KR20110017062A - Composition comprising ginsenoside for preventing and treating thrombotic diseases - Google Patents

Abstract

PURPOSE: A composition containing dammarane compound for preventing and treating thrombotic diseases is provided to suppress aggregation of platelet. CONSTITUTION: A pharmaceutical composition for preventing and treating thrombotic diseases contains ginsenoside Rg6, ginsenoside F4, ginsenoside Rk3, or ginsenoside Rh4 as an active ingredient. A health food for preventing and treating thrombotic diseases contains ginsenoside Rg6, ginsenoside F4, ginsenoside Rk3, or ginsenoside Rh4 as an active ingredient. The health food is powder, granule, tablet, capsule, or beverage.

Description

Composition comprising ginsenoside for preventing and treating thrombotic diseases

The present invention relates to a composition for the prevention and treatment of thrombotic diseases containing ginsenoside Rg ₆ , ginsenoside F ₄ , ginsenoside Rk ₃ or ginsenoside Rh ₄ as an active ingredient.

[Reference 1] Baek, Young-sook et al . , Antiplatelet, Antithrombotic Activity, Journal of Pharmacognosy, 26 (4) , pp.385-389, 1995

[2] Alan D. Michelson, MD., Platelets, Academic Press, Elsevier Science, USA, 2002

[Document 3] Republic of Korea Patent Registration No. 10-0302064

[Patent 4] Patent Office, 2004 New Technology Trend Report- Cardiovascular Disease Therapeutics, Vol. 2

[Ref. 5] Understanding Korean Ginseng , Korean Ginseng Society , p.9, 1995

[Document 6] Korean Patent Registration No. 192678, US Patent Registration No. 5776460

WY Kim, et al., Steaming of ginseng at high temperature enhances biological activity, J. Nat . Prod . , 63 , pp.1702-1704, 2000

8, SW Kwon, et al., Liquid chromatographic determination of less polar ginsenosides in processed ginseng, J. Chromatogr . A , 921 , pp.335-339, 2001

9 IH Park, et al., Cytotoxic dammarane glycosides from processed ginseng, Chem . Pharm. Bull ., 50 (4) , pp. 538-540, 2002

[10] YS Keum, et al., Antioxidant and anti-tumor promoting activities of the methanol extract of heat-processed ginseng, Caner Letters , 150 , pp. 41-48, 2000

KY Lee, et al., Ginsenoside-Rg ₅ suppresses cyclin E-dependent protein kinase activity via up-regulating p21 ^{Cip / WAF1} and down-regulating cyclin E in SK-HEP-1 cells, Anticancer Research , 17 , pp. 1067-1072, 1997

[12] SE Kim, et al., Ginsenoside-Rs ₃ , a new diol-type ginseng saponin, selectively elevates protein levels of p53 and p21 ^WAF1 leading to induction of apoptosis in SK-HEP-1 cells, Anticancer Research , 19 , pp. 487-492, 1999

[13] SE Kim, et al., Ginsenoside-Rs ₄ , a new type of ginseng saponin concurrently induces apoptosis and selectively elevates protein levels of p53 and p21 ^WAF1 in human hepatoma SK-HEP-1 cells, Eur . J. Cancer , 35 (3) , pp. 507-511, 1999

IH Park, et al., Three new dammarane glycosides from heat processed ginseng, Arch . Pharm . Res ., 25 (4) , pp. 428-432, 2002

15 IH Park, et al., Four new acetylated ginsenosides from processed ginseng (sun ginseng), Arch . Pharm . Res ., 25 (6) , pp. 837-841, 2002

16 YY Lee, et al., Anti-platelet aggregating and anti-oxidative activities of 11-O- (4'-O-Methylgalloyl) -bergenin, a New compound isolated from Crassula cv. 'Himaturi', Planta Med , 71 , pp.776-777, 2005

According to statistics from 2007, heart disease and cerebrovascular disease are the most common adult diseases with the second and third leading causes of death. In particular, cardiovascular diseases include diseases of heart disease, central blood vessels and peripheral blood vessels, and there are interactions between the diseases, and may develop into other disease groups as the disease progresses. Drug treatment and surgical treatment are used simultaneously for the treatment of cardiovascular diseases, but in the case of heart failure, hypertension, hyperlipidemia, coronary artery disease, arrhythmia or thrombosis, drugs are the main treatment methods. According to IMS Health, the market for cardiovascular disease drugs accounts for about one fifth of the market for all kinds of drugs in 2002. As society ages, the market for cardiovascular diseases is expected to grow steadily.

