KR101780262B1 - Composition comprising 4'-O-methylalpinum isoflavone for preventing and treating cardiovascular disease - Google Patents

Abstract

The present invention relates to a composition for preventing or treating cardiovascular diseases comprising 4-O-methylalpine isoflavone. The 4-O-methylalpine isoflavone of the present invention blocks a movement of a monocyte by a cholesterol oxide, and is excellent in an effect of inhibiting expression of CD137, CD105, and CD166, which are marker molecules related to atherosclerosis, thereby being effectively used for preventing or treating cardiovascular diseases including the atherosclerosis and related diseases.

Description

A composition for prevention or treatment of cardiovascular diseases comprising 4'-O-methylalpine isoflavone, which comprises 4'-O-methylalpinum isoflavone for treating and treating cardiovascular disease,

The present invention relates to a composition for preventing or treating cardiovascular diseases comprising 4'-O-methylalpine isoflavone.

Atherosclerosis, also called atherosclerosis, can occur in the coronary arteries, the aorta, and the peripheral artery, and is a problem in that it occurs in the inner wall of the vessel and gradually narrows the blood vessel. The main causes are hyperlipidemia, hypertension, diabetes, smoking, obesity, and periodic observation of my condition. Vascular anomalies caused by atherosclerosis can cause other diseases depending on the site of onset. If atherosclerosis occurs in the cerebral blood vessels, atherosclerosis in the cerebral infarction (cerebral infarction, cerebral hemorrhage) and cardiovascular (coronary artery) causes angina pectoris, myocardial infarction, and atherosclerosis in the limb blood vessels. These cardiovascular diseases are the main cause of death in Korea after malignant neoplasm (cancer).

Atherosclerosis is characterized by atheroma, atherosclerotic plaque or atherosclerotic plaque formation due to intravascular injury, and the pathogenesis of cholesterol is relatively well understood. The low density lipoprotein cholesterol present in the blood accumulates in the blood vessel walls and the cholesterol is oxidized in the blood vessels. Oxidized cholesterol induces inflammation and, as a result, monocytes migrate to the vessel wall. Monocytes are activated by cholesterol oxides in the blood vessel wall and amplify the inflammation. As a result of this series of reactions, the monocyte is continuously moved to the inner wall of the blood vessel and the atherosclerosis proceeds, and the movement of such monocytes plays a pivotal role in the development and development of atherosclerosis. Therefore, a method for inhibiting the migration of mononuclear cells to the inner wall of blood vessels is proposed as a next-generation therapy for atherosclerosis and cardiovascular diseases.

Monocytes that are present in atherosclerotic plaque promote atherosclerosis by expressing markers such as CD137 and CD105. In particular, CD137 (4-1BB) is highly expressed in monocytes existing in atherosclerotic plaques and inhibited the action of CD137 is proposed as a treatment method for atherosclerosis. In addition, CD105 (endoglin) has been reported to be involved in the destabilization of atherosclerotic plaque as a TGF-β receptor. In addition, CD166 (Alcam) is specifically expressed in atherosclerotic tissue as a marker of atherosclerosis.

27-Hydroxycholesterol (27OHChol) is the highest concentration of cholesterol oxides found in atherosclerotic plaque. 27OHChol promotes atherosclerosis by activating macrophages and by increasing the migration of monocytes. And, 27OHChol increases the expression of CD137, CD105 and CD166 in monocytes.

Isoflavone is similar to estrogen, a female hormone, and is known as a phytoestrogen because it induces estrogen secretion. It is known that there are more than 1000 different isoflavones in the plant system. Especially soybean plants, and it has recently been found that isoflavones can provide potential alternative therapies for hormone-dependent diseases including cancer, menopausal syndrome and osteoporosis.

 In studying natural products and compounds isolated therefrom in order to develop drugs for the prevention or treatment of cardiovascular diseases, the present inventors have found that 4'-O-methylalpinum isoflavone is a compound of 27 - inhibiting the movement of monocyte by hydroxycholesterol and inhibiting the expression of CD137, CD105 and CD166, thereby inhibiting cardiovascular diseases including atherosclerosis.

It is an object of the present invention to provide a composition for preventing or treating cardiovascular diseases comprising 4'-O-methylalpine isoflavone as an active ingredient.

In order to achieve the above object, the present invention provides a pharmaceutical composition for preventing or treating cardiovascular diseases comprising 4'-O-methylalpinum isoflavone as an active ingredient.

[Chemical Formula 1]

The present invention also provides a food composition for preventing or ameliorating cardiovascular diseases comprising 4'-O-methylalpine isoflavone represented by the above formula (1) as an active ingredient.

