KR20210003670A - Anti-allergy composition comprising velutin - Google Patents

Abstract

The present invention relates to a composition for preventing, treating or alleviating allergic disease, including velutin. The composition including velutin of the present invention inhibits the release of histamine from mast cells in a concentration-dependent manner to exhibit an effect of inhibiting degranulation. Therefore, the composition including velutin of the present invention can be usefully used as an antihistamine composition with alleviated side effects in the fields of cosmetics, foods or pharmaceuticals requiring an anti-allergic composition.

Description

Anti-allergy composition comprising velutin

The present invention relates to a composition for preventing, treating or improving allergic diseases, including velutin.

Flavonoids are a kind of phenolic compounds that exist in nature, and various physiological activities such as antioxidant, anti-inflammatory, antibacterial, and anti-cancer have been reported. Due to the various physiological activities of flavonoids, they are widely used as health functional foods, functional cosmetics, and pharmaceutical materials. In particular, structural diversity is possible depending on the position of the hydroxy group and the methoxy group of the flavonoid, as well as various physiological activities.

Velutin is a type of flavonoid and is found in natural products such as Viscum album, Enoki mushroom (Flammulina velutipes), Acai berry (Euterpe oleracea Mart), and Suaeda japonica. It is an antioxidant, anticancer, and HIV reverse transcriptase. It has been reported to exhibit various physiological activities such as inhibition and strong bitter taste masking effect.

On the other hand, allergy is a hypersensitivity reaction of the immune system caused by foreign antigens (allergens) in a harmless environment. Symptoms such as itching, fever, and pain, which are the main symptoms of asthma, atopic dermatitis, and allergic rhinitis, appear through allergic reactions, and these symptoms are caused by excessive activation of mast cells.

Mast cells play a critical role in the initiation of allergic reactions, and activation of mast cells is closely related to specific immunoglobulin E (IgE) antibodies as typical activators. IgE binds to Fcε receptor I (Fc epsilon receptor I, FcεRI) expressed on the surface of mast cells. Cross-linking of FcεRI with antigens on mast cells triggers intracellular signaling cascades, resulting in Lyn, Syk, protein kinase C (PKC), and mitogen activated protein kinase. Downstream mediators such as MAPK) are activated, increasing the concentration of Ca2+ (calcium ions) in the cell. In addition, aggregation of IgE-FcεRI induces exocytosis of preformed granules including histamine, chemokine, cytokine and serine protease, and leukotriene and prostaglandin Stimulates the new synthesis of These inflammatory mediators cause a variety of pathophysiological symptoms in allergic reactions, including contraction of airway smooth muscle, secretion of mucus and recruitment of other inflammatory cells. Thus, the regulation of inflammatory mediator production is important for controlling allergic reactions.

Rat basophilic leukemia (RBL)-2H3 cells are rodent mucosal mast cells that secrete inflammatory mediators. The RBL-2H3 cell line does not completely represent primary mast cells and basophils, but the RBL-2H3 cell line shares the characteristics of the two cells and can be simply subcultured. The main characteristic of mast cells is the secretion of histamine. Histamine is involved in allergic and inflammatory reactions, and plays an important role in mediating interactions between different cells. In addition, histamine is secreted by RBL-2H3 cells. Therefore, RBL-2H3 cells have generally been used for in vitro studies of allergic reactions. Phobol myristate acetate (PMA), an activating factor of PKC, and A23187, a calcium ionophore, increase the intracellular calcium ion concentration and induce activation of the signaling pathway in RBL-2H3 cells. do.

Under various reports of these allergy-related mechanisms, many studies are being conducted to develop more effective antiallergic substances.

Korean Patent Publication No. 10-2004-0087223 Korean Patent Publication No. 10-2018-0052226

The inventors of the present invention recognize the side effects of conventional antihistamines such as drowsiness, dizziness, vomiting, diarrhea, indigestion, excitement, etc. when taking it, and while studying on substances that alleviate these side effects, beloutine from mast cells It was confirmed that the release of histamine and β-hexosaminidase was inhibited, thereby exhibiting an effect of inhibiting degranulation.

Accordingly, it is an object of the present invention to provide a pharmaceutical composition for preventing or treating allergic diseases, including velutin.