Since thrombus plays a central role in the development and progression of circulatory diseases, preventing thrombus formation and effectively eliminating thrombus is a key factor in the treatment of vascular diseases. Thrombosis is a pathological response to normal haemostasis, which can occur when platelet aggregation and plasma coagulation are excessively activated. Drugs for the prevention and treatment of thrombotic diseases are broadly classified into thrombolytic, anticoagulant and antiplatelet agents. Anticoagulants, such as heparin and coumarin, which block the clotting system to inhibit the formation of blood clots, or thrombolytic agents that dissolve the blood clots formed by breaking down fibrin, a major component of blood clots, are used for the emergency treatment of thrombotic diseases. Side effects such as bleeding at the site or systemic bleeding are not suitable for long-term treatment purposes.

Among them, antiplatelet drugs are known to be effective in the prevention of thrombosis. Platelets are discoid blood cells with a diameter of about 2 to 4 μm, and hemostasis and clot stops are formed on the injured blood vessels during blood vessel breakdown, and the coagulation process is accelerated by blood coagulation factors. It has a biological defense mechanism to prevent it. However, adhesion, aggregation, and release of platelets by platelet activation in closed blood vessels result in thrombus formation and various thrombotic diseases. In addition, microthrombosis generated by abnormal activation of platelets has been found to be involved in the expression of adult diseases such as atherosclerosis, hypertension, diabetes, as well as various thrombotic diseases, and by the release of thromboxane, serotonin, etc. Edema or inflammation may follow. Therefore, antiplatelet substances with platelet aggregation inhibitory value are of great value in preventing the recurrence of severe thrombosis such as myocardial infarction and stroke, and in the treatment and prevention of various thrombosis-related diseases (antiplatelet and antithrombotic activity of Chunma, et al. , Journal of Pharmacognosy, 26 (4) , pp.385-389, 1995).

Antiplatelet agents currently in use include nonsteroidal anti-inflammatory agents such as aspirin and indomethacin, which play a role in inhibiting prostaglandin metabolism to reduce the occurrence of thromboxane or to reduce the amount of cAMP in platelets. In particular, aspirin is a drug that inhibits the synthesis of thromboxane A ₂ by irreversibly acetylating cyclooxygenase that synthesizes prostaglandins, which are platelet aggregation inducers, from arachidonic acid on cell membranes. Platelets do not have the ability to synthesize proteins, so platelet aggregation inhibitory effect is maintained until the appearance of platelets with new enzymes, which takes about 10 days. In practice, hospital prescriptions require 100 mg of aspirin per day and 2 mg of warfarin, a blood-clotting preventive, to prevent thrombosis.

In most clinical trials, however, aspirin therapy is accompanied by a gastrointestinal side effect. In aspirin-treated patients, 5.2-40% showed symptoms such as heartburn, indigestion, nausea or vomiting, and 0.7-34% in the control group. In addition, the incidence of gastric ulcer was more than doubled in the aspirin group, affecting renal toxicity and hemorrhagic stroke, and interacting with angiotensin-converting Enzyme Inhibitor. There is a need for development of antiplatelet agents that can replace the side effects of aspirin (Alan D. Michelson, MD., Platelets, Academic Press, Elsevier Science, USA, 2002).

Platelet activating factor (PAF) receptor antagonism has been studied for 20 (S) -ginsenoside Rg ₃ and ginsenoside Rg ₅ isolated from red ginseng (Korean Patent Registration No. 10-0302064). ), There is an example showing the possibility of developing a thrombosis therapy through the study. This approach aims to explore the platelet activator receptor binding antagonism of ginsenosides Rg ₃ and Rg ₅ associated with inflammatory diseases and develop them as inflammatory or inflammatory therapeutic agents. It is believed that there is. In the process of blood clot production, various substances produced in the body work, among which platelets and platelet products, endothelial cells and endothelial cells play a very important role. According to these agents and mechanisms, the thrombosis treatment system is classified into various types (Patent Office, 2004 New Technology Trend Report- Cardiovascular Disease Therapeutics, Vol. 2), and the traditional (or general) mechanism that inhibits platelet aggregation is arachidonic acid. Antagonists such as, collagen and thromboxane A ₂ .

