KR20220065116A - Composition Comprising the Extract of Senna Alata for Preventing or Inhibiting Vascular Aging - Google Patents

Abstract

The present invention relates to a food composition for vascular aging prevention or inhibition or cardiovascular disease prevention or amelioration containing a Senna alata extract as an active ingredient, a health functional food containing the food composition, and a pharmaceutical composition for cardiovascular disease prevention or treatment containing a Senna alata extract as an active ingredient. The Senna alata extract according to the present invention exhibits excellent vascular cell aging inhibition effects by inhibiting the activity of senescence associated beta galactosidase (SA-β-gal) and decreasing the expression of aging-related proteins (p16, p21, p53, and PAI-1). Accordingly, the composition containing the Senna alata extract can be used for vascular aging prevention or inhibition and can also be useful in manufacturing foods and drugs for cardiovascular disease prevention, amelioration, or treatment.

Description

Composition Comprising the Extract of Senna Alata for Preventing or Inhibiting Vascular Aging comprising Senna Alata extract

The present invention provides a food composition for preventing or inhibiting vascular aging, or preventing or improving cardiovascular disease, comprising the Senna alata extract as an active ingredient; Health functional food containing the food composition; And it relates to a pharmaceutical composition for the prevention or treatment of cardiovascular diseases comprising the Sena alata extract as an active ingredient.

Vascular endothelial cells are a single layer of cells that form the inner membrane of the vascular lumen, and play a major regulatory role in maintaining vascular homeostasis, such as vasodilation and contraction, proliferation and migration of vascular smooth muscle, and thrombus formation and dissolution. Dysfunction of vascular endothelial cells disrupts this balance, resulting in damage to the arterial wall and can be used as an early marker of atherosclerosis. Endothelial cell dysfunction can occur during the aging process.

In aged vascular endothelial cells, nitric oxide production, eNOS (endothelial nitric oxide synthase) expression, and prostacyclin production are reduced, indicating a remarkably low vascular contractility, and PAI-1 (type I plasminogen activator) The expression increases to promote thrombus formation, and increases the expression of cell adhesion proteins and chemokines such as Inter-Cellular Adhesion Molecule 1 (ICAM-1), Interleukin-1, Interleukin-8, and the inflammatory response increases, resulting in narrowing of blood vessels. will lose In fact, it is known that vascular endothelial cells of atherosclerotic lesion tissues have increased SA-β-Gal (senescence associated beta-galactosidase) activity, a well-known marker of cellular aging, compared to cells of normal tissues.

Because senescent cells do not divide smoothly, when senescent cells accumulate, damaged tissues cannot be repaired quickly, and they secrete enzymes or inflammatory cytokines that decompose surrounding tissues, thereby accelerating tissue damage. contributes to the pathogenesis of the disease; In other words, it is believed that by inhibiting or delaying the aging of vascular endothelial cells, it is possible to prevent and treat cardiovascular diseases resulting therefrom.

As such cellular senescence is considered to be an essential causative factor of aging, efforts have been made to develop a method for delaying cellular senescence. Among them, some studies have been reported to search for substances capable of regulating cellular aging and to utilize them for the prevention and treatment of diseases.

On the other hand, Senna alata ( Senna alata ) is a plant distributed in tropical and wet areas, and is known to exhibit various pharmacological activities in flowers, roots, leaves, and seeds. Specifically, antibacterial activity, anti-fungal activity, antiviral activity, etc. of Senna alata have been reported, and it has been reported that it is used in the treatment of wounds, skin infections, allergies, diarrhea, and the like. However, there is no case in which the Senna alata extract is used to inhibit cellular aging or to be applied to the prevention or treatment of cardiovascular diseases.

Under this background, the present inventors found that Sena alata extract inhibits senescence associated beta galactosidase (SA-β-gal) activity and has the effect of reducing the expression of senescence-associated proteins (p16, p21, p53 and PAI-1). By identifying the, it was confirmed that the Senna alata extract can be used as a food composition and pharmaceutical composition material for preventing or inhibiting vascular aging, or preventing, improving or treating diseases related to vascular aging, thereby completing the present invention. .

Republic of Korea Patent Registration No. 10-1522596 Republic of Korea Patent No. 10-1816601

An object of the present invention is to prevent or inhibit vascular aging, including Senna alata extract as an active ingredient; Or to provide a food composition for the prevention or improvement of cardiovascular disease.

