can be combined with one or more solid or liquid pharmaceutical excipients and / or adjuvants to form a suitable administration form or dosage form for human use.

[0032] In addition, if necessary, colorants, preservatives, fragrances, flavoring agents, sweeteners, or other materials can also be added to the pharmaceutical preparation.

[0033] The dosage of the pharmaceutical composition of the present invention depends on many factors, such as the sex, age, weight and individual response of the patient or animal, the route of administration and the frequency of administration, etc. The above dosage can be administered in a single dosage form or divided into several, for example, two, three or four dosage forms. The dosage level must be selected according to the specific route of administration, the severity of the condition to be treated and the condition and past medical history of the patient to be treated, etc. However, the practice in the art is that the dosage starts from a level lower than that required to obtain the desired therapeutic effect and is gradually increased until the desired effect is obtained.

[0034] It should be recognized that the total daily dosage of the pharmaceutical composition 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 depend on a variety of factors, including the disorder being treated and the severity of the disorder; the specific composition employed; the patient's age, weight, general health, sex and diet; the time of administration, the route of administration and the excretion rate; the duration of treatment; the drugs used in combination or concurrently; and similar factors well known in the medical field. For example, the practice in the art is that the dosage of administration starts from a level lower than that required to obtain the desired therapeutic effect and is gradually increased until the desired effect is obtained.

[0035] Generally speaking, the dosage of the pharmaceutical composition of the present invention calculated based on the active ingredient for mammals, especially humans, can range from 1 - 1000 mg / kg body weight / day, for example, from 1 - 500 mg / kg body weight / day, for example, from 50 - 500 mg / kg body weight / day, or 100 - 500 mg / kg, or 150 - 500 mg / kg, or 200 - 500 mg / kg, or 250 - 500 mg / kg.

[0036] Compared with the prior art, the present invention discloses for the first time a new medical use of Ganoderma lucidum, ginseng and kudzu root formula. Through experiments and clinical verification, this formula can increase the activities of key enzymes in the gastric and liver tissues of mice with acute alcoholic gastric mucosal injury, reduce the content of pro-inflammatory factor TNF-α in the gastric tissue, increase the production of anti-inflammatory factor IL-10 in the gastric tissue, alleviate the inflammatory injury of ethanol to the gastric mucosa, increase the levels of protective factors EGF, SS, PGE-2 in the serum, enhance the defense mechanism of the gastric mucosa and improve the repair ability of the gastric mucosa. This discovery will have a significant impact on the research and treatment of acute alcoholic gastric mucosal injury diseases and has clinical practical value. BRIEF DESCRIPTION OF THE DRAWINGS

[0037] Figure 1 It is a diagram of the HE staining result of the gastric tissue of mice in the present invention;

[0038] Figure 2 It is an AB-PAS staining diagram of the gastric tissue of mice in the present invention;

[0039] Figure 3 Effects of the present invention on the activities of key enzymes in mouse gastric tissue; 3A shows the expression level of ADH in gastric tissue; 3B shows the expression level of ALDH in gastric tissue; *P < 0.05;

[0040] Figure 4 Effects of the present invention on gastric tissue protective factors in mouse gastric tissue; 4A shows the expression level of PGE2 in gastric tissue; 4B shows the expression level of EGF in gastric tissue; 4C shows the expression level of SS in gastric tissue; *P < 0.05;

[0041] Figure 5 Effects of the present invention on inflammation-related proteins in mouse gastric tissue; 5A shows the expression level of TNF-α; 5B shows the expression level of IL-10; *P < 0.05. Detailed implementation manners

[0042] To enable those skilled in the art to better understand the technical solution of the present invention, the present invention will be described in detail below in combination with specific implementation manners. For the experimental methods without specific conditions indicated in the following examples, they are usually in accordance with conventional conditions or the conditions recommended by the manufacturer. The test materials used in the following examples are all obtained from regular biochemical reagent stores without special instructions. Unless otherwise stated, percentages and parts are calculated by weight. Unless otherwise defined, all professional and scientific terms used herein have the same meaning as those familiar to those skilled in the art. In addition, any methods and materials similar or equivalent to the described content can be applied to the present invention. The preferred implementation methods and materials described herein are only for demonstration purposes.