The 4'-O-methylalpine isoflavone of the present invention blocks the movement of monocyte by the cholesterol oxide and is excellent in the effect of inhibiting the expression of CD137, CD105 and CD166, the marker molecules related to atherosclerosis, And may be useful for the prevention or treatment of cardiovascular diseases and related diseases.

Brief Description of the Drawings Figure 1 is a schematic representation of a process for inhibiting the action of 27-hydroxycholesterol in 4'-O-methylalpine isoflavone.
FIG. 2 is a graph showing the results of cell migration analysis of a mononuclear cell line treated with 4'-O-methylalpine isoflavone.
FIG. 3 is a graph showing the results of flow cytometry of a mononuclear cell line treated with 4'-O-methylalpine isoflavone.

The present invention provides a composition for preventing or treating cardiovascular diseases comprising 4'-O-methylalpinum isoflavone as an active ingredient.

The composition comprises a pharmaceutical composition and a food composition.

Hereinafter, the present invention will be described in detail.

The 5-hydroxy-7- (4-methoxyphenyl) -2,2-dimethylpyrano [3,2-g] chromen- -methoxyphenyl) -2,2-dimethylpyrano [3,2- g] chromen-6-one) (C 21 H 18 O 5) in the following, can be represented by the formula (1).

[Chemical Formula 1]

The compound can be used in the form of a pharmaceutically acceptable salt, and includes all salts, hydrates and solvates produced by conventional methods.

As salts, acid addition salts formed by pharmaceutically acceptable free acids are useful. The acid addition salt is prepared in a conventional manner, for example, by dissolving the compound in an excess amount of an aqueous acid solution and precipitating the salt using a water-miscible organic solvent such as methanol, ethanol, acetone or acetonitrile. The same molar amount of the compound and the acid or alcohol (e.g., glycol monomethyl ether) in water may be heated and then the mixture may be evaporated to dryness, or the precipitated salt may be subjected to suction filtration. As the free acid, organic acids and inorganic acids can be used. As the inorganic acids, hydrochloric acid, phosphoric acid, sulfuric acid, nitric acid, tartaric acid and the like can be used. Examples of the organic acids include methanesulfonic acid, p- toluenesulfonic acid, acetic acid, trifluoroacetic acid, maleic acid, succinic acid, oxalic acid, benzoic acid, tartaric acid, fumaric acid, mandelic acid, propionic acid, citric acid, lactic acid, glycollic acid, gluconic acid, galacturonic acid, glutamic acid, glutaric acid, glucuronic acid, aspartic acid, ascorbic acid, carbonic acid, vanillic acid and hydroiodic acid may be used. It is not limited.

In addition, bases can be used to make pharmaceutically acceptable metal salts. The alkali metal or alkaline earth metal salt is obtained, for example, by dissolving the compound in an excess of an alkali metal hydroxide or an alkaline earth metal hydroxide solution, filtering the non-soluble compound salt, and evaporating and drying the filtrate. At this time, as the metal salt, it is preferable to produce sodium, potassium or calcium salt particularly, but not limited thereto. The corresponding silver salt can also be obtained by reacting an alkali metal or alkaline earth metal salt with a suitable silver salt (e.g., silver nitrate).

Pharmaceutically acceptable salts of such compounds include, unless otherwise indicated, salts of acidic or basic groups that may be present in the compounds. For example, pharmaceutically acceptable salts may include the sodium, calcium and potassium salts of the hydroxy group, and the other pharmaceutically acceptable salts of the amino group include hydrobromide, sulfate, hydrogen sulfate, phosphate, hydrogen phosphate, (Mesylate) and p -toluenesulfonate (tosylate) salts and the like, and a method for producing a salt known in the art ≪ / RTI >

Isoflavone is a soy protein that functions similar to estrogen, a female hormone. It is also called 'phytoestrogen', and is found in soybeans.

The 4'-O-methylalpine isoflavone of the present invention is a kind of isoflavone, which can be synthesized by an organic synthesis method or can be used without limitation such as those commercially available. Preferably, 4'-O-methylalpine isoflavone is obtained from a plant such as Cudrania tricuspidata Can be separated by conventional extraction and fractionation methods.

When the 4'-O-methylalpine isoflavone of the present invention is isolated from a plant, water, C ₁ -C ₄ alcohol or a mixed solvent thereof may be used as the extraction and fraction solvent, preferably methanol or ethanol Can be used.

The cardiovascular disease may be stroke, angina, heart failure, myocardial infarction, atherosclerosis or ischemic heart disease, preferably atherosclerosis.

