KR20170054115A - A pharmaceutical composition comprising extract from germinated gemmule of bean for preventing or treating osteoporosis - Google Patents

Abstract

The present invention relates to a pharmaceutical composition comprising an extract from germinated gemmule of beans for preventing or treating osteoporosis and, more specifically, to a pharmaceutical composition comprising an extract from germinated gemmule of beans or fractions thereof for preventing or treating osteoporosis, a method for preventing or treating osteoporosis comprising the step of administering the composition into an individual except for human, and a food composition and a feed composition comprising an extract from germinated gemmule of beans or fractions thereof for preventing or alleviating osteoporosis. The extract from germinated gemmule of beans improves proliferation and differentiation abilities of osteoblast and increases the bone density, the bone volume and the number of osteoblasts, thereby being used for preventing or treating osteoporosis.

Description

Technical Field [0001] The present invention relates to a pharmaceutical composition for preventing or treating osteoporosis including a germinated germ extract of soybean germ,

The present invention relates to a pharmaceutical composition for preventing or treating osteoporosis, which comprises a soybean germinated embryo extract. More specifically, the present invention relates to a pharmaceutical composition for preventing or treating osteoporosis comprising a soybean germinated embryo extract or a fraction thereof, To a food composition for preventing or ameliorating osteoporosis comprising a soybean germinated embryo extract or a fraction thereof, and a feed composition.

Osteoporosis is a disease in which the amount of bone is decreased and the strength of the bone is weakened so that fracture tends to occur even in a small impact. It is known that osteoporosis is a problem due to the limitation of long-term activities such as femoral fractures or vertebral fractures, which are caused by the weakening of the bone rather than the symptom itself, and it accounts for about 15% of the elderly deaths.

The causes of osteoporosis include age, lack of exercise, underweight, smoking, low calcium diet, menopause, and ovariectomy, especially in women, bone reduction continues after 30 years of age, . That is, estrogen concentration rapidly decreases in menopause, thereby increasing the activity of osteoclasts, decreasing the amount of calcium contained in the bone, and weakening the structure that maintains the bone morphology.

To treat such osteoporosis, a hormone, alendronate, calcitonin, raloxifene, Na-F, calcitriol or a bisphosphonate preparation is administered. However, since the treatment of osteoporosis requires a long-term treatment period, long-term use of conventional drugs increases the possibility of side effects such as urinary stone, endometrial cancer, and breast cancer. In particular, adverse effects such as breast cancer, myocardial infarction, and venous thrombosis have been reported in hormone therapy including estrogen and calcitonin, which are currently widely used for osteoporosis treatment (JAMA 2002, 288, 872-881, J. Obstet., Bynaecol. 2002, 24, 711-715). In addition, while bisphosphonate has been attracting attention as a new alternative therapeutic agent as a bone resorption inhibitor for inhibiting osteoclast, it has been reported that the absorption rate is low, and lesions are observed in the upper airway when the oral administration is incorrect (Clin. Proc. 2002, 77, 1031-1043).

Accordingly, it is urgently required to develop a therapeutic agent having less toxicity and side effects. In recent years, research on alternative therapies using natural ingredients derived from natural products such as herbal medicines and foods, which can promote bone formation while minimizing bone loss, A manufacturing method is being developed. However, development of a natural pharmaceutical composition or a raw material thereof, which has excellent effects on prevention and improvement of osteoporosis and has less side effects than conventional synthetic pharmaceutical compositions, has not yet been developed.

Under these circumstances, the inventors of the present invention have made intensive efforts to develop a therapeutic agent for osteoporosis using a natural-derived component. As a result, it has been found that the extract of soybean germinated embryo produced by germination of discarded soybean embryo in the process of manufacturing processed products using soybean, The present invention has been accomplished on the basis of this finding that it can be used as a prophylactic and therapeutic agent for osteoporosis by promoting bone formation by increasing bone density, bone volume and number of osteocytes.

