KR20190063616A - Composition for promoting angiogenesis comprising extract of stichopus japonicas red - Google Patents

Abstract

The present invention relates to a composition for promoting angiogenesis containing an extract of Stichopus japonicus Red as an active component. The extract of Stichopus japonicus Red of the present invention exhibits an angiogenesis promoting activity, thereby being able to be usefully used in fields requiring promotion of angiogenesis, for example, treatment, prevention, and alleviation of angiogenesis-dependent diseases.

Description

Technical Field [0001] The present invention relates to a composition for promoting angiogenesis including red sea ginseng extract,

The present invention relates to a composition for promoting angiogenesis comprising an extract of Red Sea Ginseng as an active ingredient.

Angiogenesis is a biological process that provides new blood vessels to tissues or organs, which means that new blood vessels are created from existing blood vessels and play an important role in tissue regeneration, wound healing and female reproductive cycle. Angiogenesis is tightly regulated by neovascularization regulators, and consists of a complex and sequential process of multiple stages rather than merely proliferation of vascular endothelial cells. The blood vessels circulate the blood between the organs and tissues of the heart and the human body. If the oxygen or nutrients are not properly supplied to the tissue due to stenosis or bleeding in a part of the blood vessels, the cells are seriously damaged. In addition, diseases such as arteriosclerosis, myocardial infarction, or angina pectoris are caused by an inability to smoothly supply blood, so substances that promote angiogenesis may be usefully used in the treatment of such diseases. Accordingly, the development of a material having an effect of promoting angiogenesis using natural substances having relatively few side effects for treatment of ischemic diseases and the like has attracted much attention.

On the other hand, sea cucumber is a generic term for sea cucumbers belonging to the echinoderms of echinoderms. It is called 'sea cucumber' in the meaning of ginseng in the sea. There are about 1,500 species of sea cucumber worldwide, and there are 4 or 14 species in Korea. The sea cucumbers in Korea are classified into blue sea cucumbers, black sea cucumbers and red sea cucumbers according to their color, and they are known to have different prey and habitat. The sea cucumber contains a large amount of minerals such as calcium, iron or phosphorus. Especially, it has high calcium and iron content. It helps the hematopoietic action of the child and the pregnant woman during the growing season. It is known to be effective in prevention of obesity and adult diseases as a low calorie food. It has been used as a nourishing lotion and as a preservative food. Recently, as the excellent effects of sea cucumbers have been widely known, many studies on sea cucumbers as medicines, cosmetics and functional food materials are being carried out. Technology related to commercialization of sea cucumbers including red sea ginseng began in the late 2000s, and the field of technology development related to high functionality commercialization utilizing these technologies is a field with unlimited growth potential. Korean Patent Laid-Open Publication No. 2012-0063709 discloses a composition for enhancing immunity comprising an extract of sea cucumber as an active ingredient, and Korean Patent Laid-Open Publication No. 2011-0135908 discloses an anti-inflammatory agent composition and an anticancer agent composition containing Red Sea Ginseng Extract. However, the composition for promoting a new blood vessel containing the red sea ginseng extract of the present invention as an effective ingredient has not yet been disclosed.

It is an object of the present invention to provide a composition for promoting angiogenesis.

Another object of the present invention is to provide a pharmaceutical composition, a cosmetic composition and a food composition for promoting angiogenesis.

In order to accomplish the above object, one aspect of the present invention provides a composition for promoting angiogenesis comprising an extract of Red Sea Ginseng as an active ingredient.

In the present specification, the term "red sea ginseng" means a sea cucumber whose scientific name is Stichopus japonicus Red or Stichopus japonicus Orange. It belongs to Aspiodchirotida, Stichopodidae, and Stichopus. Specifically, the body color is red, yellowish brown or dark reddish brown, and the abdomen is red. The polian vesicle is thin and long, the tip is sharp, and the shrinkage is severe, so that the body shape is similar to the sphere, It can be defined as a sea cucumber having a gelatin filament of about 25 탆 in thickness on an egg. The red sea ginseng generally live mainly in rocks and gravel areas (Ondasung, Jeju, Ulleung, Guiza Island, etc.).

The term "extract" means a product obtained by extracting with an appropriate extraction solvent, and the resultant product may be a solution obtained by diluting an extract, a diluent or concentrate of the extract, a fraction of the extract, a dried product obtained by drying the extract, All included.

