KR20230172387A - Composition for ameliorating intestinal health or treating inflammatory bowel disease comprising Daehong Crataegus pinnatifida Bunge extract as effective component - Google Patents

Abstract

The present invention relates to a composition for improving intestinal health or treating inflammatory bowel disease containing Dahongsansa extract as an active ingredient. The Dahongsansa extract has little cytotoxicity, has intestinal protective activity, and contains necroptosis markers. Since it reduces the expression level and has the effect of restoring intestinal length in animal models with inflammatory bowel disease, it can be usefully used as a health functional food or medicine for intestinal health.

Description

Composition for improving intestinal health or treating inflammatory bowel disease comprising Daehong Crataegus pinnatifida Bunge extract as effective component}

The present invention relates to a composition for improving intestinal health or treating inflammatory bowel disease containing Dahongsansa extract as an active ingredient.

In addition to the functions of food digestion, absorption, and excretion, the human intestines have a very close and important relationship with immunity, to the extent that approximately 70% of the total immune cells of the entire body are gathered, and are immune to mechanical stress, carcinogens from the external environment, and It is continuously damaged by chemical stress and biological stress from harmful bacteria in the intestines, which has a negative impact on intestinal epithelial cell damage, stem cell production, and mucus production, posing a threat to intestinal health. Due to westernized and inappropriate eating habits, the number of patients with intestinal diseases such as inflammatory bowel disease and irritable bowel syndrome is increasing, and many metabolic diseases, such as diabetes, degenerative brain diseases such as Parkinson's and Alzheimer's diseases, are also reported to be ultimately related to intestinal health. The correlation with adult diseases is becoming important.

In particular, inflammatory bowel disease (IBD) is classified into two diseases, ulcerative colitis and Crohn's disease, which are clinically similar but different in terms of histological findings, endoscopy, and immunology. IBD is a disease of inflammatory cells. Activation is known to be an important etiological factor. Persistent or inappropriate activation of the intestinal immune system plays an important role in the pathophysiology of chronic mucosal inflammation, especially through infiltration of neutrophils, macrophages, lymphocytes, and mast cells, ultimately leading to mucosal destruction and ulceration. Infiltrated and activated neutrophils are an important source of reactive oxygen or reactive nitrogen species, which are cytotoxic substances that induce cellular oxidative stress by cross-linking proteins, lipids and nucleic acids and cause epithelial dysfunction and damage. . In inflammatory diseases, various inflammatory cytokines are secreted from the intestinal mucosa. TNF-α is highly expressed in the colonic lumen and colonic epithelial cells of patients with ulcerative colitis, and recent studies have shown that TNF-α plays an important role in the pathogenesis of ulcerative colitis. Infliximab, an anti-TNF-α antibody, is known to be effective not only in the treatment of boils but also in the treatment of previously untreated Crohn's disease. Currently, inflammatory bowel disease treatments include 5-aminosalicylic acid (5-ASA) drugs that block the production of prostaglandins, such as sulfasalazine, or steroid-type immunosuppressants. .

Meanwhile, the mountain temple is round and red in appearance and has white spots. The outer surface is yellow-brown or grayish-red-brown and has many fine wrinkles. On one side, there is a concave area with a diameter of 5 mm, and sometimes calyxes remain around it, and on the other side, fruit buds or marks remain. On the side cut side, there is a five-chambered ovary, and each chamber contains one seed. The seeds are shaped like balls divided into equal parts, 6 to 8 mm long, the outer surface is light brown and hard, and the quality is hard. It is also called Sansa, Sanlihong, Honggwa, Buksansa, Jeokgwaja, Sanjohong, Honggwaja, Yangguja, Dangguja, Mosa, Sanligwa, Seo, Seo, Yanggu, Jeoksil, Husa, Guja, and Gyemae.

The flowers of the mountain temple bloom white in May, and the fruit is a pome fruit that is round and has white spots. In oriental medicine, it is known to have effects such as stomach health, digestion, astringency, and pain relief. In Europe, the fruit of the European hawthorn is also called crataegus and is used as a tonic. Pharmacological ingredients include caffeic acid, citric acid, corosolic acid, crateegolic acid, euscaphic acid, hyperin, and hyperoic acid. Hyperoside, procatechuic acid, pyrogallol, oleanolic acid, ursolic acid, vitexin, etc. have been reported.