As such, antiplatelet agents in the blood coagulation process can block blood coagulation at an initiation stage, and thus, have become an important antithrombotic agent. In the future, antiplatelet agents have to be solved, including improved drug efficacy, improved safety, improved tolerability, and effective treatment of peripheral arterial disease. In recent years, studies are being actively conducted to obtain a platelet aggregation inhibitor component having no such side effects and excellent effects from natural products. So far, antiplatelet aggregation effects have been reported in many natural products, but the development of products at the commercialization level is still insignificant, and there is still a great need for research for the detection of antiplatelet aggregation substances.

Ginseng is a herbal medicine using the root of the perennial herb, Panax ginseng CA Meyer, belonging to the genus Araliaceae, and from ancient times, it has been recorded to have pharmacological effects such as tonic, analgesic, healthy, calming and anti-tumor. Due to the recent development of analytical techniques, a single component has been reported to explain these various pharmacological effects. Among the various components contained in ginseng, a component that can represent the medicinal effect of ginseng by exhibiting strong pharmacological activity is a ginseng-derived glycoside saponin called ginsenoside. It is a secondary metabolite of ginseng, a ginseng specific component, and its content is known to occupy about 3-6%. Saponins with high content in Korean ginseng are ginsenosides Rb ₁ , Rb ₂ , Rc, Rd, Rg ₁ , and Re. These saponin components have various medicinal effects, and their types and strengths vary according to their structure (Understanding Korean Goryeo Ginseng, Journal of Korean Ginseng Society , p.9, 1995).

Ginsenosides undergo structural changes by physicochemical processing due to heat and pressure, and the new ginsenosides exhibit strong physiological activity even at trace amounts. Red ginseng, a type of traditional numerical medicine made through steam drying for the purpose of improving preservation, contains red ginseng-specific saponins produced by structural changes of ginsenosides and not found in ginseng. Ginsenoside Rg ₂ , Ginsenoside Rg ₃ , Ginsenoside Rg ₅ , Ginsenoside Rh ₂ , Ginsenoside Rh ₃ , Ginsenoside Rh ₄ , Ginsenoside Rs ₁ , Ginsenoside Rs ₂ , Ginsenoside Rs ₃ and the like have been reported, many of them reported to have a strong pharmacological effect even by a small amount.

As such, the increase in the pharmacological action of shame natural products has been reinterpreted as being due to the change of ingredients by processing today, and the methods are diversified. Park et al. Developed a new ginseng processing method that can maximize the content of red ginseng-specific saponins by optimizing various physicochemical conditions necessary for processing ginseng (Korean Patent Registration No. 192678, US Patent No. 5776460). It has been reported that ginseng prepared according to this processing method shows significantly higher efficacy than red ginseng (WY Kim, et al., Steaming of ginseng at high temperature enhances biological activity, J. Nat . Prod . , 63 , pp.1702). -1704, 2000). The reason for this is that the content of red ginseng-specific saponins in trace amounts of red ginseng increases more than hundreds of times and new kinds of non-polar saponins are made. Kwon et al. Also contain the content of non-polar saponins in ginseng, a new heat-treated ginseng. This is reported to be significantly higher (SW Kwon, et al., Liquid chromatographic determination of less polar ginsenosides in processed ginseng, J. Chromatogr . A , 921 , pp. 335-339, 2001).