Another object of the present invention is to provide a health functional food comprising the food composition.

Another object of the present invention is to provide a pharmaceutical composition for the prevention or treatment of cardiovascular diseases comprising the Sena alata extract as an active ingredient.

This will be described in detail as follows. Meanwhile, each description and embodiment disclosed in the present application may be applied to each other description and embodiment. That is, all combinations of the various elements disclosed in the present application fall within the scope of the present application. In addition, it cannot be seen that the scope of the present application is limited by the detailed description described below.

As an aspect for achieving the above object, the present invention provides a food composition for preventing or inhibiting vascular aging, or preventing or improving cardiovascular disease, comprising the Senna alata extract as an active ingredient; And it provides a health functional food comprising the food composition.

The term "extract" of the present invention refers to an extract obtained by the Senna alata extraction treatment, a diluted or concentrated liquid of the extract, a dried product obtained by drying the extract, a prepared or purified product of the extract, or a mixture thereof, such as an extract It includes extracts of all formulations that can be formed by themselves and using extracts.

The extract of the present invention may be extracted from the stem, root, leaf, flower, seed, outpost, or a combination thereof, of Sena alata, specifically, it may be extracted from the whole plant of Sena alata.

The method of extracting the extract of the present invention is not particularly limited, and may be extracted according to a method commonly used in the art. Non-limiting examples of the extraction method include hot water extraction method, cold extraction method, solvent extraction method, steam distillation method, elution method, compression method, ultrasonic extraction method, filtration method, reflux extraction method, etc. can be performed.

The extract of the present invention is extracted from Senna alata using an appropriate solvent, and may include, for example, a crude extract, a polar solvent-soluble extract, or a non-polar solvent-soluble extract. The type of extraction solvent used to prepare the extract is not particularly limited, and any solvent known in the art may be used. Non-limiting examples of the extraction solvent include water; lower alcohols having 1 to 4 carbon atoms, such as methanol, ethanol, propanol, isopropanol, butanol, propyl alcohol, and butyl alcohol; polyhydric alcohols such as glycerin, butylene glycol and propylene glycol; hydrocarbon solvents such as methyl acetate, ethyl acetate, acetone, benzene, hexane, diethyl ether, and dichloromethane; Or a mixture thereof may be used. Specifically, the extraction solvent may be methanol. In addition, it is possible to prepare a solvent extract by extracting one or more times using the solvent.

Specifically, in an embodiment of the present invention, the Senna alata extract is obtained by adding 100 to 300 ml of methanol to 20 to 50 g of dried and pulverized Senna alata, and 10 to 30 at 40 to 60° C. and 1200 to 1800 psi conditions. It may have been extracted using an automatic extractor for minutes. In addition, in another embodiment of the present invention, the Senna alata extract is dried and pulverized by adding 1 to 2 L of methanol to 100 to 300 g of Senna alata and using a sonicator for 2 to 4 days at 40 to 60 ° C. may have been extracted.

The extract of the present invention is used as it is, or concentrated or dried (eg, freeze-dried, Hot air drying, spray drying, etc.) can be used. In addition, the extract may be further subjected to a conventional fractionation process, and may be purified using a conventional purification method.

Specifically, in the present invention, Senna alata may be extracted with a methanol solvent, the extract is filtered, and the extract may be concentrated by evaporating the methanol, but is not limited thereto.

As used herein, the term “active ingredient” refers to a component capable of exhibiting the desired activity alone, or in combination with a carrier that has no activity by itself.

As used herein, the term "cardiovascular disease" refers to vascular diseases of the circulatory system, heart disease or cerebrovascular disease, and specifically, arteriosclerosis, hyperlipidemia, hypertension, angina, myocardial infarction, stroke, cardiac arrhythmias, It may be selected from the group consisting of heart failure, endocarditis, ischemic cerebrovascular disease, and ischemic heart disease, but is not limited thereto.

As used herein, the term “vascular disease of the circulatory system” refers to cases in which diseases occur in various organs of the large and small circulatory systems, including the heart, involved in blood circulation.