[0043] The endpoints and any values within the ranges disclosed in this document are not limited to the exact ranges or values. These ranges or values should be understood to include values close to these ranges or values. For numerical ranges, the values between the endpoints of each range, between the endpoints of each range and individual point values, and between individual point values can be combined with each other to obtain one or more new numerical ranges, and these numerical ranges should be regarded as specifically disclosed in this document.

[0044] The technical solution of the present invention will be further described in detail below in combination with specific examples and drawings. It should be understood that the following examples are only used to explain the present invention and are not used to limit the present invention.

[0045] Example 1

[0046] 6 parts of Ganoderma lucidum, 9 parts of Pueraria lobata, 9 parts of Panax ginseng, 15 parts of Poria cocos, 220 mg / kg of oligomeric walnut peptide.

[0047] Example 2

[0048] 3 parts of Ganoderma lucidum, 15 parts of Pueraria lobata, 12 parts of Panax ginseng, 20 parts of Poria cocos, 110 mg / kg of oligomeric walnut peptide.

[0049] Example 3

[0050] 10 parts of Ganoderma lucidum, 6 parts of Pueraria lobata, 9 parts of Panax ginseng, 12 parts of Poria cocos, 300 mg / kg of oligomeric walnut peptide.

[0051] Example 4

[0052] 9 parts of Ganoderma lucidum, 15 parts of Pueraria lobata, 9 parts of Panax ginseng, 18 parts of Poria cocos, 440 mg / kg of oligomeric walnut peptide.

[0053] Example 5

[0054] 5 parts of Ganoderma lucidum, 12 parts of Pueraria lobata, 12 parts of Panax ginseng, 15 parts of Poria cocos, 150 mg / kg of oligomeric walnut peptide.

[0055] Investigate the synergistic protective effect of the pharmaceutical composition on mice with acute alcoholic gastric mucosal injury

[0056] Experimental animals: C57BL / 6J mice, male, 15 mice, body weight 18 - 22 g, provided by Beijing Speyford Biotechnology Co., Ltd.

[0057] Drug sources: Panax ginseng, Ganoderma lucidum, Poria cocos, Pueraria lobata, purchased from "Yixintang" pharmacy in Yunnan and identified by the School of Traditional Chinese Medicine of Yunnan University of Traditional Chinese Medicine; walnut peptide, Shaanxi Parne Biotechnology Co., Ltd.; Tumor Necrosis Factor-α (TNF-α), Interleukin 10 (IL-10), Prostaglandin E2 (PGE2), Epidermal Growth Factor (EGF), Somatostatin (SS) enzyme-linked immunosorbent assay (ELISA) kits, Jiangsu Enzyme-linked Biotechnology Co., Ltd.; Alcohol dehydrogenase (ADH), Acetaldehyde dehydrogenase (ALDH) kits, Beijing Solarbio Science & Technology Co., Ltd.

[0058] Experimental method: C57BL / 6J mice were randomly divided into 3 groups, with 5 mice in each group: blank group (Control, C), model group (Model, M), and Ganoderma lucidum, Panax ginseng, and Pueraria lobata formula group (M+LSD). After 3 days of adaptive feeding (day, d), the mice were fasted for 24 hours without water deprivation. The model group (Model, M) and the Ganoderma lucidum, Panax ginseng, and Pueraria lobata formula group (M+LSD) were intragastrically administered 56% ethanol at a dose of 10 mL / kg, and the blank group (Control, C) was given an equal amount of normal saline. The intervention was carried out every other day, once a day, for a total of 4 times. At the same time, the drug intervention experiment was started on the first day. The Ganoderma lucidum, Panax ginseng, and Pueraria lobata formula group was administered the drug composition of Example 1 at a dose of 6.24 g / kg per day, and the model group and the blank group were both given an equal amount of normal saline for 7 days.

[0059] The experimental data were expressed as "mean ± standard deviation" ", and SPSS 29.0 software was used for one-way ANOVA for comparison among multiple groups, and the LSD method was used for pairwise comparison between groups. GraphPad Prism 10.4.1 software was used for drawing, and P<0.05 was considered statistically significant.

[0060] (1) Observe the effect of drug administration on the histopathology of mice

[0061] Observation of histopathological changes under HE staining: After the drug intervention experiment, gastric tissues were taken and fixed with 4% paraformaldehyde for standby. The fixed tissues were dehydrated step by step, then embedded in paraffin, sectioned (about 4 μm thick), deparaffinized, and stained with hematoxylin-eosin (HE), and then observed under a microscope for pathological changes of the tissues.