Many studies have reported the various pharmacological effects of ginseng, especially the major ginsenosides of ginseng, including new substances isolated for the first time from ginseng, show potent anticancer activity (IH Park, et al., Cytotoxic dammarane glycosides). from processed ginseng, Chem . Pharm . Bull., 50 (4) , pp.538-540, 2002; YS Keum, et al., Antioxidant and anti-tumor promoting activities of the methanol extract of heat-processed ginseng, Caner Letters , 150 , pp. 41-48, 2000; KY Lee, et al., Ginsenoside-Rg ₅ suppresses cyclin E-dependent protein kinase activity via up-regulating p21 ^{Cip / WAF1} and down-regulating cyclin E in SK-HEP-1 cells, Anticancer Research , 17 , pp.1067- 1072, 1997; SE Kim, et al., Ginsenoside-Rs ₃ , a new diol-type ginseng saponin, selectively elevates protein levels of p53 and p21 ^WAF1 leading to induction of apoptosis in SK-HEP-1 cells, Anticancer Research , 19 , pp. 487-492, 1999; SE Kim, et al., Ginsenoside-Rs ₄ , a new type of ginseng saponin concurrently induces apoptosis and selectively elevates protein levels of p53 and p21 ^WAF1 in human hepatoma SK-HEP-1 cells, Eur. J. Cancer , 35 (3) , pp. 507-511, 1999; IH Park, et al., Three new dammarane glycosides from heat processed ginseng, Arch . Pharm . Res ., 25 (4) , pp. 428-432, 2002; IH Park, et al., Four new acetylated ginsenosides from processed ginseng (sun ginseng), Arch . Pharm . Res ., 25 (6) , pp. 837-841, 2002).

In particular, attention is paid to the anticancer effects of these ingredients against ginsenoside Rg ₆ , ginsenoside F ₄ , ginsenoside Rk _3, or ginsenoside Rh ₄ , which are the main components of ginseng. However, to date, aside from the anti-cancer effects, studies on the various pharmacological effects and mechanisms of various components contained in ginseng are insignificant, and especially circulatory diseases competing for the 1st and 2nd causes of death in Korea. The study of the pharmacological effects on humans is still in its infancy.

Therefore, the present inventors separated the ginsenoside Rg ₆ , ginsenoside F ₄ , ginsenoside Rk ₃ or ginsenoside Rh ₄ from the extract obtained by specially processing ginseng to determine the antiplatelet action, the substances are arachidone The present invention was completed by confirming the excellent activity of inhibiting aggregation of platelets induced by acid or thromboxane A ₂ .

In order to carry out the above object, the present invention is ginsenoside Rg ₆ represented by the following structural formula (I), ginsenoside F ₄ represented by the structural formula (II), ginsenoside Rk ₃ represented by the structural formula (III) or One or more ginsenosides, preferably ginsenosides Rg ₆ , selected from the group consisting of ginsenosides Rh ₄ represented by the formula (IV) Or it provides a pharmaceutical composition for the prevention and treatment of thrombotic disease containing ginsenoside F ₄ as an active ingredient.

The present invention also relates to ginsenoside Rg ₆ represented by the following structural formula (I), ginsenoside F ₄ represented by the structural formula (II), ginsenoside Rk ₃ represented by the structural formula (III) or structural formula (IV). Health functional food for the prevention and improvement of thrombotic disease containing at least one ginsenoside selected from the group consisting of ginsenoside Rh ₄ , preferably ginsenoside Rg ₆ or ginsenoside F ₄ as an active ingredient. To provide.

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Thrombotic disease as defined herein includes thrombotic disease due to platelet aggregation induced by arachidonic acid, thromboxane A ₂ , and more specifically, atherosclerosis, embolism, ischemic heart disease, stroke, angina pectoris, cerebral infarction, cranial Internal bleeding, aneurysms, atherothrombosis, neurosis or myocardial infarction, preferably atherosclerosis, stroke, angina, cerebral infarction, intracranial hemorrhage or atherothrombosis, more preferably stroke, cerebral infarction or atherothrombosis.

Hereinafter, the method for obtaining the ginsenoside Rg ₆ , ginsenoside F ₄ , ginsenoside Rk ₃ or ginsenoside Rh ₄ of the present invention will be described in detail.