As used herein, the term “heart disease” refers to a disease related to the heart that is responsible for supplying blood to the human body, and refers to ischemic heart disease, cardiac arrhythmias, heart failure, and the like. The ischemic heart disease typically includes angina pectoris and myocardial infarction, and refers to an acute or chronic heart disorder that occurs because the blood supply to the myocardium is reduced or stopped as the coronary artery disease progresses. Other heart diseases include endocarditis, cardiac arrhythmias, or heart failure. In particular, heart failure refers to a state in which the heart cannot sufficiently rescue blood returning to the heart through the venous system.

As used herein, the term "cerebrovascular disease" refers to a disease that occurs suddenly due to abnormalities in cerebral blood vessels and causes brain dysfunction and collapses. It may be distinguished from ischemic cerebrovascular disease in which blood flow is impaired or blocked, but is not limited thereto.

In the present invention, the cardiovascular disease may be caused by vascular aging, and specifically, as the age increases, the endothelial cells constituting the blood vessels age and function abnormalities accumulate in the process.

In the present invention, the composition can inhibit the aging of vascular cells by reducing the activity of senescence associated beta galactosidase (SA-β-gal), which is a marker of aging, in senescent vascular cells.

In the present invention, the composition inhibits the aging of vascular cells by reducing the expression of senescence markers p16, p21, p53 and PAI-1 (plasminogen activator inhibitor-1) in aged vascular cells, and is capable of normalizing the function of vascular cells. can The p53, p16 and p21 are proteins that inhibit cell growth or cell division, and the PAI-1 protein promotes aging and thrombus formation by increasing inflammation.

As used herein, the term “prevention” refers to any action that inhibits or delays vascular aging or cardiovascular disease by ingesting the food composition of the present invention.

As used herein, the term “improvement” refers to any action that improves or benefits vascular aging or cardiovascular disease by ingesting the food composition of the present invention.

SA-β-gal, p16, p21, p53 and PAI-1 are known aging markers, and the correlation between vascular aging and cardiovascular disease is well known in the art ( J Atheroscler Thromb, 2020; 27: 47-59 and Experimental Gerontology , 2005; 40: 634-642). Therefore, the composition comprising the Sena alata extract according to the present invention as an active ingredient inhibits SA-β-gal activity and reduces the expression of senescence-related proteins (p16, p21, p53 and PAI-1), thereby improving vascular cells Since it has an anti-aging effect, it is evident that the composition comprising the Senna alata extract has a preventive or inhibitory effect on vascular aging and a prevention, improvement or treatment effect of cardiovascular disease associated therewith.

The food composition of the present invention prevents or inhibits vascular aging; Or, as long as it has the effect of preventing or improving cardiovascular disease, the Sena alata extract may be included in various amounts.

In addition to the Senna alata extract, the food composition of the present invention may further include any compound or natural extract known to have the activity and safety has already been verified in the art in order to enhance the convenience of taking or ingestion. there is.

Such compounds or extracts include compounds or extracts and pharmaceuticals listed in compendial documents such as pharmacopeias of each country ('Korean Pharmacopoeia' in Korea) and health functional food regulations of each country ('health functional food standards and specifications', announced by the Ministry of Food and Drug Safety in Korea). Each country's laws governing the manufacture and sale of compounds or extracts and health functional foods that have been approved for items in accordance with the laws of each country (the 'Pharmaceuticals Act' in Korea) that regulate manufacturing and sales ('Health Functional Food Act' in Korea) Depending on the functionalities recognized compounds or extracts may be included.

As used herein, the term "food" refers to meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages, There are vitamin complexes, nutritional supplements, functional foods, food additives, health functional foods, health foods, and health foods, and includes all foods in a normal sense.

The health food means a food having an active health maintenance or promotion effect compared to general food, and the health supplement food means a food for the purpose of health supplementation. In some cases, the terms health functional 　 food, health food, and health supplement food are preferred.

As used herein, the term "health functional food" is the same term as food for special health use (FoSHU), and it is a food with high medical and medical effects processed to efficiently exhibit bioregulatory functions in addition to nutrition supply. it means. Here, "function (sex)" means to obtain a useful effect for health purposes such as regulating nutrients or physiological action with respect to the structure and function of the human body. The food of the present invention can be manufactured by a method commonly used in the art, and during the preparation, raw materials and components commonly added in the art can be added. In addition, if the formulation of the above 　 food is also a formulation that is recognized as a food, it can be manufactured without limitation. The 　food 　 composition of the present invention can be prepared in various types of dosage forms, and unlike general drugs, 　 food as a raw material has the advantage of not having side effects that may occur during long-term administration of the drug, and has excellent portability, Food can be ingested as an adjuvant for preventing, suppressing, or improving vascular aging or cardiovascular disease.