The atherosclerosis is mainly caused by cholesterol, and the endothelial layer is cracked or thinned due to physical or biological damage of arterial endothelial cells, so that LDL cholesterol in the blood penetrates into the gap of the inner membrane and penetrates into the lower endothelium. Monocyte) enters the tissue, and the monocytes that enter the tissue play the role of macrophages that eat fat. As a result of this series of reactions, the mononuclear cells continuously move to the inner wall of the vessel, gradually become narrower vessels, .

The 4'-O-methylalpine isoflavone of the present invention can be used for prevention or treatment of cardiovascular diseases including atherosclerosis by inhibiting the action of cholesterol oxide.

The cholesterol oxide may preferably be 27-hydroxycholesterol, and its inhibition process is schematically shown in Fig.

As shown in Fig. 1, 4'-O-methylalpine isoflavone blocks the migration of mononuclear cells by 27-hydroxycholesterol, and inhibits the expression of CD137, CD105 and CD166, the marker molecules involved in atherosclerosis.

The composition of the present invention may contain, in addition to the 4'-O - methylalpine isoflavone of the present invention, one or more known active ingredients having an effect of preventing or treating cardiovascular diseases.

The compositions of the present invention may further comprise suitable carriers, excipients and diluents conventionally used in the manufacture of pharmaceutical compositions. In addition, it can be formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, oral formulations such as syrups and aerosols, external preparations, suppositories and sterilized injection solutions according to a conventional method. Suitable formulations known in the art are preferably those as disclosed in Remington ' s Pharmaceutical Science, recently, Mack Publishing Company, Easton PA. Examples of carriers, excipients and diluents which may be included include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, Cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. When the composition is formulated, it is prepared using a diluent such as a filler, an extender, a binder, a wetting agent, a disintegrant, a surfactant, or an excipient usually used. Solid formulations for oral administration include tablets, pills, powders, granules, capsules and the like, which may contain at least one excipient such as starch, calcium carbonate, sucrose, lactose, Gelatin and the like. In addition to simple excipients, lubricants such as magnesium stearate and talc may also be used. Examples of the liquid preparation for oral use include suspensions, solutions, emulsions, and syrups. In addition to water and liquid paraffin, simple diluents commonly used, various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included . Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Examples of the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. Examples of the suppository base include witepsol, macrogol, tween 61, cacao butter, laurin, glycerogelatin and the like.

The term "administering" as used herein is intended to provide any individual of the invention with a composition of the invention in any suitable manner.

The preferred dosage of the pharmaceutical composition of the present invention varies depending on the condition and the weight of the individual, the degree of disease, the type of drug, the route of administration and the period of time, but can be appropriately selected by those skilled in the art. For the desired effect, the 4'-O-methylalpine isoflavone of the present invention may be administered in an amount of 1 mg / kg to 1000 mg / kg per day, and may be administered once a day or divided into several times You may.

The pharmaceutical composition of the present invention may be administered to a subject in various routes. All modes of administration may be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intra-uterine dural or intracerebral injection.

The composition of the present invention can be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy, and biological response modifiers for the prevention and treatment of cardiovascular diseases.

In the present invention, the term "food" refers to a food having a biological control function such as prevention and improvement of disease, bio-defense, immunity, recovery after disease, suppression of aging and the like.

The composition of the present invention may be added to food for the purpose of preventing or improving cardiovascular diseases. When 4'-O-methylalpine isoflavone of the present invention is used as a food additive, the 4'-O-methylalpine isoflavone may be directly added or used together with other food or food ingredients, Can be appropriately used. The amount of the active ingredient to be mixed can be suitably determined according to the intended use (prevention, health or therapeutic treatment). Generally, the 4'-O-methylalpine isoflavone of the present invention is added in an amount of not more than 15% by weight, preferably not more than 10% by weight based on the raw material. However, in the case of long-term intake for the purpose of health and hygiene or for the purpose of controlling health, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount exceeding the above range.

There is no particular limitation on the kind of the food. Examples of the foods to which the above substances can be added include dairy products including meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen noodles, gums, ice cream, various soups, drinks, tea, Alcoholic beverages, and vitamin complexes, and includes foods in a conventional sense.

The beverage composition of the present invention may contain, as an additional ingredient, various flavors or natural carbohydrates such as ordinary beverages. The natural carbohydrates may be monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, natural sweeteners such as dextrin and cyclodextrin, synthetic sweeteners such as saccharine and aspartame, and the like. The ratio of the natural carbohydrate is generally about 0.01 to 10 g, preferably about 0.01 to 0.1 g per 100 ml of the composition of the present invention.