In the present invention, dried ginseng is put into an autoclave and heated at 70 ° C. to 150 ° C., preferably at 100 ° C. to 130 ° C., for 1 hour to 6 hours, preferably for 3 hours, and then the heat treated ginseng weight ( kg) 1 to 10 times, preferably 2 to 5 times water, C ₁ to C ₄ lower alcohol or a mixed solvent thereof, preferably ethanol, for 1 to 10 hours, preferably in a water bath After 3 hours to 6 hours cold extract, hot water extraction, ultrasonic extraction, reflux extraction, heating extraction, and the like, preferably obtained by reflux extraction filtered and dried under reduced pressure to obtain a concentrated syrup-like processed ginseng extract, Suspended by adding water 1 to 20 times, preferably 5 times to 15 times the weight of ginseng extract (g), and extracting 1 to 5 times by adding the same amount of saturated n-butanol to obtain a processed ginseng butanol fraction. Could The.

In addition, the dried ginseng butanol fractions obtained above were dried under reduced pressure and repeatedly subjected to preparative high performance liquid chromatography equipped with silica gel column chromatography and reversed phase column (C ₁₈ ) to ginsenoside Rg ₆ , ginsenoside F ₄ , Ginsenoside Rk ₃ and ginsenoside Rh ₄ can be obtained.

The present invention provides a thrombus containing one or more ginsenosides selected from the group consisting of ginsenoside Rg ₆ , ginsenoside F ₄ , ginsenoside Rk _3, or ginsenoside Rh ₄ obtained by the above-described preparation method as an active ingredient. It provides a pharmaceutical composition for the prevention and treatment of sexual diseases.

Pharmaceutical compositions containing a compound of the present invention comprise from 0.1 to 50% by weight of said compound relative to the total weight of the composition.

However, the composition as described above is not necessarily limited thereto, and may vary according to the condition of the patient and the type and extent of the disease.

Since the compound of the present invention has little toxicity and side effects, it is a drug that can be used safely even when taken for a long time for the purpose of prevention.

The compositions of the present invention may further comprise suitable carriers, excipients and diluents conventionally used in the manufacture of pharmaceutical compositions.

The compositions of the present invention can be used in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like, oral formulations, external preparations, suppositories, and sterile injectable solutions, respectively, according to conventional methods. Carriers, excipients and diluents that may be included in the composition comprising the compound include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate , Cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. When formulated, diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, and surfactants are usually used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and such solid preparations may contain at least one excipient such as starch, calcium carbonate, sucrose, or the like. ) Or lactose, gelatin and the like are mixed. In addition to simple excipients, lubricants such as magnesium styrate talc are also 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. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like can be used. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

The preferred dosage of the composition of the present invention varies depending on the condition and the weight of the patient, 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. However, for the desired effect, the compound of the present invention is preferably administered at 0.5 g / kg to 5 g / kg, preferably 1 g / kg to 3 g / kg per day. The administration may be carried out once a day or divided into several doses. Therefore, the above dosage does not limit the scope of the present invention in any aspect.

The composition of the present invention may be administered to mammals such as rats, mice, livestock, humans, and the like in various routes. All modes of administration can be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intrauterine dural or intracerebroventricular injection.

In addition, the present invention contains one or more ginsenosides selected from the group consisting of ginsenoside Rg ₆ , ginsenoside F ₄ , ginsenoside Rk ₃ or ginsenoside Rh ₄ obtained by the above method as an active ingredient. To provide a health functional food for the prevention and improvement of thrombotic disease.

The composition containing the compound of the present invention may be used in various ways, such as drugs, foods and drinks for the prevention and improvement of thrombotic diseases. Examples of the food to which the compound of the present invention may be added include various foods, beverages, gums, teas, vitamin complexes, health supplements, and the like, and may be used in the form of powders, granules, tablets, capsules, or beverages. have.

The compounds of the present invention may be added to foods or beverages for the purpose of preventing and ameliorating thrombotic diseases. At this time, the amount of the compound in the food or beverage is generally the health food composition of the present invention can be added to 1 to 5% by weight of the total food weight, the health beverage composition is 0.02 to 10 g, preferably based on 100 ml It can be added at a ratio of 0.3 to 1 g.