Specifically, the health function 　 food is a food prepared by adding Senna alata extract to food materials such as beverages, teas, spices, gum, and confectionery, or manufactured in pills, tablets, capsules, powders, suspensions, etc., when ingested It means to bring a specific effect on health, but unlike general drugs, it has the advantage that there are no side effects that may occur when taking the drug for a long time because it is made from food.

The food composition of the present invention is very useful because it can be ingested on a daily basis, and thus the prevention, suppression or improvement effect of vascular aging or cardiovascular disease can be expected.

The food composition of the present invention may be prepared in any form, for example, beverages such as tea, juice, carbonated beverages, and ionic beverages, processed oils such as milk and yogurt, gums, rice cakes, Korean sweets, bread, It can be prepared into foods such as confectionery and noodles, tablets, capsules, pills, granules, liquids, powders, pieces, pastes, syrups, gels, jellies, and preparations such as bars. In addition, the food composition of the present invention may have any product classification in terms of legal and functional classification as long as it conforms to the enforcement laws at the time of manufacturing and distribution. For example, it is a health functional food according to the 'Health Functional Food Act' of Korea, or confectionery, tea, beverage, and special food according to each food type according to the Food Ordinance of the Korea 'Food Sanitation Act' (the 'Food Standards and Specifications' notified by the Ministry of Food and Drug Safety). It may be used food and the like.

In addition, the food composition of the present invention may contain food additives in addition to the active ingredient. Food additives can be generally understood as substances that are added and mixed or infiltrated into food in manufacturing, processing, or preserving food. These food additives are classified into sweeteners, flavor enhancers, preservatives, emulsifiers, flavorings, and the like in terms of use.

The sweetener is used to impart an appropriate sweetness to food, and may be natural or synthesized. For example, sweeteners such as neotame, lactitol, D-ribose, mannitol, D-maltitol, and sodium saccharin may be used.

The flavor enhancer is a food additive that enhances the taste or flavor of food, and both natural and synthetic ones may be used. For example, the flavor enhancer may include 5'-disodium guanylate, L-glutamic acid, glycine, betaine, and the like.

As the preservative, sodium metabisulfite, sulfurous anhydride, sorbic acid, benzoic acid, grapefruit seed extract, etc. may be used.

The emulsifier is a food additive that homogeneously mixes or maintains two or more immiscible phases such as water and oil, and sodium gluconate, glycerin fatty acid ester, lecithin, magnesium stearate, alginic acid, etc. may be used.

The flavoring is a food additive that gives a unique flavor to food or reinforces the original flavor of food lost during the manufacturing process. .

The food composition of the present invention may contain, in addition to the food additives described above, physiologically active substances or minerals known in the art for the purpose of supplementing and reinforcing functionality and nutrition and guaranteed stability as food additives. Examples of the physiologically active substances include catechins included in green tea, vitamins such as vitamin B1, vitamin C, vitamin E, and vitamin B12, tocopherol, dibenzoylthiamine, and the like, and minerals include calcium preparations such as calcium citrate, stearic acid Magnesium preparations, such as magnesium, iron preparations, such as iron citrate, chromium chloride, potassium iodide, selenium, germanium, vanadium, zinc, etc. are mentioned.

The food composition of the present invention may contain the above-mentioned food additives in an appropriate amount to achieve its purpose depending on the type of product, and with respect to other food additives that may be included in the food composition of the present invention, You can refer to the Food Additives Ordinance.

When the Sena alata extract is used as a food additive, the Senna alata extract may be added as it is or used with other foods or food ingredients, and may be appropriately used according to a conventional method. The mixing amount of the active ingredient may be suitably determined according to the purpose of its use.

As another aspect for achieving the above object, the present invention provides a pharmaceutical composition for the prevention or treatment of cardiovascular diseases comprising Senna alata extract as an active ingredient.

The terms "extract", "active ingredient", "cardiovascular disease" and "prevention" of the present invention are the same as described above.

As used herein, the term “treatment” refers to any action that improves, alleviates, or benefits cardiovascular disease by administering the pharmaceutical composition of the present invention to an individual.

In the present invention, the composition can inhibit the aging of vascular cells by reducing the activity of senescence associated beta galactosidase (SA-β-gal), which is a marker of aging, in senescent vascular cells.