Another object of the present invention is to provide a food composition for preventing or improving allergic diseases, including beloutine.

Another object of the present invention is to provide a cosmetic composition for preventing or improving allergic diseases, including beloutine.

In order to achieve the object of the present invention, the present invention provides a pharmaceutical composition for preventing or treating allergic diseases, including velutin.

In addition, the present invention provides a food composition for preventing or improving allergic diseases, including beloutine.

In addition, the present invention provides a cosmetic composition for preventing or improving allergic diseases, including beloutine.

In one embodiment of the present invention, the beloutine may have a therapeutic concentration of 1 to 200 μg/ml.

In one embodiment of the present invention, the allergic disease may be an allergic inflammatory disease.

In one embodiment of the present invention, the allergic disease may be one selected from the group consisting of asthma, hay fever (allergic rhinitis), and atopic eczema caused by an immediate allergic reaction.

In one embodiment of the present invention, the composition may inhibit degranulation of mast cells.

In one embodiment of the present invention, the composition may inhibit the release of β-hexosaminidase or histamine.

According to the present invention, the composition containing beloutin inhibits the release of β-hexosaminidase and histamine from mast cells to inhibit degranulation, such as drowsiness, dizziness, vomiting, diarrhea, indigestion, excitement, etc. It has been shown that the side effects of antihistamines can be alleviated. Therefore, the composition containing beloutine of the present invention can be usefully used as an antihistamine composition with improved side effects in the field of cosmetics or pharmaceuticals requiring an antiallergic composition.

Figure 1 shows the structure of the beloutin of the present invention and the similar compounds genkwanin (Genkwanin) and luteolin (luteolin).
Figure 2 shows the excellent β-hexosaminidase and histamine release inhibitory effect upon beloutine treatment of the present invention.
3 shows the ¹ H-NMR spectrum of the prepared beloutin.
Figure 4 shows the ¹ H-NMR spectrum of the prepared genkwanin.

The present invention provides a pharmaceutical composition for preventing or treating allergic diseases, including velutin.

In the present invention, the beloutine may have a therapeutic concentration of 1 to 200 μg/ml, preferably 50 to 150 μg/ml, and most preferably 100 μg/ml, but is not limited thereto.

In one embodiment of the present invention, the allergic disease may be an allergic inflammatory disease, preferably due to an immediate allergic reaction, and more preferably, asthma and hay fever (allergic rhinitis) due to an immediate allergic reaction. ) And atopic eczema may be one selected from the group consisting of.

The term allergic disease as used in the present invention refers to a disease caused by allergies. Allergy refers to an antigen-antibody reaction that occurs when a certain kind of substance (antigen or allergen) enters an individual, an antibody is made against it, and then the same substance, the antigen, enters the body. The main diseases are bronchial asthma, allergic rhinitis, urticaria, abruption, drug allergy, serum diseases, etc., and various types of diseases are indicated depending on the type of allergen or tissue causing an allergic reaction. In addition, in the present invention, the allergic inflammatory disease refers to an inflammatory disease induced by the allergic reaction.

The term immediate-type allergic reaction used in the present invention refers to type Ⅰ of four types of allergic reactions of type Ⅰ, type Ⅱ, type Ⅲ, and type Ⅳ. In type I reaction, when IgE is bound to the surface of mast cells or basophilic cells and specific antigens are bound to this IgE antibody, granule prolapse (degranulation) occurs in mast cells or basophilic cells, histamine, serotonin. , It refers to a phenomenon in which chemical substances such as bradykinin are released, resulting in increased vascular permeability and promotion of mucus secretion. Symptoms included in this include anaphylaxis, bronchial asthma, allergic rhinitis, and hives.

The composition of the present invention can inhibit degranulation of mast cells and inhibit β-hexosaminidase or histamine release. When Mast cells are activated, they cause hypersensitivity reactions (allergies), and these hypersensitivity reactions cause diseases such as atopy and asthma. Mast cells have numerous granules in cells, and the granules contain a number of inflammatory substances such as histamine and TNF-α. When mast cells are activated, these granules fuse with the cell membrane, and substances in the granules are secreted to the outside of the cells, causing an irritable inflammatory reaction. This process is called degranulation. β-hexosaminidase is a substance that is secreted simultaneously with histamine when mast cell degranulation occurs, and is widely used as an index to determine the degree of mast cell degranulation.