The term "red sea ginseng extract" means an extract obtained from red sea ginseng. For example, it may be an extract obtained from red sea ginseng in which whole red sea gins or a part such as gut is removed.

The extraction solvent may be selected from the group consisting of water, a lower alcohol having 1 to 4 carbon atoms (such as methanol, ethanol, propanol or butanol), an organic solvent (hexane, acetone, chloroform, methyl acetate, ethyl acetate and the like) And preferably the extraction solvent can be used for the mixing of water and alcohol. The extraction solvent may more preferably be ethanol of 50 to 80%, 55 to 80%, 60 to 80%, 60 to 75%, 55 to 75%, or 60 to 75%, and most preferably 60 to 75 % Ethanol.

The red sea ginseng extract of the present invention may more preferably be obtained by extracting red sea gins with ethanol and further extracting with ethyl acetate. The ethyl acetate extraction can be carried out, for example, by adding ethyl acetate to the ethanol extract of red sea gull, stirring the mixture, and then removing the supernatant and remaining ethyl acetate. In addition, the ethanol extract of Red Sea Ginseng used in the ethyl acetate extraction may preferably be a concentrate of 20% to 40% of the result obtained by extracting Red Sea Gins through ethanol. In one embodiment, the red sea gums extracted from ethanol and further extracted with ethyl acetate showed high thermal stability and the highest content of anilino acetic acid, an indicator of red sea ginseng.

The red sea ginseng extract may be prepared by a general extraction method known in the art, and examples of the extract include immersion, hot water extraction, hot water extraction, cold extraction, reflux cooling extraction, steam distillation extraction, alcohol extraction, / Or ultrasonic extraction may be used. Preferably, the red sea ginseng extract may be prepared by one or more extraction methods selected from the group consisting of immersion, hot water extraction and alcohol extraction, and more preferably, the red sea ghrecipes extract may be prepared by immersion and heating.

Specifically, the red sea ginseng extract comprises (a) immersing red sea ginseng in a solvent; (b) heating the resultant obtained in step (a); And (c) cooling the resultant heated in the step (b). In the step (a), the solvent may be preferably used for the mixing of water and ethanol.

In the above extraction method, the immersion may preferably be performed at room temperature for 24 hours or more. More preferably, the immersion step may be to immerse the ground red sea gull in a 50 to 80% alcohol solvent at room temperature for not less than 24 hours but not longer than 48 hours. In the above extraction method, the heating step may preferably be heating at a temperature of 100 DEG C or more for 3 hours or more. In the above extraction method, the cooling may preferably be performed at a temperature of 60 DEG C or lower for 3 hours or more.

Removing the red ginseng guts and washing the red ginseng before immersing the ginseng in a solvent; Rapidly freezing the washed red sea ginseng; Freezing the red sea germ in a refrigerated state, and pulverizing the red sea germ.

The extraction method may further include a step of filtering and / or concentrating the extract after the cooling step. The filtration can be preferably performed using a 10 mu m filter cloth. The concentration step may preferably be performed at a temperature of 60 DEG C or lower. The concentration may be such that the solid content is 20% or more and less than 40%, 25% or more and 40% or less, 25% or more and 35% or less, or 30% or more and less than 40% For example, 28% or more and 32% or less.

The term "active ingredient" means an ingredient that alone exhibits the desired activity or is capable of exhibiting activity with a carrier that is itself inactive.

The term "angiogenesis" refers to all phenomena in which blood vessels are newly formed as a phenomenon of new blood vessels. For example, the cells constituting existing blood vessels may proliferate and migrate to create new blood vessels And may include a phenomenon in which blood vessels are newly generated by precursor cells that make blood vessels

The term "promoting angiogenesis" means promoting angiogenesis as described above, so that blood vessel formation such as capillary blood vessels can be performed in a shorter time and / or more blood vessels can be produced. The composition for promoting angiogenesis of the present invention may specifically be a composition for skin board regeneration, wound and burn treatment, artificial skin transplantation and transplantation blood vessel preparation. In the case of skin plate regeneration, scarring and burn treatment, artificial skin grafting and transplantation blood vessel preparation, induction of tissue formation is required and smooth supply of blood is essential. The composition of the present invention is used to promote angiogenesis .

Another aspect provides a pharmaceutical composition comprising the above angiogenesis promoting composition.