In the present invention, the composition inhibits the aging of vascular cells by reducing the expression of senescence markers p16, p21, p53 and PAI-1 (plasminogen activator inhibitor-1) in aged vascular cells, and is capable of normalizing the function of vascular cells. can The p53, p16 and p21 are proteins that inhibit cell growth or cell division, and the PAI-1 protein promotes aging and thrombus formation by increasing inflammation.

SA-β-gal, p16, p21, p53 and PAI-1 are known as aging markers, and the correlation between vascular aging and cardiovascular disease is well known in the art. Therefore, the composition comprising the Sena alata extract according to the present invention as an active ingredient inhibits SA-β-gal activity and reduces the expression of senescence-related proteins (p16, p21, p53 and PAI-1), thereby improving vascular cells Since it has an anti-aging effect, it is evident that the composition comprising the Senna alata extract has a preventive or therapeutic effect on cardiovascular diseases.

The pharmaceutical composition of the present invention may further include a pharmaceutically acceptable carrier, excipient or diluent according to a conventional method. Pharmaceutically acceptable carriers are well known in the art depending on the route of administration or formulation, and specifically, reference may be made to the pharmacopoeia of each country including the 'Korean Pharmacopoeia'. Carriers, excipients and diluents that may be included in the composition of the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. In addition, carriers, excipients and diluents that may be included in the composition of the present invention may be non-natural carriers, but are not limited thereto.

The pharmaceutical composition of the present invention may be formulated in the form of oral dosage forms such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, external preparations, suppositories, or sterile injection solutions according to conventional methods. . Specifically, it can be prepared using a diluent or excipient such as a filler, extender, binder, wetting agent, disintegrant, surfactant, etc. usually used when formulating. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc., and such solid preparations include at least one excipient, for example, starch, calcium carbonate, sucrose, lactose, gelatin, etc. to the compound. It can be prepared by mixing. In addition to simple excipients, lubricants such as magnesium stearate and talc may also be used. Liquid formulations for oral administration include suspensions, internal solutions, emulsions, and syrups. In addition to commonly used simple diluents such as water and liquid paraffin, various excipients such as wetting agents, sweeteners, fragrances, and preservatives are included. can Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized formulations and suppositories. Non-aqueous solvents and suspending agents include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate. As the base of the suppository, Witepsol, Macrogol, Tween 61, cacao butter, laurin fat, glycerogelatin, etc. can be used. Specific formulations of pharmaceutical compositions are known in the art, and reference may be made to, for example, Remington's Pharmaceutical Sciences (19th ed., 1995). This document is considered a part of this specification.

The pharmaceutical composition of the present invention is administered in a pharmaceutically effective amount. The pharmaceutically effective amount means an amount sufficient to treat a disease with a reasonable benefit/risk ratio applicable to medical treatment and not to cause side effects, and the effective dose level is determined by the patient's health condition, disease type, severity, The activity of the drug, the sensitivity to the drug, the administration method, administration time, administration route and excretion rate, treatment period, factors including drugs used in combination or concurrently, and other factors well known in the medical field may be determined according to factors. Dosage and frequency do not limit the scope of the present invention in any way.

The pharmaceutical composition of the present invention may be administered to mammals such as rats, dogs, cats, cattle, horses, pigs, and humans through various routes, preferably humans. Any mode of administration may be contemplated and may be administered, for example, by oral, intravenous, intramuscular or subcutaneous injection, but is not limited thereto.

The Sena alata extract according to the present invention inhibits senescence associated beta galactosidase (SA-β-gal) activity and reduces the expression of senescence-associated proteins (p16, p21, p53 and PAI-1), thereby inhibiting excellent vascular cell senescence. Since it has an effect, the composition comprising the Senna alata extract can be used for the prevention or inhibition of vascular aging, as well as for the prevention, improvement or treatment of cardiovascular diseases, and can be usefully used in the manufacture of foods and pharmaceuticals.

1 is a graph showing the results of measuring the cytotoxicity of Sena alata extract.
2 is a photomicrograph confirming the cellular aging inhibitory effect of Sena alata extract through SA-β-gal activity staining.
3 is a photograph of Western blotting results showing the expression levels of p16, p21, p53, and plasminogen activator inhibitor-1 (PAI-1) proteins in aged vascular endothelial cells treated with Sena alata extract.