As for technologies related to mountain temples, Korean Patent No. 2349702 discloses a composition for skin whitening containing mountain temple extracts or fractions thereof, and Korean Patent Publication No. 2005-0079256 further includes mixed extracts of Houttuynia cordata, bokbunja and mountain lions. Although compositions for the prevention and treatment of allergic diseases have been disclosed for the mixture, a composition for improving intestinal health or treating inflammatory bowel disease containing the Dahongsansa extract of the present invention as an active ingredient has not yet been disclosed.

The present invention was developed in response to the above-mentioned needs, and the present invention provides a composition for improving intestinal health or treating inflammatory bowel disease containing Dahongsansa extract as an active ingredient, and the Dahongsansa extract has little cytotoxicity and The present invention was completed by confirming that it has intestinal protective activity, reduces the expression level of necroptosis markers, and has the effect of restoring intestinal length in an animal model induced by inflammatory bowel disease.

In order to achieve the above object, the present invention provides a health functional food composition for improving intestinal function or inflammatory bowel disease containing Daehong Crataegus pinnatifida Bunge extract as an active ingredient.

In addition, the present invention provides a pharmaceutical composition for the prevention or treatment of irritable bowel syndrome or inflammatory bowel disease, comprising an extract of Dahongsansa as an active ingredient.

In addition, the present invention provides a herbal medicine composition for preventing or treating irritable bowel syndrome or inflammatory bowel disease, containing Dahongsansa extract as an active ingredient.

In addition, the present invention provides a feed additive for improving intestinal function or inflammatory bowel disease containing Daehongsansa extract as an active ingredient.

In addition, the present invention provides a veterinary composition for the prevention or treatment of irritable bowel syndrome or inflammatory bowel disease containing Dahongsansa extract as an active ingredient.

The present invention relates to a composition for improving intestinal health or treating inflammatory bowel disease containing Dahongsansa extract as an active ingredient. The Dahongsansa extract has little cytotoxicity, has intestinal protective activity, and contains necroptosis markers. It has the effect of reducing the expression level and restoring intestinal length in animal models caused by inflammatory bowel disease.

Figure 1 shows the cell viability results confirmed by treating HCE cells (Human Primary Colonic Epithelial cells) with Daehongsansa extract (A) and Crataegus extract (B).
Figure 2 shows the results of confirming the intestinal cell protective activity of the Dahongsansa extract. #### indicates a statistically significant increase in the number of Annexin V/PI positive cells (number of necrotic cells) in the STZ(+) group that induced cell necrosis compared to the normal control group, p<0.0001, **, *** indicates a statistically significant decrease in the number of Annexin V/PI positive cells (number of necrotic cells) in the group treated with the Dahongsansa extract of the present invention compared to the STZ(+) group, ** indicates p<0.01 and *** is p<0.001.
Figure 3 is a Western blot result confirming the change in protein expression level of cell necrosis marker according to treatment of Dahongsansa extract.
Figure 4 shows the results of confirming body weight changes after treating an animal model with induced inflammatory bowel disease with Dahongsansa extract. #### indicates a statistically significant decrease in body weight of the control group (inflammatory bowel disease induced group) compared to the normal group, p<0.0001, * indicates a statistically significant decrease in body weight of the control group (inflammatory bowel disease induced group), and * indicates a statistically significant decrease in body weight of the control group (inflammatory bowel disease induced group), compared to the inflammatory bowel disease induced group. The body weight of the group administered the extract significantly increased, p<0.05.
Figure 5 is a photograph (A) confirming the intestinal length and a graph (B) showing the intestinal length after treating an animal model in which inflammatory bowel disease was induced with Dahongsansa extract. #### indicates a statistically significant decrease in the intestine length of the control group (inflammatory bowel disease induced group) compared to the normal group, p<0.0001, *, **** compared to the inflammatory bowel disease induced group, according to the present invention. This means that the intestinal length of the group administered with Dahongsansa extract or hawthorn extract significantly increased, * means p < 0.05, and **** means p < 0.0001. $$ indicates that the long length of the Dahongsansa extract-administered group of the present invention increased statistically significantly compared to the comparison group, which was the Sansa extract-administered group, p<0.01.

The present invention relates to a health functional food composition for improving intestinal function or inflammatory bowel disease containing Daehong Crataegus pinnatifida Bunge extract as an active ingredient.

The Dahongshan Temple extract can be prepared by a method including the following steps, but is not limited to this:

(1) Extracting by adding an extraction solvent to dried Dahongsansa;

(2) filtering the extract of step (1); and

(3) Preparing an extract by drying the filtered extract of step (2).