The health beverage composition of the present invention, in addition to containing the compound as an essential ingredient in the indicated proportions, has no particular limitation on the liquid component and may contain various flavors or natural carbohydrates as additional ingredients, such as ordinary drinks. Examples of the above-mentioned natural carbohydrates include monosaccharides such as disaccharides such as glucose and fructose such as maltose, sucrose and the like and polysaccharides such as dextrin, cyclodextrin and the like Sugar, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those described above, natural flavoring agents (tauumatin, stevia extract (e.g., Rebaudioside A, glycyrrhizin, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. The proportion of said natural carbohydrates is generally about 1-20 g, preferably about 5-12 g per 100 mL of the composition of the present invention.

In addition to the above-mentioned composition, the composition of the present invention can be used as a flavoring agent such as various nutrients, vitamins, minerals (electrolytes), synthetic flavors and natural flavors, coloring agents and intermediates (cheese, chocolate etc.), pectic acid and its salts, Salts, organic acids, protective colloid thickening agents, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonating agents used in carbonated beverages and the like. In addition, the compositions of the present invention may contain flesh for the production of natural fruit juices and vegetable beverages. These components can be used independently or in combination. The proportion of such additives is not so critical, but is generally selected in the range of 0 to about 20 parts by weight per 100 parts by weight of the composition of the present invention.

Ginsenosides Rg ₆ , Ginsenosides F ₄ , Ginsenosides Rk ₃ or Ginsenosides Rh ₄ of the present invention shows the activity of inhibiting the aggregation of platelets induced by arachidonic acid or thromboxane A ₂ , thrombus It can be used in pharmaceutical compositions and health functional foods useful for the prevention and treatment of sexual diseases.

Hereinafter, the present invention will be described in detail by reference examples, examples and experimental examples.

However, the following Reference Examples, Examples, and Experimental Examples are merely illustrative of the present invention, and the content of the present invention is not limited to the following Reference Examples, Examples, and Experimental Examples.

Example  One. Gin Senocide Rg ₆ , Gin Senocide  F ₄ , Gin Senocide Rk ₃  And Gin Senocide  Rh ₄ Manufacture

1-1. Processed ginseng extract and Fraction  Produce

Specially processed ginseng was prepared by the method of Kim et al. (WY Kim, et al., Steaming of ginseng at high temperature enhances biological activity. J. Nat . Prod . , 63 , pp.1702-1704, 2000).

1 kg of dried ginseng was put into an autoclave and heated with steam at 120 ° C. for 3 hours. 2 liters of ethanol was added to the heated ginseng, refluxed for 4 hours in a water bath, filtered, and the solvent was dried under reduced pressure to obtain 190 g of a concentrated syrup (hereinafter, referred to as SG) on a concentrated syrup.

100 g of the processed ginseng extract (SG) was suspended in 1 L of water, followed by extraction three times by adding 1 L of saturated n-butanol to obtain 50 g of the processed ginseng butanol fraction.

1-2. Gin Senocide Rg ₆ , Gin Senocide  F ₄ , Gin Senocide Rk ₃ And Gin Senocide Rh ₄ Separation of tablets

50 g of the processed ginseng butanol fraction obtained in Example 1-1 was dried under reduced pressure, followed by silica gel column chromatography (ethyl acetate: methanol: water = 20: 1: 1, v / v / v). 7.5 g of a mixture containing side Rg ₆ , ginsenoside F ₄ , ginsenoside Rk ₃ and ginsenoside Rh ₄ was obtained, and preparative high performance liquid chromatography (aceto) equipped with a reversed phase column (C ₁₈ ) was obtained from the mixture. Nitrile / water, gradient elution), repeatedly applying 13 mg, 17 mg, 15 mg and 19 mg of the desired ginsenoside Rg ₆ , ginsenoside F ₄ , ginsenoside Rk ₃ and ginsenoside Rh ₄ , respectively. Yields were obtained. For each ginsenoside obtained, its structure was confirmed by using nuclear magnetic resonance spectroscopy (NMR) and mass spectroscopy (MS), and used as a sample of the following experimental example.