In addition to the above, the composition of the present invention may further contain various nutrients, vitamins, electrolytes, flavors, colorants, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, A carbonating agent used in a carbonated beverage, and the like. In addition, the composition of the present invention may comprise flesh for the production of natural fruit juices, fruit juice drinks and vegetable drinks. These components may be used independently or in combination. Although the ratio of such additives is not critical, it is generally selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the composition of the present invention.

Hereinafter, preferred examples and formulations of the present invention will be described in order to facilitate understanding of the present invention. However, the following experimental examples and preparation examples are provided only for the purpose of easier understanding of the present invention, and the present invention is not limited thereto.

Experimental Example  1. Cell migration assay

THP-1 cells were purchased from American Type Culture Collection (ATCC), cultured according to the method recommended by ATCC, and 27-hydroxycholesterol (27OHChol) was purchased from Research Plus.

Then, Transwell Permeable Supports (Costar) was used to measure the migration of the cells. First, THP-1 cells were cultured in media (control), 27OHChol (2 μg / ml), 270Chol + 4'-O-methylalpine isoflavone 0.3125 μg / ml, 27OHChol + 4'- μg / ml, and 27OHChol + 4'-O-methylalpine isoflavone, respectively. After 48 hours of culture, the medium was separated and the separated medium was dispensed into a transwell chamber. THP-1 cells (5 x 10 ⁵ cells / 100 μL of 0.1% BSA) were dispensed into the upper chamber, and the upper chamber containing THP-1 cells was inserted into the transwell chamber with the medium. The cells were then incubated in a CO ₂ incubator for 2 hours and the chamber was separated.

The number of cells transferred to the transwell chamber was measured using a Vi-Cell cell counter. The results are shown in FIG.

As shown in FIG. 2, the medium of monocytes treated with 27OHChol increased monocyte migration, but monocyte migration was blocked in the medium of monocytes treated with 4'-O-methylalpine isoflavone and 27OHChol Respectively.

(*** P <0.001 vs. control group; # P <0.05 vs. 27 OHChol; ### P <0.001 vs. 27 OHChol)

Experimental Example  2. Flow cell  analysis

THP-1 cells were purchased from American Type Culture Collection (ATCC), cultured according to the method recommended by ATCC, and 27-hydroxycholesterol (27OHChol) was purchased from Research Plus.

Then, fluorescence labeled cells were measured using FACS canto II (BD) for flow cytometry analysis of the cells. First, THP-1 cells were cultured in media (control), 27OHChol (2 μg / ml), 270Chol + 4'-O-methylalpine isoflavone 0.3125 μg / ml, 27OHChol + 4'- μg / ml, and 27OHChol + 4'-O-methylalpine isoflavone 5 μg / ml, respectively, and cultured for 48 hours. After culturing, each cultured cell was harvested, washed with PBS, and washed cells were suspended in the primary antibody solution for CD105, CD137 and CD166 and left at 4 ° C for 2 hours. After 2 hours, the cells were washed with PBS and the secondary antibody against the primary antibody labeled with the fluorescent substance was treated and left at 4 DEG C for 1 hour. The cells were then washed with PBS, suspended in 500 μl of 1% paraformaldehyde and analyzed by flow cytometry. The results are shown in FIG.

As shown in FIG. 3, the expression of CD105, CD137 and CD166 was increased by about 5-fold in the monocytes stimulated with 27OHChol. However, when 4'-O-methylalpine isoflavone and 27OHChol were treated together, mononuclear cells CD105, CD137 And CD166 expression decreased in proportion to the concentration of 4'-O-methylalpine isoflavone.

This is because the 4'-O-methylalpine isoflavone of the present invention blocks the migration of mononuclear cells by 27-hydroxycholesterol and inhibits the expression of CD137, CD105 and CD166, which are the marker molecules involved in atherosclerosis, Can be used for the prevention or treatment of cardiovascular diseases and related diseases.

Hereinafter, a pharmaceutical composition for prevention or treatment of cardiovascular diseases comprising the 4'-O-methyl alpine isoflavone of the present invention as an active ingredient, and a preparation example of a food composition for preventing or improving is described, but the present invention is intended to be limited I would like to explain more specifically.

Formulation Example 1. Preparation of pharmaceutical preparations

1. Manufacturing of powder

4'-O-methylalpine isoflavone 20 mg

Lactose 100 mg

Talc 10 mg

The above components are mixed and filled in airtight bags to prepare powders.

2. Preparation of tablets

4'-O-methylalpine isoflavone 10 mg

Corn starch 100 mg

Lactose 100 mg

Magnesium stearate 2 mg

After mixing the above components, tablets are prepared by tableting according to the usual preparation method of tablets.