In step (1), the extraction solvent is preferably selected from water, C ₁ -C ₄ lower alcohol, or a mixture thereof, more preferably water or ethanol, but is not limited thereto. In the above manufacturing method, the extraction method can be any conventional method known in the art, such as hot water extraction, immersion extraction, reflux cooling extraction, and ultrasonic extraction. It is preferable to extract the extraction solvent by adding 1 to 20 times the weight of Dahongsansa. The extraction temperature is preferably 20 to 100°C, but is not limited thereto. In addition, the extraction time is preferably 1 to 5 hours, and more preferably 2 to 4 hours, but is not limited thereto. In step (3), the drying is preferably reduced pressure drying, vacuum drying, boiling drying, spray drying, or freeze drying, more preferably reduced pressure drying, but is not limited thereto.

The Daehongsansa extract is preferably, but not limited to, the Daehong variety (variety protection application number 2010-10).

The improvement in intestinal function is preferably an improvement in irritable bowel syndrome, and the inflammatory bowel disease is preferably any one selected from ulcerative colitis, Crohn's disease, intestinal lesions accompanying Behcet's disease, bleeding rectal ulcers, and ileal pouchitis. However, it is not limited to this.

In the present invention, irritable bowel syndrome (IBS) refers to a functional disorder caused by hypersensitive contraction of the colonic muscles, although there is no other disease or anatomical abnormality, and causes abdominal pain, abdominal discomfort, defecation disorders, etc. accompany.

The health functional food composition is preferably manufactured in a dosage form selected from powder, granule, pill, tablet, capsule, candy, syrup and beverage, but is not limited thereto. The health functional food composition of the present invention can be prepared by adding the Daehongsansa extract as is or mixing it with other foods or food ingredients, and can be prepared appropriately according to conventional methods. Examples of foods to which the Dahongsansa extract can be added include caramel, meat, sausages, bread, chocolate, candies, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, It may be in any form selected from tea, drink, alcoholic beverage, and vitamin complex, and includes all health functional foods in the conventional sense. That is, there are no particular restrictions on the type of food. The health functional food composition includes various nutrients, vitamins, minerals (electrolytes), synthetic and natural flavors, colorants and enhancers (cheese, chocolate, etc.), pectic acid and its salts, alginic acid and its salts, organic acids, and protective colloidal thickeners. , pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, etc. Additionally, it may contain pulp for the production of natural fruit juice and vegetable drinks. The above components can be used independently or in combination.

The health functional food composition of the present invention may contain various flavoring agents or natural carbohydrates as additional ingredients, and the natural carbohydrates include monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, dextrin, and cyclodextrin. Polysaccharides such as xylitol, sorbitol, and erythritol are sugar alcohols. The ratio of the natural carbohydrate is not very important, but is preferably 0.01 to 0.04 g, more preferably 0.02 to 0.03 g, per 100 g of the composition of the present invention, but is not limited thereto. As sweeteners, natural sweeteners such as thaumatin and stevia extract, and synthetic sweeteners such as saccharin and aspartame can be used.

Additionally, the present invention relates to a pharmaceutical composition for the prevention or treatment of inflammatory bowel disease containing Dahongsansa extract as an active ingredient.

The inflammatory bowel disease is preferably, but not limited to, any one selected from the group consisting of ulcerative colitis, Crohn's disease, intestinal lesion accompanying Behçet's disease, bleeding rectal ulcer, and ileal pouchitis.

The pharmaceutical composition containing the Dahongsansa extract according to the present invention is preferably, but not limited to, any one formulation selected from capsules, powders, granules, tablets, suspensions, emulsions, syrups, and aerosols. In addition to the Dahongsansa extract, it may further include pharmaceutically acceptable carriers, excipients, or diluents. Carriers, excipients, or diluents that may be included in the pharmaceutical composition containing the Dahongsansa extract include lactose, dextrose, and sucrose. , oligosaccharides, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate. , propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. When formulated, it can be prepared using diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrants, and surfactants. The preferred dosage of the Dahongshan Temple extract of the present invention varies depending on the patient's condition and weight, degree of disease, drug form, administration route and period, but can be appropriately selected by a person skilled in the art. However, for a desirable effect, the pharmaceutical composition containing the Dahongsansha extract of the present invention is preferably administered at 0.0001 to 100 mg/kg per day, preferably at 0.001 to 10 mg/kg. Administration may be administered once a day, or may be administered several times. The above dosage does not limit the scope of the present invention in any way.