In the pharmaceutical composition according to the present invention, each term is as mentioned above in the composition for promoting angiogenesis, unless otherwise specified.

The pharmaceutical composition may be used to treat or prevent angiogenesis-dependent diseases. The term "treatment" refers to any action in which the symptoms of an angiogenesis-dependent disease are ameliorated or cured by administration of a composition according to the present invention. The term "prophylactic" also refers to any act that inhibits or delays the symptoms of an angiogenesis dependent disease by administration of the composition according to the present invention.

The term "angiogenesis-dependent disease" is a disease accompanied by symptoms in which the blood supply is not smooth or the angiogenesis is not properly performed, and the disease which can be prevented or treated by the angiogenesis- promoting effect of the red sea ginseng extract of the present invention The kind thereof is not particularly limited. For example, the angiogenesis-dependent diseases may include wounds, burns, arteriosclerosis, myocardial infarction, angina, varicose veins or cerebrovascular dementia. Such angiogenesis-dependent diseases require promotion of angiogenesis in their prevention and treatment. For example, in the case of a wound or burn, angiogenesis is required for the supply of nutrients for the recovery of damaged tissue, and in the case of arteriosclerosis, myocardial infarction, angina pectoris, varicose vein or cerebrovascular dementia, angiogenesis is required to improve the blood flow disorder . The red sea ginseng extract of the present invention having an activity of promoting angiogenesis has an effect of increasing the formation ability of a new blood vessel by increasing the proliferation and mobility of vascular endothelial cells and can be usefully used for the prevention or treatment of angiogenesis dependent diseases .

The content of the composition for angiogenesis in the pharmaceutical composition of the present invention can be appropriately adjusted according to the symptoms of the disease, the progress of the symptoms, the condition of the patient, and the like. For example, the amount of the composition of the present invention may vary from 0.0001 to 99.9 wt%, 0.1 to 90 wt%, 1 to 80 wt%, 1 to 70 wt%, 1 to 60 wt%, or 1 to 50 wt% ≪ / RTI >

The pharmaceutical compositions of the present invention may further comprise suitable carriers, excipients and diluents conventionally used in the manufacture of pharmaceutical compositions. Examples of carriers, excipients and diluents that may be contained in the composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, But are not limited to, cellulose, methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral.

The pharmaceutical composition of the present invention may be an oral dosage form, and may be formulated into a suitable dosage form for topical administration using techniques well known to those skilled in the art. In the case of topical dosage form, the formulations may include external preparations, foams, suppositories, and the like.

The pharmaceutical composition of the present invention is administered to a subject in a pharmaceutically effective amount. As used herein, the term "pharmaceutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment for medical treatment and the effective dosage level will vary depending on the species and severity, The sex, the activity of the drug, the sensitivity to the drug, the time of administration, the route of administration and the rate of release, the duration of the treatment, factors including co-administered drugs, and other factors well known in the medical arts. The composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents. And can be administered singly or multiply. It is important to take into account all of the above factors and to administer the amount in which the maximum effect can be obtained in a minimal amount without adverse effect, and can be easily determined by those skilled in the art.

Another aspect provides a cosmetic composition comprising the composition for promoting angiogenesis.

In the cosmetic composition according to the present invention, each term including a composition for promoting angiogenesis is as mentioned in the above composition for promoting angiogenesis, unless otherwise specified.

The cosmetic composition according to one embodiment is present in an amount of from 0.0001 to 99.9% by weight, from 0.1 to 90% by weight, from 1 to 80% by weight, from 1 to 70% by weight, from 1 to 60% by weight, or from 1 to 50% And a composition for promoting angiogenesis.

It is needless to say that the above-mentioned cosmetic composition can be produced in the form of a general emulsified formulation and a solubilized formulation or the like, using a generally known production method. At this time, the cosmetic composition may be appropriately selected and formulated according to the purpose. For example, it can be manufactured in various formulations such as cosmetics, essences, creams, packs, patches, skin adhesive gels, foundations, make-up bases, and the like, and can be applied to a common method for producing cosmetics.