The term "prevention" used in the present invention means any action that suppresses or delays the onset of an allergic disease by administration of the pharmaceutical composition according to the present invention.

The term "treatment" used in the present invention refers to any action in which symptoms caused by an allergic disease are improved or advantageously changed by administration of the pharmaceutical composition according to the present invention.

The pharmaceutical composition according to the present invention includes beloutine as an active ingredient, and may include a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier is commonly used in formulation, and includes, but is limited to, saline, sterile water, Ringer's solution, buffered saline, cyclodextrin, dextrose solution, maltodextrin solution, glycerol, ethanol, liposome, and the like. It is not, and other conventional additives such as antioxidants and buffers may be further included if necessary. In addition, diluents, dispersants, surfactants, binders, lubricants, and the like may be additionally added to form an injectable formulation, pill, capsule, granule, or tablet such as an aqueous solution, suspension, or emulsion. Regarding suitable pharmaceutically acceptable carriers and formulations, it can be preferably formulated according to each component using a method disclosed in Remington's literature. The pharmaceutical composition of the present invention is not particularly limited in its formulation, but may be formulated as an injection or oral ingestion.

The pharmaceutical composition of the present invention can be administered orally or parenterally according to a desired method (for example, intravenously or subcutaneously), and the dosage is the patient's condition and weight, the degree of disease, the drug form, It depends on the route and time of administration, but may be appropriately selected by those skilled in the art.

The composition according to the present invention is administered in a pharmaceutically effective amount. In the present invention, “a pharmaceutically effective amount” means an amount sufficient to treat a disease at a reasonable benefit/risk ratio applicable to medical treatment, and the effective dose level is the type, severity, and activity of the drug of the patient. , Sensitivity to drugs, time of administration, route of administration and rate of excretion, duration of treatment, factors including drugs used concurrently, and other factors well known in the medical field. The composition according to the present invention may be administered as an individual therapeutic agent or administered in combination with other therapeutic agents, may be administered sequentially or simultaneously with a conventional therapeutic agent, and may be administered single or multiple. It is important to administer an amount capable of obtaining the maximum effect in a minimum amount without side effects in consideration of all the above factors, and this can be easily determined by a person skilled in the art.

Specifically, the effective amount of the composition according to the present invention may vary depending on the age, sex, and weight of the patient, and may increase or decrease according to the route of administration, the severity of the disease, sex, weight, age, and the like.

The term "pharmaceutically acceptable salts thereof" used in the present invention can be prepared by a method conventional in the art, for example, hydrochloric acid, hydrogen bromide, sulfuric acid, sodium hydrogen sulfate, phosphoric acid, carbonic acid Salts with inorganic acids such as formic acid, acetic acid, oxalic acid, benzoic acid, citric acid, tartaric acid, gluconic acid, gastisic acid, fumaric acid, lactobionic acid, salicylic acid, or organic acid such as acetylsalicylic acid (aspirin) Forms salts of these acids that are scientifically acceptable, or reacts with alkali metal ions such as sodium and potassium to form their metal salts, or reacts with ammonium ions to form another form of pharmaceutically acceptable salts thereof. Means to form.

In another aspect, the present invention provides a food composition for preventing or improving allergic diseases, including beloutine.

The food composition is a concept including a health functional food.

As used herein, the term "improvement" refers to any action that at least reduces the severity of a parameter related to the condition being treated, for example, symptoms.

In the food composition of the present invention, beloutine may be added to food as it is or may be used together with other foods or food ingredients, and may be appropriately used according to a conventional method. The amount of beloutin mixed may be appropriately determined depending on the purpose of use (for prevention or improvement). In general, in the manufacture of food or beverage, the composition of the present invention is added in an amount of 15% by weight or less, preferably 10% by weight or less based on the raw material. However, in the case of long-term intake for the purpose of health and hygiene or for health control purposes, the amount may be less than the above range.