It is an object of the present invention to provide a pharmaceutical composition for preventing or treating osteoporosis, which comprises a soybean germinated embryo extract or a fraction thereof.

It is another object of the present invention to provide a method for preventing or treating osteoporosis, which comprises the step of administering the composition to a subject other than a human.

It is still another object of the present invention to provide a food composition for preventing or ameliorating osteoporosis comprising a soybean germinated embryo extract or a fraction thereof.

It is another object of the present invention to provide a feed composition for preventing or ameliorating osteoporosis comprising a soybean germinated embryo extract or a fraction thereof.

One aspect of the present invention provides a pharmaceutical composition for preventing or treating osteoporosis, which comprises a soybean germinated embryo extract or a fraction thereof.

The term "soybean germinated embryo" of the present invention means a germinated embryo obtained by germinating an embryo isolated from soybean. A method for producing a soybean germinated embryo has been known from previous studies of the present inventors (Korean Laid-Open Patent Application No. 2013-0057173). Specifically, the soybean germinated embryo may be obtained by germinating an embryo isolated from soybean, But is not limited thereto. The therapeutic effect of the germinated germ extract of the present invention on osteoporosis has not been known at all and has been identified for the first time by the present inventors.

The soybean germinated embryo extract of the present invention may be obtained by extracting the soybean germinated embryo with water, a lower alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof.

The term "extract " of the present invention means an extract obtained by extracting a soybean germinated embryo, a diluted solution or concentrate of the extract, a dried product obtained by drying the extract, a preparation or a purified product of the extract, Extracts themselves and extracts of all formulations which can be formed using extracts.

In the soybean germinated embryo extract of the present invention, the method for extracting the soybean germinated embryo is not particularly limited and may be carried out according to a method commonly used in the art. Non-limiting examples of the extraction method include hydrothermal extraction, ultrasonic extraction, filtration, and reflux extraction. These may be performed alone or in combination with two or more methods.

In the present invention, the kind of the extraction solvent used for extracting the soybean germinated embryo is not particularly limited, and any solvent known in the art can be used. Non-limiting examples of the extraction solvent include water, alcohol, and a mixed solvent thereof. These solvents may be used alone or in combination. When an alcohol is used as a solvent, an alcohol having 1 to 4 carbon atoms can be specifically used.

The fraction of the soybean germinated embryo extract of the present invention may be obtained by fractionating the above extract with water, a lower alcohol having 1 to 4 carbon atoms, a non-polar solvent, or a mixed solvent thereof.

The term "fraction " of the present invention means a product obtained by performing fractionation to separate a specific component or a specific component group from a mixture containing various components.

The fractionation method for obtaining the fraction in the present invention is not particularly limited and may be carried out according to a method commonly used in the art. As a non-limiting example of the above-mentioned fractionation method, there can be mentioned a method of treating a soybean germinated embryo extract of the present invention with a predetermined solvent to obtain a fraction from the extract.

The kind of the fraction solvent used for obtaining the fraction in the present invention is not particularly limited, and any solvent known in the art can be used. Non-limiting examples of the fraction solvent include polar solvents such as water and alcohol; And non-polar solvents such as hexane, ethyl acetate, chloroform, and dichloromethane. These may be used alone or in combination of one or more. When an alcohol is used in the fraction solvent, an alcohol having 1 to 4 carbon atoms can be specifically used.

The soybean germinated embryo extract of the present invention may be one containing isoflavone or soyasaponin.

The term "isoflavone" of the present invention is a kind of polyphenol compound having a structure similar to estrogen, and is also called phytoestrogen. The isoflavone is highly absorbed into the body and binds to the estrogen receptor to activate its action.

Specific examples of the isoflavone include (E) - ((2R, 3S, 4S, 5R, 6S) -3,4,5-trihydroxy- 6- (3- (4-hydroxyphenyl) 2- (4-hydroxyphenyl) -4-oxo-4H-chromen-7-yloxy) -tetrahydro-2H- pyran- 4-oxo-4H-chromen-7-yl 6-O - [(2E) -2-butenoyl] hexopyranoside.