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

Example 1: Senna Alata ( Senna alata ) extract preparation

After drying Senna alata outpost (ie, whole plant), it was ground to a certain size. When using an automatic extractor (DIONEX Corporation, ASE300 ACCELERATED SOLVENT EXTRACTOR), 200 ml of methanol was added to 30 to 40 g of crushed Senna Alata, and extraction was performed at 50 ° C. and 1500 psi for 20 minutes, and a sonicator (BRANSON Ultrasonics corporation, Ultrasonic) cleaner), 1.5 L of methanol was added to 200 g of crushed Senna Alata, and extraction was performed at 50° C. for 3 days. After filtration of the obtained extract, the methanol extract was prepared by evaporating and concentrating the methanol extract.

Experimental Example 1: Human umbrical vein endothelial cell (HUVEC) culture

HUVEC cells were cultured in a 37° C., 5% carbon dioxide incubator after dispensing the cells on a 100 cm plate using an EBM-2 growth culture medium. When about 80% of the cells in the plate grew (cells 1.3 x 10 ⁶ ), the cells were separated by treatment with Trypsin-EDTA (0.25%) and passaged.

Experimental Example 2: Measurement of cytotoxicity

In order to measure the effect of the Senna alata extract according to the present invention on cell survival, 0, 12.5, 25, 50, 100, or 200 μg/ml of Senna alata (SA) extract was added to cultured vascular endothelial cells. and the cell viability was measured after 48 hours.

Specifically, 1 x 10 ⁴ vascular endothelial cells were dispensed per well in a 96-well plate, and then cultured in a 37°C, 5% carbon dioxide incubator for one day. The SA extract was treated at the above concentration and incubated. After 10uL of D-Plus™ CCK cell viability assay kit solution based on 200uL medium, absorbance was measured at 450nm 1 hour after dispensing, and cytotoxicity was investigated with CCK.

As a result, as shown in Figure 1A, it was confirmed that the SA extract increases the cell viability. However, it was confirmed that toxicity appeared at a high concentration of 200 ug/mL.

In order to determine whether the SA extract exhibits toxicity in senescence-induced cells, vascular endothelial cells were treated with H ₂ O ₂ to induce oxidative stress-induced aging. Specifically, vascular endothelial cells were treated with 0, 12.5, 25 or 50 ug/mL of the SA extract, and then treated with H ₂ O ₂ 100 ug/mL after 24 hours. After 72 hours, cell viability was checked.

Specifically, 1 x 10 ⁴ vascular endothelial cells were dispensed per well in a 96-well plate, and then cultured in a 37°C, 5% carbon dioxide incubator for one day. After pre-treatment of the SA extract for 24 hours, H ₂ O ₂ Treatment and culture for 72 hours. After 10uL of D-Plus™ CCK cell viability assay kit solution based on 200uL medium, absorbance was measured at 450nm 1 hour after dispensing, and cytotoxicity was investigated with CCK.

As a result, as shown in FIG. 1B, the group treated with H ₂ O ₂ showed cytotoxicity and decreased cell viability, but in the group treated with the SA extract, cytotoxicity decreased and cell viability increased. was confirmed.

Experimental Example 3: Confirmation of cell aging inhibitory efficacy of Sena alata extract through SA-β-gal activity staining

When oxidative stress induces aging in vascular endothelial cells, the senescence-associated beta galactosidase (SA-β-gal) staining method was used to observe the anti-aging effect of the SA extract to confirm the anti-aging effect. The cytoplasm of senescent cells by the SA-β-gal staining method is stained blue.

Specifically, 1 x 10 ⁴ vascular endothelial cells were dispensed per well in a 96-well plate, and then cultured in a 37°C, 5% carbon dioxide incubator for one day. The vascular endothelial cells were treated with 0, 12.5, 25, or 50 ug/mL of SA extract, and after 24 hours, 100 ug/mL of H ₂ O ₂ was treated. After 72 hours, the cells were washed with 1x DPBS, and the cells were fixed with 1x Fix solution (RT, 10 min). After washing, 1x staining solution was added, sealed with parafilm, and reacted at 37° C. overnight. After the reaction was completed, it was washed and the cells stained blue were observed under a microscope.