It is needless to say that the cosmetic composition according to one embodiment may further include various additives in addition to the composition for promoting angiogenesis. The additive may be, for example, a stabilizer, an emulsifier, a thickening agent, a moisturizer, a liquid crystal film strengthening agent, a pH adjusting agent, an antibacterial agent, a water-soluble polymer, a coating agent, a metal ion sequestrant, an amino acid, an organic amine, a polymer emulsion, At least one aqueous additive selected from an antioxidant, an antioxidant, a preservative, a perfume, and the like; And one or more oily additives selected from oils, waxes, hydrocarbonaceous oils, higher fatty acid oils, higher alcohols, synthetic ester oils, and silicone oils; And so on.

Another aspect provides a health functional food composition comprising the composition for promoting angiogenesis.

In the food composition according to the present invention, each term including the composition for promoting angiogenesis is the same as mentioned above, unless otherwise specified.

The food composition can improve or prevent angiogenesis-dependent diseases.

The term "improvement" refers to all actions that at least reduce the degree of a parameter associated with a condition for which the angiogenesis dependent disease is being treated, for example, a symptom.

The term "health functional food " means food prepared and processed in the form of tablets, capsules, powders, granules, liquids, and circles using raw materials and components having useful functions in the human body. The term "functional" as used herein means that the structure and function of the human body have a beneficial effect on health uses such as controlling nutrients or physiological actions.

The food composition according to the present invention can be prepared by a method commonly used in the art and can be prepared by adding raw materials and components that are conventionally added in the art. In addition, unlike general medicines, there is an advantage that there is no side effect that may occur when a medicine is taken for a long time by using food as a raw material, and since it is excellent in portability, the food composition of the present invention is effective for improving or improving the therapeutic effect of an angiogenesis- It can be taken as an adjuvant for.

The amount of the composition for promoting angiogenesis contained as an active ingredient in the food composition according to one embodiment can be suitably determined according to the intended use (prevention, improvement, or therapeutic treatment). In general, the composition for promoting angiogenesis of the present invention may be contained in an amount of 0.001 to 20% by weight, 0.001 to 15% by weight or 0.001 to 10% by weight in the raw material composition when the food is prepared. In the case of health drinks, 0.01 to 2 g, specifically 0.02 to 2 g, more specifically 0.3 to 1 g, may be added based on 100 mL. However, in the case of long-term ingestion intended for health and hygiene purposes or for the purpose of controlling health, the amount can also be used in the above-mentioned range. The composition for promoting angiogenesis according to the present invention, which is added to a food composition in the course of preparing the food composition, can be appropriately added or decreased as needed.

The food composition may be any one selected from the group consisting of pills, tablets, granules, powders, capsules, and liquid solutions.

The red sea ginseng extract of the present invention exhibits angiogenesis promoting activity and thus can be usefully used for the treatment, prevention or amelioration of an angiogenesis-dependent disease in an area in which angiogenesis promotion is required.

1 is a graph showing the results of analysis of cell survival rate of Red Sea Ginseng extract according to an embodiment of the present invention.
2 is a microscopic examination of the angiogenesis inducing effect of the red sea ginseng extract using Chicken Chorioallantoic Membrane (CAM) assay.

Hereinafter, the present invention will be described in more detail with reference to examples. However, these examples are for illustrative purposes only, and the scope of the present invention is not limited by these examples.

Example  One. Red sea ginseng  Preparation of extract

After removing the viscera of the raw material, red sea gums, the red sea gums were pulverized using a pestle. 800 g of pulverized red sea goby was immersed in 4 kinds of extracting solvents (distilled water, 20% ethanol, 50% ethanol and 70% ethanol) of 3.2 kg each for 36 hours. Each of the dipped products was then heated at 100 DEG C for 3 hours and then at a temperature of 60 DEG C or less for 3 hours. The cooled product was filtered with a filter paper filter (10 mu m filter paper (ADVANTEC No. 2) to remove the suspension. At this time, 0.1% diatomaceous earth was added as a filter aid. The filtered extract was concentrated at 50 ° C to a concentration of about 30% by a rotary evaporator, and sterilized at 95 ° C for 5 minutes. The obtained concentrate was used as a sample for analysis of angiogenesis induction activity of the extract after refrigerated storage.

Analysis of red sea ginseng extracts by four kinds of solvents revealed that red sea ginseng contained the most abundant index components in 70% ethanol extract. 70% ethanol extract was used in the experiment to analyze the vascular promoting effect of red sea ginseng extract.