Additionally, the present invention relates to a herbal medicine composition for the prevention or treatment of irritable bowel syndrome or inflammatory bowel disease, containing Dahongsansa extract as an active ingredient.

The herbal medicine composition according to the present invention refers to a conventional dried medicinal material used in the manufacture of herbal medicine. It may be an extract obtained by extracting the herbal medicine composition of the present invention using an extraction solvent selected from water, C ₁ to C ₄ lower alcohol, or a mixture thereof according to a conventional method, or it may be an extract in powder form obtained by drying it, especially There are no restrictions on dosage form. For example, it can be prepared in the form of pills, tablets, capsules, or liquid.

In addition, the present invention relates to a feed additive for improving intestinal function or inflammatory bowel disease containing Dahongsansa extract as an active ingredient.

The feed additive of the present invention corresponds to supplementary feed under the Feed Management Act. In the present invention, the term 'feed' may mean any natural or artificial diet, meal, etc., or a component of the meal, for or suitable for eating, ingestion, and digestion by animals. The type of feed is not particularly limited, and feed commonly used in the art can be used. Non-limiting examples of the feed include plant feeds such as grains, roots and fruits, food processing by-products, algae, fiber, pharmaceutical by-products, oils and fats, starches, cucurbits or grain by-products; Examples include animal feeds such as proteins, inorganic substances, fats and oils, minerals, oils and fats, single-cell proteins, zooplanktons or food. These may be used alone or in combination of two or more types.

In addition, the present invention relates to a veterinary composition for the prevention or treatment of irritable bowel syndrome or inflammatory bowel disease containing Dahongsansa extract as an active ingredient.

The veterinary composition of the present invention may further include appropriate excipients and diluents according to conventional methods. Excipients and diluents that may be included in the veterinary composition of the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, Cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methyl hydroxy benzoate, propyl hydroxy benzoate, talc, magnesium stearate, ethanol, stearyl alcohol, liquid paraffin, sorbitan monostearate, Examples include polysorbate 60, methylparaben, propylparaben, and mineral oil. The veterinary composition according to the present invention may further include fillers, anti-aggregants, lubricants, wetting agents, spices, emulsifiers, preservatives, etc. The veterinary composition according to the present invention provides rapid and sustained release of the active ingredient after administration to an animal. or may be formulated using methods well known in the art to provide sustained release, the dosage form being powders, granules, tablets, capsules, suspensions, emulsions, solutions, syrups, aerosols, soft or hard gelatin capsules, It may be in the form of a suppository, sterile injectable solution, or sterile topical medication. The effective amount of the veterinary composition according to the present invention can be appropriately selected depending on the individual animal. Severity of the disease or condition, sensitivity to the active ingredient of the present invention depending on the individual's age, weight, health condition or gender, administration route, administration period, factors including other compositions mixed or used simultaneously with the composition, and other physiological or It can be determined based on factors well known in the veterinary field.

Hereinafter, the present invention will be described in more detail using examples. These examples are only for illustrating the present invention in more detail, and it is obvious to those skilled in the art that the scope of the present invention is not limited thereto.

Example 1. Preparation of Dahongshan Temple extract

10 times the weight of dried Dahongsansa was added with 70% ethanol, and refluxed extraction was performed at 84 ± 2°C for 3 hours. Afterwards, the obtained Dahongsansa extract was filtered using a 25㎛ filter. Afterwards, it was concentrated under reduced pressure at 50°C and dried.

Example 2. Confirmation of cytotoxicity

To confirm the cytotoxicity of Dahongshansa extract, MTT (Mitochondrial NADH-dehydrogenase) assay was performed.

1.0 × 10 ⁴ HCE cells (Human Primary Colonic Epithelial cells) were prepared in 96 wells. After 24 hours, the Dahongsansa extract extracted with ethanol as a solvent was treated and cultured for further 24 and 48 hours. Absorbance was measured at 490 nm. Each experiment was performed three times.

As a result, as shown in Figure 1, both the Dahongsansa extract and the comparative example Sansa extract (conventional variety) did not show cytotoxicity in normal colon cells up to 1600㎍/㎖.

Example 3. Confirmation of enterocyte protective activity of Daehongsansa extract

2 × 10 ⁶ HCE (Human Primary Colonic Epithelial) cells per well were prepared in a 6-well plate. After 24 hours, the Dahongshansa extract extracted with 70% ethanol was treated with medium for 30 minutes, and STZ (20 μM of Smac, 10 μM of z-VAD, and 10 ng/ml of TNFα complex) was treated for 12 hours to incubate the cells. Necroptosis was induced. Afterwards, cell death values were quantified using FACS (Annexin V/PI staining, BD biosciences).