[Chemical Formula 1]

(2)

The term " soyasaponin "of the present invention is a kind of phytosterol contained in soybeans, and it is a phytosterol which is based on aglycone structure such as steroid or triterpenoid, Respectively. Specific examples of the soy saff saponin include triacetyl soyasaponin Ab, Diacetyl soyasaponin Aa, Soyasaponin Ab, Soyasaponin Ae, Soyasaponin Af, Soyasaponin Aa, Soyasaponin Ac, Soyasaponin Ag, Soyasaponin Ad, Soyasaponin Ah, Soyasaponin Ba, Soyasaponin Bc, Soyasaponin Bb, Soyasaponin Bd , Soyasaponin Be, Soyasaponin? G, Soyasaponin? Or Soyasaponin? G.

In one embodiment of the present invention, the soybean germinated embryo extract was applied to an UPLC QTOF-mass spectrometer, and major compounds were selected. As a result, it was confirmed that soybean saponin, which is a new substance of isoflavone series and saponin series, was contained (Table 1).

The soybean germinated embryo extract of the present invention may improve osteoblast proliferation and differentiation ability.

The term "osteoblast ", also referred to as osteoblast, refers to a cell capable of calcining bone tissue by synthesizing and secreting bone mineral and depositing inorganic salts such as Ca and Mg ions on the substrate. do. It is mainly observed at the site of osteogenesis by ossification or the like.

The term "proliferative ability" of the present invention means the ability of a cell to divide and proliferate. Improvement of the proliferative capacity means an increase in the number of cells, and an increase in the number of cells can improve the effect of the characteristics of the cells. For the purpose of the present invention, the cell refers to osteoblast, and osteoblast growth can be promoted thereby promoting bone regeneration.

The term "differentiation ability" of the present invention means the ability of a less specialized cell to develop into a specific cell, and the improvement of the differentiation ability means that the specific biological activity of the cell is improved. For the purpose of the present invention, the cells refer to osteoblasts. When the osteoblast differentiation capability is improved, the bone matrix is matured and mineralized, thereby accumulating calcium and promoting bone regeneration.

In one embodiment of the present invention, it was confirmed that the soybean germinated embryo extract does not show cytotoxicity even at a concentration of 500 μg / mL, but rather enhances the proliferative capacity of osteoblasts (FIG. 1). In addition, it was confirmed that the degree of differentiation of osteoblast was further improved when the extract of 500 / / mL was treated (Fig. 2). This suggests that the extract is a safe substance free from harm to human body and promotes bone formation by activating the proliferation and differentiation of osteoblasts.

The soybean germinated embryo extract of the present invention may increase bone density, bone volume, and osteocyte count.

The term "bone density " of the present invention means a measure of the bone mineral content and is a criterion for determining the rigidity of the bone. The World Health Organization defines osteoporosis as a bone mineral density of less than 2.5% standard deviation of young adults, ie, less than 3%. Bone density is reduced by 2% every 5 years after reaching the peak around 30 years, and it decreases by 3 times faster than postmenopause.

The term "bone mass " of the present invention means the thickness of the bone, and when the bone volume is reduced, the bone becomes thin and tender.

The term "bone area " of the present invention means the size of the bone, and when the area of the bone is reduced, the bone is thin and torn.

In one embodiment of the present invention, the above-mentioned extract was administered to an animal model in which osteoporosis was induced by menopause. As a result, it was confirmed that the bone density, bone volume and bone area of cancellous bone of distal femoral epiphysis were increased (Figs. 3 to 5 ). This suggests that the extract has the effect of increasing osteopenia by increasing the bone density, volume and area to make the bone hard and to activate the mineral metabolism of the bone tissue.

The term "osteoporosis" of the present invention means a state in which the amount of bone is decreased and the strength of bone is weakened due to a qualitative change, whereby fracture is likely to occur.