[0062] As Figure 1 shown, the gastric mucosa epithelial structure of the mice in the blank group was intact, the glands were arranged neatly, the structure of the mucosa layer was intact and clear, and no obvious edema or inflammatory cell infiltration was seen. Compared with the blank group, the normal structure of the gastric mucosa tissue of the mice in the model group disappeared, the epithelial cells were arranged disorderly or partially exfoliated, there was inflammatory cell infiltration, and the mucosa layer was edematous. After drug administration, the gastric mucosa tissue structure of the mice in the Ganoderma lucidum, Panax ginseng, and Pueraria lobata formula group was intact, the glandular cells were arranged relatively neatly, and there was no obvious edema in the mucosa layer, which was relatively close to the normal gastric tissue structure. It is suggested that Ganoderma lucidum, Panax ginseng, and Pueraria lobata formula can effectively relieve gastric mucosa damage.

[0063] Observation of histopathological changes of mouse gastric tissue under AB-PAS staining Alcian blue-periodic acid-Schiff (AB-PAS) staining can combine with the colorless fuchsin in the Schiff reagent to form a purplish-red dye deposit and show color at the corresponding site. The paraffin sections of gastric tissues were stained with AB-PAS, sealed, observed and photographed with a digital slide scanner, and specific lesions were observed.

[0064] AsFigure 2 As shown, the gastric mucosa of the mice in the blank group was normal in morphology, with regular and tight cell arrangement, intact cell and nuclear polarity, and no obvious blue-stained foci, indicating that there were no glands secreting mucus. Compared with the blank group, the gastric mucosa structure of the mice in the model group was severely damaged, with disordered cell arrangement, a large number of inflammatory cell infiltrations, obvious vacuolization of cells, mucus lakes of different sizes formed in the glandular lumen, and deep blue staining in the glands, indicating the formation of intestinal metaplasia glands secreting acidic mucus. Compared with the model group, the abnormal cell morphology of the gastric mucosa tissue of the mice in the Ganoderma lucidum, Panax ginseng and Pueraria lobata formula group improved, the inflammatory infiltration area decreased, and the vacuolization, mucus lake and tissue atypia decreased, indicating that the Ganoderma lucidum, Panax ginseng and Pueraria lobata formula can improve the gastric mucosa defense barrier.

[0065] (2) Observe the effects of drug administration on the activities of key enzymes ADH and ALDH in mouse gastric tissue

[0066] After homogenizing and centrifuging the mouse gastric tissue, the supernatant was taken, and samples were prepared according to the instructions of the detection kit. The activity of alcohol dehydrogenase (ADH) was detected by the enzyme-coupled rate method, and the activity of aldehyde dehydrogenase (ALDH) was detected by the colorimetric method combined with the catalytic reaction of aldehyde dehydrogenase.

[0067] Figure 3 As can be seen, the activities of key enzymes ADH and ALDH in the gastric tissue of the mice in the model group were significantly lower than those in the blank control group. After drug administration, the activities of alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) in the Ganoderma lucidum, Panax ginseng and Pueraria lobata formula group were significantly increased. It can be seen that drug administration can effectively increase the expression of ADH and ALDH in mice, regulate the ethanol metabolism enzyme system, and thus improve the damage.

[0068] (3) Observe the effects of drug administration on the protective factors of mouse gastric tissue

[0069] Use enzyme-linked immunosorbent assay to detect the gastric tissue protective factors PGE2, EGF, and SS. Specifically:

[0070] 1) Sample treatment: About 20 mg of gastric tissue samples were rinsed with normal saline to remove possible blood and impurities. Then, normal saline equivalent to 9 times its mass was added, and the tissue was homogenized into a homogenate using a tissue grinder. Then, it was centrifuged at 4 °C for 15 minutes, and the supernatant was aspirated and kept on ice.

[0071] 2) Immunoassay: ELISA kits were used to quantitatively analyze PGE2, EGF, and SS in gastric tissues. The kits were left at room temperature for 30 minutes to reach equilibrium. Standard curve wells, blank control wells, and test wells for each sample were prepared according to the instructions. 50 μL of the corresponding standard was added to each standard well; 45 μL of diluent and 5 μL of the sample were added to each sample well. Then, 100 μL of the corresponding antibody was added to each sample well and standard well, except for the blank well. The plate was placed in a sealed incubator at 37°C for 1 hour to ensure sufficient antibody-binding reaction.