The health functional food composition of the present invention is not particularly limited in other ingredients other than containing the above ingredients as an essential ingredient in the indicated ratio, and may contain various flavoring agents or natural carbohydrates, etc. as additional ingredients as in ordinary beverages. . Examples of the above-described natural carbohydrates include monosaccharides such as glucose, fructose, and the like; Disaccharides such as maltose, sucrose, and the like; And polysaccharides, for example, common sugars such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those described above, natural flavoring agents (taumatin, stevia extract (for example, rebaudioside A, glycyrrhizin, etc.)) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. The proportion of the natural carbohydrate can be appropriately determined by the choice of a person skilled in the art.

In addition to the above, the health functional food composition of the present invention includes various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and heavy weight agents (cheese, chocolate, etc.), pectic acid and salts thereof, Alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonates used in carbonated beverages, and the like may be contained. These components may be used independently or in combination. The proportion of these additives can also be appropriately selected by a person skilled in the art.

In another aspect, the present invention provides a cosmetic composition for preventing or improving allergic diseases, including beloutine.

In the present invention, the cosmetic composition is a formulation selected from the group consisting of solutions, suspensions, emulsions, pastes, gels, creams, powders, soaps, surfactant-containing cleansing, oils, powder foundations, emulsion foundations, wax foundations, and sprays. May have, but is not limited thereto.

The cosmetic composition of the present invention may include components commonly used in cosmetic compositions, and may include conventional adjuvants such as antioxidants, stabilizers, solubilizers, vitamins, pigments, and fragrances, and carriers. .

In addition, the composition of the present invention may be used by mixing an organic sunscreen. The organic sunscreens include glycerylpava, dromethrizoltrisiloxane, dromethrizol, digaloyltrioleate, disodiumphenyldibenzimidazoletetrasulfonate, diethylhexylbutamidotriazone, diethylamino Hydroxybenzoylhexylbenzoate, DieA-methoxycinnamate, mixture of Lawson and dihydroxyacetone, methylenebis-benzotriazolyltetramethylbutylphenol, 4-methylbenzylidene camphor, menthylanthranylate, benzophenone -3 (oxybenzone), benzophenone-4, benzophenone-8 (dioxyfebenzone), butylmethoxydibenzoylmethane, bisethylhexyloxyphenolmethoxyphenyltriazine, sinoxate, ethyldihydroxypropylpaba, Octocrylene, ethylhexyldimethylpaba, ethylhexylmethoxycinnamate, ethylhexylsalicylate, ethylhexyltriazone, isoamyl-p-methoxycinnamate, polysilicon-15 (dimethicodiethylbenzalmalonate) , Terephthallylidene dicamphor sulfonic acid and salts thereof, TEA-salicylate, and one or more selected from the group consisting of aminobenzoic acid (pava) may be used.

Hereinafter, the present invention will be described in more detail through examples. However, the following examples are for illustrating the present invention, and the scope of the present invention is not limited thereto.

<Example 1: Synthesis of beloutine and genkwannin>

Among the three flavone derivatives, belutin and genkwannin were chemically synthesized, and luteolin was purchased from ChemOne, Korea.

Belutin and Genkwanin Preparation: Prepared from 2',4',6'-trihydroxyacetophenone through four sequential methylation, aldol condensation, cyclization, and deprotection reactions (Fig. 3 and 4).

Preparation of Belloutine: ¹ H NMR (DMSO-d ₆ , 400 MHz), δ : 12.97 (s, 1H), 10.00 (s, 1H), 7.61-7.59 (m, 2H, J = 4.62 Hz), 6.96-6.93 (m, 2H, J = 9.96 Hz), 6.80 (d, 1H, J = 2.24 Hz), 6.38-6.37 (d, 1H, J = 2.20 Hz), 3.93 (s, 3H), 3.91 (s, 3H) .

Preparation of Genkwanin: ¹ H NMR (DMSO-d ₆ , 400 MHz), δ : 12.96 (s, 1H), 7.96 (d, 2H, J = 8.84 Hz), 6.93 (d, 2H, J = 8.84 Hz) , 6.84 (s, 1H), 6.77 (d, 1H, J = 2.25 Hz), 6.37 (d, 1H, J = 2.25 Hz), 3.87 (s, 1H)

<Example 2: Preparation for experiment>

RBL-2H3 cells were in DMEM medium (Dulbecco's modified Eagle's medium, Hyclone Logan, UT, USA) supplemented with 10% heat inactivated fetal bovine serum (FBS), 1% penicillin and 1% streptomycin (Hyclone). Cultured.