Experimental Example  1. Platelet aggregation inhibitory effect

1-1. Adjusted platelet rich plasma ( adjusted platelet rich plasma Pharmacy

The experimental animals used in the present invention were anesthetized with male ethyl Sprague-Dawley male rats (240 ± 20 g) 2 weeks old with diethyl ether, followed by 10% v of 2.2% sodium citrate. Heart blood was drawn using a plastic syringe with / v. After blood collection, the blood was centrifuged at 200 x g for 10 minutes using a centrifuge (HA1000-6, Hanil Co., Ltd.), and the supernatant of the buffy coat upper layer was separated from the serum tube (10 × 250 mm, Mediland Co., Ltd., Korea) to obtain platelet rich plasma (PRP). The remaining lower layer was centrifuged again at 1500 x g for 10 minutes to obtain platelet-depleted platelet-removed plasma (PPP). After mixing PRP and PPP in an appropriate ratio, check it with a platelet analyzer (Platelet Analyzer, Excell 18 MWI, Inc., DANAM Electronics, USA) and adjust the platelet count to 600 ~ 700 × 10 ⁶ / mL and dilute with physiological saline again. The platelet count was adjusted to 400-450 × 10 ⁶ cells / mL (YY Lee, et al., Anti-platelet aggregating and anti-oxidative activities of 11-O- (4'-O-Methylgalloyl) -bergenin, a New compound isolated from Crassula cv.'Himaturi ', Planta Med , 71 , pp. 776-777, 2005).

1-2. Detection of Platelet Aggregation Inhibitory Activity

The platelet aggregation inhibitory effect was searched using the adjusted platelet rich plasma (aPRP) prepared in Experimental Example 1-1. After incubating 0.5 mL of aPRP at 37 ° C. for 3 minutes using a platelet aggregation tester (490-X, Chrono-Log Corp., USA), the ginsenoside obtained in the example was dimethylsulfoxide ( 0.005 mL of the sample dissolved in dimethylsulfoxide (final dimethyl sulfoxide concentration was less than 1%) was added thereto, followed by further incubation for 30 seconds. After right into arachidonic acid (arachidonic acid (AA), Sigma , A8798), thromboxane A ₂ (thromboxane A ₂₎ similar drug, U46619 (Sigma, D8174) as a platelet aggregation inducer to put 0.005 mL, respectively, 1,100 rpm The platelet aggregation degree was determined as a change in turbidity while stirring was continued and culture was continued at 37 ° C. Final concentration of AA was 30 ~ 60 × 10 ^-6 M and U46619 was 2 ~ 4 × 10 ^-6 M, and each aPRP was tested at the highest concentration. Particularly, in the case of arachidonic acid and U46619, since aggregation does not occur directly in rat platelets, the aggregation inducing substance (AA and U46619) was added after administration of collagen at a concentration of 0.8 to 1.2 × 10 ^-6 g / mL. Aspirin, an antithrombotic agent, was used as a control group.

As a result, as shown in Table 1, ginsenoside Rg ₆ was able to confirm a slight inhibitory activity against the aggregation induced by U46619, aspirin, a control group for the aggregation of platelets induced by arachidonic acid Inhibition at the level equivalent to In particular, ginsenoside F ₄ , ginsenoside Rk ₃ and ginsenoside Rh ₄ had a relatively low effect on platelet aggregation by arachidonic acid, but showed significantly higher inhibitory effect on platelet aggregation by U46619. Ginsenoside F ₄ showed the highest activity against platelet aggregation by U46619, which was about five times higher than aspirin, and Ginsenoside Rk ₃ and Ginsenoside Rh ₄ also showed U46619 induced platelet aggregation. When compared with aspirin, about 2.5 and 3.9 times higher activity was confirmed. In conclusion, it was confirmed that ginsenoside Rg ₆ effectively inhibits platelet aggregation by arachidonic acid and ginsenosides F ₄ , Rk ₃ , and Rh ₄ are effective in inhibiting platelet aggregation by thromboxane A ₂ . These ginsenosides demonstrate the effect of inhibiting the secondary aggregation step of platelets.

50% inhibitory concentration (IC ₅₀ , μM) Arachidonic acid U46619 Ginsenoside Rg ₆ 76 286 Ginsenoside F ₄ 114 87 Ginsenoside Rk ₃ 128 187 Ginsenoside Rh ₄ > 200 119 aspirin 63 468

Hereinafter, the formulation examples of the composition comprising the ginsenoside Rg ₆ , ginsenoside F ₄ , ginsenoside Rk ₃ or ginsenoside Rh ₄ of the present invention will be described, but the present invention is not intended to be limited thereto. To explain.