[0072] 3) Plate washing and color development: After incubation, the plate was washed 5 times to remove unbound antibodies and other non-specific binding substances. Then, chromogenic substrates A and B were added to each well, and the plate was incubated in the dark in a sealed incubator at 37°C for 15 minutes for color development reaction. After color development was completed, the stop solution was quickly added to stop the reaction.

[0073] 4) Content determination: The absorbance values of each well were read using an enzyme-linked immunosorbent assay (ELISA) reader at a wavelength of 450 nm. This value reflects the concentration of each detection index, and the content of the target protein in the sample can be quantitatively analyzed by combining with the standard curve.

[0074] In acute gastric mucosal injury, the level of SS showed a trend of "first increasing and then decreasing". An increase in SS secretion was observed in the early stage, which inhibited gastric acid as a protective reaction. A decrease in SS secretion was seen in the later stage, exacerbating gastric acid erosion and mucosal ischemia and delaying healing; while EGF could counteract the destructive effect of pepsin on the gastric mucosa and simultaneously inhibit excessive gastric acid secretion, thus providing protection for the gastric mucosa and maintaining the integrity of its structure and function; PGE2 is an inhibitor of monocyte pro-inflammatory factor expression, which can relieve oxidative stress and play an anti-inflammatory and antipyretic role. At the same time, PGE2 can protect the gastric mucosa by inhibiting gastric acid secretion, regulating gastric wall mucus secretion, and improving gastric mucosal blood circulation.

[0075] As Figure 4 shown, the levels of EGF, SS, and PGE-2 in the gastric tissues of mice in the model group were significantly lower than those in the blank group. After administration, the levels of EGF, SS, and PGE-2 in the gastric tissues of mice in the Ganoderma lucidum, Panax ginseng, and Pueraria lobata formula group were significantly increased. It is suggested that the Ganoderma lucidum, Panax ginseng, and Pueraria lobata formula of the present invention can play a protective role by increasing the levels of protective factors.

[0076] (4) Observe the effect of drug administration on inflammatory factors in mouse gastric tissues

[0077] Mouse serum was taken and the serum inflammatory factors IL-10 and TNF-α were detected by enzyme-linked immunosorbent assay. The specific method was as follows: 40 μL of sample diluent, 10 μL of sample, and 100 μL of enzyme-labeled reagent were added to each sample well; 50 μL of each standard solution and 100 μL of enzyme-labeled reagent were added to the standard wells; 40 μL of sample diluent was added to the blank well; the plate was sealed with a sealing film and incubated in a constant temperature incubator at 37 °C in the dark for 1 h; the plate was quickly washed 5 times with a multi-channel pipette, and the plate was patted dry after each wash; then 50 μL of chromogenic reagent A and 50 μL of chromogenic reagent B were added respectively, and incubated in the dark at a constant temperature for 15 min; the optical density (OD value) of each well was measured successively with an enzyme-labeled instrument at a wavelength of 450 nm, and the concentration was calculated according to the standard curve (see Figure 5 ).

[0078] As Figure 5 shown, compared with the model group, the content of TNF-α in the gastric tissues of mice in the blank group and the Ganoderma lucidum, Panax ginseng and Pueraria lobata formula group was significantly decreased, while the content of IL-10 was significantly increased. TNF-α is a pro-inflammatory factor for judging the degree of inflammation, which can induce the release of platelet-activating factor, and then accelerate the generation of harmful substances such as oxygen free radicals and leukotrienes, causing hypoxia and ischemia of the intestinal mucosa in the body and aggravating the damage of gastric mucosa. The anti-inflammatory factor IL-10 can inhibit the chemotactic functions of macrophages and neutrophils and reduce the activity of pro-inflammatory factors. It is shown that the Ganoderma lucidum, Panax ginseng and Pueraria lobata formula of the present invention has good anti-inflammatory activity on mice with acute alcoholic gastric mucosal injury.

[0079] Finally, it should be noted that the above description is only the preferred embodiment of the present invention. Those skilled in the art can make various similar representations under the inspiration of the present invention without violating the purpose and claims of the present invention. Such transformations all fall within the protection scope of the present invention.