<Example 3: Confirmation of the effect of beloutin on reducing the release of β-hexosaminidase>

RBL-2H3 (mast cells) cells were seeded in a 24-well plate using DMEM media containing 10% FBS and cultured for 24 hours. Then, remove the culture medium and add beloutin to a concentration of 2-100 μg/ml in a siraganian buffer (NaCl 119mM, KCl 5mM, Glucose 5.6mM, PIPES 25mM, MgCl ₂ , 0.4mM, CaCl ₂ 1mM, BSA 0.1%, pH 7.2) was diluted to the cells and treated for 1 hour. In order to compare the effect of beloutin, genkwanin and luteolin were treated at a concentration of 100 μg/ml as controls. Then, the solution was removed from the cells, washed twice using PIPES, and treated with 50 nM PMA (phorbol myristate acetate) and 1 μM A23187 (calcium ionophore) for 30 minutes to induce an immune response. Then, take 20 μL of the supernatant and transfer it to a 96-well plate, and dilute the substrate 4-p-nitrophenyl-N-acetyl-β-D-glucosaminide in 0.05 M sodium citrate buffer (pH 4.5) to a concentration of 2 mM. Mix with 80 μL and react at 37 degrees for 30 minutes. Thereafter, 200 μL of a stop buffer (0.1 M NaHCO ₃ , pH 10) was added to terminate the reaction, and absorbance was measured at 405 nm.

As a result, as shown in FIG. 2, the release of β-hexosaminidase was significantly increased when degranulation was activated compared to normal mast cells (-) that were not treated with anything, and the release of β-hexosaminidase was decreased as the concentration of beloutine treatment increased. Confirmed. On the other hand, it was confirmed that genkwanin and luteolin showed lower effects than beloutin even when treated at the same concentration of 100 μg/ml.

<Example 4: Confirmation of the effect of beloutin on reducing histamine release>

RBL-2H3 (mast cells) cells were seeded in a 24-well plate using DMEM media containing 10% FBS and cultured for 24 hours. Then, remove the culture medium and add beloutin to a concentration of 2-100 μg/ml in a siraganian buffer (NaCl 119mM, KCl 5mM, Glucose 5.6mM, PIPES 25mM, MgCl ₂ , 0.4mM, CaCl ₂ 1mM, BSA 0.1%, pH 7.2) was diluted to the cells and treated for 1 hour. In order to compare the effect of beloutin, genkwanin and luteolin were treated at a concentration of 100 μg/ml as controls. Then, the solution was removed from the cells, washed twice using PIPES, and treated with 50 nM PMA (phorbol myristate acetate) and 1 μM A23187 (calcium ionophore) for 30 minutes to induce an immune response. Then, a 1 mL supernatant was taken, mixed with 2 mL of 1 N·NaOH, and 0.1 mL of 1% OPT (o-phtha-laldehyde), and reacted at room temperature for 5 minutes. Then, 0.1 mL 3 N·HCl was added to terminate the reaction, and excitation at 360 nm and fluorescence intensity emitted at 450 nm were measured.

As a result, as shown in FIG. 2, it was confirmed that histamine release was significantly increased when degranulation was activated compared to normal cells (-) that were not treated with anything, and histamine release was decreased as the beloutine concentration increased. On the other hand, it was confirmed that genkwanin and luteolin showed lower effects than beloutin even when treated at the same concentration of 100 μg/ml.

Claims (8)

A pharmaceutical composition for preventing or treating allergic diseases, including velutin.
The pharmaceutical composition of claim 1, wherein the beloutine is at a therapeutic concentration of 1 to 200 μg/ml.
The pharmaceutical composition according to claim 1, wherein the allergic disease is an allergic inflammatory disease.
The pharmaceutical composition according to claim 1, wherein the allergic disease is one selected from the group consisting of asthma, hay fever (allergic rhinitis), and atopic eczema caused by immediate allergic reaction.
The pharmaceutical composition of claim 1, wherein the composition inhibits degranulation of mast cells.
The pharmaceutical composition of claim 1, wherein the composition inhibits the release of β-hexosaminidase or histamine.
Food composition for preventing or improving allergic diseases, including beloutine.
A cosmetic composition for preventing or improving allergic diseases, including beloutine.

Images