As a result, as shown in FIG. 2 , in the group treated with H ₂ O ₂ , the SA-β staining was increased, and it was observed that the blue-stained cells decreased according to the concentration of the SA extract when the SA extract was treated. became Based on this, it was confirmed that the Sena alata extract according to the present invention can inhibit vascular aging and prevent or improve cardiovascular diseases induced by vascular aging.

Experimental Example 4: Confirmation of the effect of Sena alata extract on protein expression of aging-related genes

When aging is induced, protein expression of p16, p21, p53, and plasminogen activator inhibitor-1 (PAI-1) is increased. Accordingly, it was attempted to determine the effect of the SA extract on the expression of the aging-related protein.

Specifically, 1 x 10 ⁵ vascular endothelial cells were dispensed per 1 well in a 96 well plate, and then cultured in a 37°C, 5% carbon dioxide incubator for one day. The vascular endothelial cells were treated with 0, 12.5, 25, or 50 ug/mL of SA extract, and after 24 hours, 100 ug/mL of H ₂ O ₂ was treated. After 72 hours, proteins were extracted and Western blotting was performed to observe the changes in the expression of aging-related proteins. Cells were washed with DPBS, and 50-100uL of lysis buffer (% full of cells, the amount of lysis buffer determined according to the cell state) was added. The process of vortexing on ice for 30 minutes was continued to obtain a cell crushed solution. The supernatant was recovered by centrifugation at 13000 rpm and 4° C. for 15 minutes. The amount of protein in the sample was quantified by the Bradford method (bradford reagent, sigma). The protein of the obtained sample was separated with 10-15% acrylamide gel and transferred to a PVDF membrane. To the membrane to which the protein was transferred, p16, p21, p53, and PAI-1 were attached as primary antibodies, and anti-mouse was attached as a secondary antibody to react with the antibody. The protein expression level of the reacted PVDF membrane was checked using ECL™ Select Western Blotting Detection Reagent (Amersham, RPN2235).

As a result, as shown in FIG. 3 , in the group treated with H ₂ O ₂ , the expression of p16, p21 and PAI-1 proteins increased due to cellular senescence, and in the group treated with the SA extract, p16, p21, p53 and PAI-1 protein expression was decreased. Based on this, it was confirmed that the Sena alata extract according to the present invention can inhibit vascular aging and prevent or improve cardiovascular diseases induced by vascular aging.

Claims (13)

Prevention or inhibition of vascular aging, including Senna alata extract as an active ingredient; Or a food composition for preventing or improving cardiovascular disease. 2. The method of claim 1
The Sena alata extract is water, a lower alcohol solvent having 1 to 4 carbon atoms, or a mixed solvent thereof, the food composition. According to claim 1,
The cardiovascular disease is caused by vascular aging, the food composition. According to claim 1,
The cardiovascular disease is selected from the group consisting of arteriosclerosis, hyperlipidemia, hypertension, angina, myocardial infarction, stroke, cardiac arrhythmias, heart failure, endocarditis, ischemic cerebrovascular disease and ischemic heart disease, food composition According to claim 1,
The composition reduces the activity of Senescence associated beta galactosidase (SA-β-gal) in aged vascular cells, a food composition. According to claim 1,
The composition reduces the expression of p16, p21, p53 and PAI-1 (plasminogen activator inhibitor-1), a food composition. The prevention or inhibition of vascular aging comprising the food composition according to any one of claims 1 to 6; Or a health functional food for the prevention or improvement of cardiovascular disease. Senna alata ( Senna alata ) A pharmaceutical composition for the prevention or treatment of cardiovascular diseases comprising an extract as an active ingredient. 9. The method of claim 8
The Sena alata extract is water, a lower alcohol solvent having 1 to 4 carbon atoms, or a mixed solvent thereof, a pharmaceutical composition. 9. The method of claim 8,
The cardiovascular disease is caused by vascular aging, the pharmaceutical composition. 9. The method of claim 8,
The cardiovascular disease is selected from the group consisting of arteriosclerosis, hyperlipidemia, hypertension, angina, myocardial infarction and ischemic heart disease, a pharmaceutical composition 9. The method of claim 8,
The composition reduces the activity of senescence associated beta galactosidase (SA-β-gal) in senescent vascular cells, a pharmaceutical composition. 9. The method of claim 8,
The composition reduces the expression of p16, p21, p53 and PAI-1 (plasminogen activator inhibitor-1) in aged vascular cells, a pharmaceutical composition.

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