As a result, more than 70% of the intestinal cells in the STZ-treated group died, and in the group treated with Daehongsansa extract and STZ (STZ+Dahongsansa), cell necrosis of colonic epithelial cells decreased in a concentration-dependent manner of the Daehongsansa extract. It was determined that the hawthorn extract had intestinal cell protective activity (Figure 2).

Example 4. Confirmation of changes in expression level of necroptosis markers according to treatment of Daehongsan Temple extract

2 × 10 ⁶ HCE (Human Primary Colonic Epithelial) cells per well were prepared in a 6-well plate. After 24 hours, the Dahongshansa extract extracted with 70% ethanol was treated with medium for 30 minutes, and STZ (20 μM of Smac, 10 μM of z-VAD, and 10 ng/ml of TNFα complex) was treated for 12 hours to incubate the cells. Necroptosis was induced. Afterwards, Western blot was performed to confirm changes in protein expression levels of cell necrosis markers.

As a result. As shown in Figure 3, the cell necrosis markers p-RIP1 (receptor-interacting serine/threonine-protein kinase-1), pRIP3 (receptor-interacting serine/threonine-protein kinase- 3) and the protein expression level of MLKL (mixed-lineage kinase domain-like pseudokinase) decreased.

Example 5. Confirmation of changes in intestinal length according to treatment of Daehongshansa extract using a DSS-induced inflammatory bowel disease animal model

C57BL/6-female mice (5 per group) were given 2.5% DSS (Dextran sulfate sodium) in a drinking container and allowed to drink voluntarily (the DSS drinking container was replaced with a new one every 2 days). Dried Dahongshansa extract was dissolved in water and administered orally at a single concentration (400mg/kg/day) for 3 weeks, with DSS administered for the last week. After a total of 3 weeks, an autopsy was performed on the animal, and body weight and colon length were measured.

As a result, when 200 mg/kg/day of Dahongsansa extract was administered orally for 3 weeks as shown in Figure 4, the body weight that had been reduced by DSS administration was significantly recovered, and as shown in Figure 5, the colon length increased. The change showed that both the hawthorn extract-administered group and the Daehongsansa extract increased the intestine length compared to the intestinal disease-induced group, and in particular, the Daehongsansa extract-administered group showed a statistically significantly greater increase in intestine length than the hawthorn extract-administered group.

Claims (11)

A health functional food composition for improving intestinal function or inflammatory bowel disease containing Daehong Crataegus pinnatifida Bunge extract as an active ingredient. The health functional food composition for improving intestinal function or inflammatory bowel disease according to claim 1, wherein the extraction solvent of the Dahongsansa extract is water, C ₁ to C ₄ lower alcohol, or a mixture thereof. The health functional food composition for improving intestinal function or improving inflammatory bowel disease according to claim 1, wherein the improving intestinal function is improving irritable bowel syndrome. The method of claim 1, wherein the inflammatory bowel disease is any one selected from ulcerative colitis, Crohn's disease, intestinal lesions accompanying Behcet's disease, hemorrhagic rectal ulcers, and ileal pouchitis. Health functional food composition for improvement. The health functional food for improving intestinal function or inflammatory bowel disease according to claim 1, wherein the composition is manufactured in any one formulation selected from powder, granule, pill, tablet, capsule, candy, syrup, and beverage. Composition. A pharmaceutical composition for the prevention or treatment of irritable bowel syndrome or inflammatory bowel disease, comprising an extract of Dahongsansa as an active ingredient. The pharmaceutical composition for preventing or treating irritable bowel syndrome or inflammatory bowel disease according to claim 6, wherein the composition further comprises a pharmaceutically acceptable carrier, excipient, or diluent in addition to the active ingredient. The method of claim 6, wherein the composition is prepared in any one formulation selected from capsules, powders, granules, tablets, suspensions, emulsions, syrups, and aerosols for the prevention or treatment of irritable bowel syndrome or inflammatory bowel disease. Pharmaceutical composition. A herbal medicine composition for the prevention or treatment of irritable bowel syndrome or inflammatory bowel disease, comprising an extract of Dahongsansa as an active ingredient. A feed additive for improving intestinal function or inflammatory bowel disease containing Dahongsansa extract as an active ingredient. A veterinary composition for the prevention or treatment of irritable bowel syndrome or inflammatory bowel disease, comprising an extract of Dahongsansa as an active ingredient.