3. Preparation of capsules

4'-O-methylalpine isoflavone 10 mg

Crystalline cellulose 3 mg

Lactose 14.8 mg

Magnesium stearate 0.2 mg

The above components are mixed according to a conventional capsule preparation method and filled in gelatin capsules to prepare capsules.

4. Preparation of injections

4'-O-methylalpine isoflavone 10 mg

180 mg mannitol

Sterile sterilized water for injection 2974 mg

Na ₂ HPO ₄ 2H ₂ O 26 mg

(2 ml) per 1 ampoule in accordance with the usual injection preparation method.

5. Manufacture of liquids

4'-O-methylalpine isoflavone 20 mg

10 g per isomer

5 g mannitol

Purified water quantity

Each component was added and dissolved in purified water according to the usual liquid preparation method, and the lemon flavor was added in an appropriate amount. Then, the above components were mixed, and purified water was added thereto. The whole was added with purified water to adjust the total volume to 100 ml, And sterilized to prepare a liquid preparation.

Formulation Example 2. Preparation of food preparation

1. Manufacture of health food

4'-O-methylalpine isoflavone 100 mg

Vitamin mixture quantity

70 g of vitamin A acetate

Vitamin E 1.0 mg

Vitamin B1 0.13 mg

0.15 mg of vitamin B2

Vitamin B6 0.5 mg

0.2 g of vitamin B12

Vitamin C 10 mg

Biotin 10 g

Nicotinic acid amide 1.7 mg

Folate 50 g

Calcium pantothenate 0.5 mg

Mineral mixture quantity

1.75 mg of ferrous sulfate

0.82 mg of zinc oxide

Magnesium carbonate 25.3 mg

Potassium monophosphate 15 mg

Secondary calcium phosphate 55 mg

Potassium citrate 90 mg

Calcium carbonate 100 mg

Magnesium chloride 24.8 mg

Although the composition ratio of the above-mentioned vitamin and mineral mixture is comparatively mixed with a composition suitable for health food as a preferred embodiment, the compounding ratio may be arbitrarily modified, and the above ingredients are mixed according to a conventional method for producing healthy foods , Granules can be prepared and used in the manufacture of health food compositions according to conventional methods.

2. Health drink  Produce

4'-O-methylalpine isoflavone 100 mg

Vitamin C 15 g

Vitamin E (powder) 100 g

19.75 g of ferrous lactate

3.5 g of zinc oxide

Nicotinic acid amide 3.5 g

Vitamin A 0.2 g

Vitamin B1 0.25 g

Vitamin B2 0.3g

Water quantification

The above components were mixed according to a conventional health drink manufacturing method, and the mixture was stirred and heated at 85 DEG C for about 1 hour. The resulting solution was filtered and sterilized in a sterilized 2 L container, &Lt; / RTI &gt;

Although the compositional ratio is relatively mixed with a component suitable for a favorite drink, it is also possible to arbitrarily modify the compounding ratio according to the regional or national preference such as the demand class, the demanding country, and the use purpose.

Claims (10)

A pharmaceutical composition for preventing or treating cardiovascular diseases comprising 4'-O-methylalpinum isoflavone as an active ingredient. The pharmaceutical composition for preventing or treating cardiovascular diseases according to claim 1, wherein the 4'-O-methyl alpine isoflavone is represented by the following formula (1).
[Chemical Formula 1]

The pharmaceutical composition for preventing or treating cardiovascular diseases according to claim 1, wherein the 4'-O-methylalpine isoflavone is isolated from Cudrania tricuspidata fruit. The pharmaceutical composition according to claim 3, wherein the 4'-O-methylalpine isoflavone is separated by using water, a C ₁ -C ₄ alcohol or a mixed solvent thereof. Gt; The pharmaceutical composition for preventing or treating cardiovascular diseases according to claim 1, wherein the cardiovascular disease is at least one selected from the group consisting of stroke, angina pectoris, heart failure, myocardial infarction, atherosclerosis and ischemic heart disease. The pharmaceutical composition for preventing or treating cardiovascular diseases according to claim 5, wherein the cardiovascular disease is atherosclerosis. The pharmaceutical composition for preventing or treating cardiovascular diseases according to claim 1, wherein the 4'-O-methyl alpine isoflavone inhibits monocyte migration. The pharmaceutical composition according to claim 7, wherein the movement of the mononuclear cells is caused by cholesterol oxide. A food composition for prevention or improvement of cardiovascular diseases comprising 4'-O-methylalpinum isoflavone as an active ingredient. 10. The food composition according to claim 9, wherein the 4'-O-methylalpine isoflavone is represented by the following formula (1).
[Chemical Formula 1]

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