Example  2. Red sea ginseng  Cytotoxicity analysis of extracts

In order to evaluate the safety of red sea ginseng extract, cell viability of human epithelial cell extracts was determined by MTT assay. Specifically, HaCaT cells as a keratinocyte cell line were inoculated in a 12-well plate at a concentration of 1 × 10 ⁴ cells / well and cultured for 24 hours, followed by washing with PBS. Then, a sample of 70% ethanolic red sea ginseng extract diluted with each concentration (50, 100, 150, 200 ㎍ / ml) was added to each well (control was untreated) and cultured for 24 hours. After culturing for 24 hours, 40 쨉 l of 0.5% MTT solution was added to each well, followed by incubation in a 5% CO ₂ incubator at 37 캜 for 1 hour. After removing the culture medium, 150 μl of DMSO was added and shaken for 10 minutes. Absorbance was measured at 595 nm using an ELISA plate reader.

As a result, as shown in Fig. 1, the control group and the red sea germ extract treatment group had no significant effect on cell proliferation. Especially, it was confirmed that red sea ginseng extract was not toxic even at a high concentration of 200 μg / μl. As a result, it was confirmed that the treatment of the extract of Red Sea Ginseng does not show toxicity to the cells.

Example  3. Red sea ginseng  Identification of induction of angiogenesis by extract

Chicken Chorioallantoic Membrane (CAM) assay was used to confirm the angiogenesis promoting activity of the extract of Red Sea Ginseng according to the present invention. Specifically, embryos were purchased and sterilized with 70% ethanol, and incubated for 3 days in an incubator maintained at 37 ° C and 90% humidity. At this time, the temperature of the incubator was 37-38 ° C, and the humidity was checked over time so as to be maintained at 90% or more. On the 3rd day (2nd day), the egg of the embryo was scratched with a knife, and then horizontally laid. Then, a hole was made with a 5 ml syringe and 2 ml of albumin was extracted. The embryos were sealed with a glass tape so that the embryos did not dry out and were not infected. The embryos were placed with the holes facing downward and then cultured again. On the 4th day (3 rd day), a circular window with a diameter of 2-3 ㎝ was placed on the side of the air sac of the embryo (opposite to the syringe hole). CAM was generated at the fifth day (4.5 times) and it was observed that the diameter was about 2-5 mm. 200 ㎍ / ㎖ of Red Sea Ginseng Extract and the control sample were dissolved in secondary distilled water, and then 10 ㎕ of each of them was placed on a four-divided Thermanox cover slip and dried in a clean bench. The glass tape of the embryo was cut with a knife, and the CAM was found and checked. The Thermanox, which had been treated with the tweezers, was turned upside down and carefully placed on the glass tape. On the 7th day (6.5-fold), the glass tape was stabbed with a knife, and 1 ml of intralipose (fat emulsion) was taken with a syringe, and bubbles were removed and injected into the lower part of the CAM. At this time, clear blood vessels were observed on the white background. When injecting intralipose with a syringe, the blood vessels were not injured. Observed embryos were photographed with a camera.

As a result, new blood vessels were observed by treatment with each of the red sea ginseng extracts, and it was confirmed that the extract of red sea gins extract had effective activities to induce angiogenesis to a degree similar to that of the vitamin C treatment group and the commercial product treatment group (FIG. 2).

Claims (8)

A composition for promoting angiogenesis, comprising an extract of Red sea ginseng as an active ingredient. The method according to claim 1,
Wherein the red ginseng extract is extracted with at least one solvent selected from the group consisting of water, an alcohol having 1 to 4 carbon atoms, and a hydrocarbon-based solvent. The method according to claim 1,
Wherein the red ginseng extract is extracted using 50 to 80% ethanol as a solvent. The method according to claim 1,
The red sea ginseng extract,
(a) immersing red ginseng in a mixed solvent of water and ethanol;
(b) heating the resultant obtained in step (a); And
(c) cooling the resultant heated in the step (b). A pharmaceutical composition for preventing or treating angiogenesis-dependent diseases, comprising the composition for promoting angiogenesis according to any one of claims 1 to 4. 6. The method of claim 5,
Wherein said angiogenesis-dependent disease is a disease selected from the group consisting of wounds, burns, arteriosclerosis, myocardial infarction, angina, varicose veins and cerebrovascular dementia. A cosmetic composition comprising the composition for promoting angiogenesis according to any one of claims 1 to 4. A functional food composition for preventing or ameliorating an angiogenesis-dependent disease comprising the composition for promoting angiogenesis according to any one of claims 1 to 4.

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