Formulation example  One. Powder  Produce

Ginsenoside Rg ₆ 20 mg

Lactose 100 mg

Talc 10 mg

The above ingredients are mixed and filled in an airtight cloth to prepare a powder.

Formulation example  2. Preparation of Tablets

Ginsenoside F ₄ 10 mg

Corn starch 100 mg

Lactose 100 mg

2 mg magnesium stearate

After mixing the above components, tablets are prepared by tableting according to a conventional method for preparing tablets.

Formulation example  3. Manufacture of capsule

Ginsenoside Rk ₃ 10 mg

3 mg of crystalline cellulose

Lactose 14.8 mg

Magnesium Stearate 0.2 mg

According to a conventional capsule preparation method, the above ingredients are mixed and filled into gelatin capsules to prepare capsules.

Formulation example  4. Preparation of injections

Ginsenoside Rh ₄ 10 mg

Mannitol 180 mg

Sterile distilled water for injection 2974 mg

Na ₂ HPO ₄ , 12H ₂ O 26 mg

According to the conventional method for preparing an injection, the amount of the above ingredient is prepared per ampoule (2 ml).

Formulation example  5. Liquid  Produce

Ginsenoside Rg ₆ 20 mg

10 g of isomerized sugar

5 g of mannitol

Purified water

After dissolving each component in purified water according to the usual method of preparing a liquid solution, adding lemon flavor appropriately, mixing the above components, adding purified water, adjusting the whole to 100 ml by adding purified water, and then filling into a brown bottle. The solution is prepared by sterilization.

Formulation example  6. Manufacture of health food

Ginsenoside F ₄ 1000 mg

Vitamin mixture proper amount

70 μg of Vitamin A Acetate

Vitamin E 1.0 mg

0.13 mg vitamin B1

0.15 mg of vitamin B2

0.5 mg vitamin B6

0.2 [mu] g vitamin B12

10 mg vitamin C

Biotin 10 μg

Nicotinic acid amide 1.7 mg

50 ㎍ of folic acid

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

15 mg of potassium phosphate monobasic

Secondary calcium phosphate 55 mg

Potassium citrate 90 mg

100 mg of calcium carbonate

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.

Formulation example  7. Manufacture of health drinks

Ginsenoside Rk ₃ 1000 mg

Citric acid 1000 mg

100 g of oligosaccharide

Plum concentrate 2 g

Taurine 1 g

Purified water was added to a total of 900 ml

The above components were mixed according to a conventional health drink manufacturing method, and the mixture was heated at 85 DEG C for about 1 hour with stirring, and the solution thus prepared was filtered to obtain a sterilized 2-liter container, which was sealed and sterilized, ≪ / RTI >

Although the composition ratio is a mixture of the components suitable for the preferred beverage as a preferred embodiment, the blending ratio may be arbitrarily varied according to the regional and national preferences such as the demand level, the demanding country, and the intended use.

Claims (6)

Pharmaceutical composition for the prevention and treatment of thrombotic diseases containing at least one ginsenoside selected from the group consisting of ginsenoside Rg ₆ , ginsenoside F ₄ , ginsenoside Rk ₃ or ginsenoside Rh ₄ . The method of claim 1, wherein the ginsenoside is ginsenoside Rg ₆ Or ginsenoside F ₄ pharmaceutical composition. The pharmaceutical composition according to claim 1, wherein the thrombotic disease is a thrombotic disease due to platelet aggregation induced by arachidonic acid or thromboxane A ₂ . The pharmaceutical composition of claim 1, wherein the thrombotic disease is atherosclerosis, embolism, ischemic heart disease, stroke, angina, cerebral infarction, intracranial hemorrhage, aneurysm, atherothrombosis, neurosis or myocardial infarction. Health function for the prevention and improvement of thrombotic diseases containing at least one ginsenoside selected from the group consisting of ginsenoside Rg ₆ , ginsenoside F ₄ , ginsenoside Rk ₃ or ginsenoside Rh ₄ as an active ingredient food. A dietary supplement according to claim 5 which is a powder, granule, tablet, capsule or beverage.