Claims

1. A pharmaceutical composition for improving acute alcoholic gastric mucosal injury, characterized in that, It consists of the following raw materials: 3 - 10 parts by weight of Ganoderma lucidum, 6 - 15 parts by weight of Pueraria lobata, 6 - 15 parts by weight of Panax ginseng, 12 - 20 parts by weight of Poria cocos, and 110 - 440 mg / kg of oligomeric walnut peptide.

2. The pharmaceutical composition according to claim 1, wherein The raw material composition is: 3 - 10 parts by weight of Ganoderma lucidum, or 3 - 10 parts by weight, or 5 - 10 parts by weight, or 3 - 6 parts by weight, or 6 - 9 parts by weight, or 6 parts by weight; and / or 6 - 15 parts by weight of Pueraria lobata, or 6 - 12 parts by weight of Pueraria lobata, or 6 - 9 parts by weight of Pueraria lobata, or 9 - 15 parts by weight of Pueraria lobata, or 9 - 12 parts by weight of Pueraria lobata, or 9 parts by weight; and / or 6 - 15 parts by weight of Panax ginseng, or 6 - 12 parts by weight of Panax ginseng, or 6 - 9 parts by weight of Panax ginseng, or 9 - 15 parts by weight of Panax ginseng, or 9 - 12 parts by weight of Panax ginseng, or 9 parts by weight; and / or 12 - 20 parts by weight of Poria cocos, or 15 - 20 parts by weight of Poria cocos, or 15 - 18 parts by weight of Poria cocos, or 12 - 18 parts by weight of Poria cocos, or 12 - 15 parts by weight of Poria cocos, or 15 parts by weight; and / or 110 - 440 mg / kg of oligomeric walnut peptide, or 110 - 380 mg / kg of oligomeric walnut peptide, or 110 - 350 mg / kg of oligomeric walnut peptide, or 150 - 300 mg / kg of oligomeric walnut peptide, or 200 - 250 mg / kg of oligomeric walnut peptide, or 220 mg / kg of oligomeric walnut peptide.

3. The pharmaceutical composition according to claim 1, wherein The Ganoderma lucidum is one or a combination of Ganoderma lucidum powder and Ganoderma lucidum extract; and / or The Pueraria lobata is one or a combination of Pueraria lobata powder and Pueraria lobata extract; and / or The Panax ginseng is one or a combination of Panax ginseng powder and Panax ginseng extract; and / or The Poria cocos is one or a combination of Poria cocos powder and Poria cocos extract.

4. Use of the pharmaceutical composition according to any one of claims 1 - 3 in the preparation of a drug for improving acute alcoholic gastric mucosal injury.

5. The application according to claim 4, wherein, The pharmaceutical composition as an active ingredient can be used in combination in the preparation of the drug, or can be compounded with other components having anti - inflammatory activity. Among them, the active component composition is: Ganoderma lucidum, Pueraria lobata, Panax ginseng, Poria cocos, oligomeric walnut peptide.

6. The application according to claim 4, characterized in that, The drug does not contain other effective anti - inflammatory active ingredients except the pharmaceutical composition. Among them, the active component composition is: Ganoderma lucidum, Pueraria lobata, Panax ginseng, Poria cocos, oligomeric walnut peptide.

7. The application according to claim 7, characterized in that, The drug uses the pharmaceutical composition as an active ingredient and is made into a pharmaceutically acceptable dosage form with pharmaceutically acceptable excipients. Among them, the active component composition is: Ganoderma lucidum, Pueraria lobata, Panax ginseng, Poria cocos, oligomeric walnut peptide.

8. A pharmaceutical preparation, characterized in that, The pharmaceutical composition according to any one of claims 1 - 4 is made into a pharmaceutically acceptable dosage form with pharmaceutically acceptable excipients.

9. The pharmaceutical preparation according to claim 4, wherein The dosage form of the pharmaceutical preparation is an oral dosage form or an injection dosage form, and can also adopt controlled - release or sustained - release dosage forms well - known in the modern pharmaceutical industry.

10. The pharmaceutical preparation according to claim 4, characterized in that, The excipients include one or more of solvents, disintegrants, flavoring agents, preservatives, coloring agents, binders, lubricants, diluents, and drug carriers.

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