The term "prevention" of the present invention means all actions that inhibit or delay the onset of osteoporosis by administration of a pharmaceutical composition containing the extract of the present invention.

The term "treatment" of the present invention means any suspicion of osteoporosis by administration of the pharmaceutical composition, and any action that alters or alleviates symptoms of the onset.

The pharmaceutical composition of the present invention may contain the extract in an amount of 0.0001 to 50% by weight based on the weight of the total composition, and may include, but is not limited to, 0.01 to 10% by weight.

The pharmaceutical composition of the present invention may further comprise a pharmaceutically acceptable carrier, excipient or diluent conventionally used in the production of a pharmaceutical composition, and the carrier may include a non-naturally occuring carrier .

The term "pharmaceutically acceptable" of the present invention means that it exhibits properties that are not toxic to the cells or humans exposed to the composition.

Specifically, the pharmaceutical composition may be formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like, oral preparations, suppositories and sterilized injection solutions according to a conventional method . In the present invention, the carrier, excipient and diluent which may be contained in the pharmaceutical composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, Calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid formulations for oral administration include tablets, pills, powders, granules, capsules and the like, which may contain at least one excipient such as starch, calcium carbonate, sucrose or lactose lactose, gelatin, and the like. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid preparations for oral use may include various excipients such as wetting agents, sweetening agents, fragrances, preservatives, etc. in addition to water and liquid paraffin, which are simple diluents commonly used in suspension, liquid solutions, emulsions and syrups have. Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Examples of the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. As a base for suppositories, witepsol, macrogol, tween 61, cacao paper, laurin, glycerogelatin and the like can be used.

Another aspect of the present invention provides a method for preventing or treating osteoporosis, comprising administering the composition to a subject other than a human.

The term "administering" of the present invention means introducing a given substance into an individual in a suitable manner.

The term "individual" of the present invention means all animals such as rats, mice, livestock, etc., including humans who have developed or are capable of developing osteoporosis. As a specific example, it may be a mammal including a human.

The method for preventing or treating osteoporosis of the present invention may specifically include a step of administering a pharmaceutically effective amount of a pharmaceutical composition for prevention or treatment of osteoporosis comprising a soybean germinated embryo extract or a fraction thereof to a subject other than a human. have.

The term "pharmaceutically effective amount" of the present invention means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment and not causing side effects. The effective dose level is determined by the sex, age And other medical fields, including drugs used in combination or concurrently, with respect to body weight, health status, type of disease, severity, activity of the drug, sensitivity to the drug, method of administration, administration time, route of administration, Can be readily determined by those skilled in the art according to well known factors.

Specifically, the composition of the present invention may be administered at a dose of 0.0001 to 100 mg / kg body weight per day, more specifically 0.001 to 100 mg / kg body weight, based on the solid content. The administration may be carried out once per day, or divided into several doses.

In the method for preventing or treating osteoporosis of the present invention, the administration route and method of administering the composition are not particularly limited, and any route of administration and administration may be used as long as the composition containing the composition can reach the desired site It is possible to follow the method. Specifically, the composition may be administered orally or parenterally through various routes. Non-limiting examples of routes of administration include oral, rectal, topical, intravenous, intraperitoneal, intramuscular, intraarterial, transdermal, Intramuscularly or through inhalation or the like.

Another aspect of the present invention provides a food composition for preventing or ameliorating osteoporosis comprising a soybean germinated embryo extract or a fraction thereof.

The term "improvement" of the present invention means all the actions that osteoporosis is improved or changed by administration of the composition.

The term "food" of the present invention is intended to encompass all kinds of foods, such as meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gums, ice creams, Vitamin complex, health functional food, and health food, all of which include foods in a conventional sense.

The term "functional food" as used herein means the same term as "food for special health use" (FoSHU). In addition to nutrition, It means food. Here, the term "function (surname)" means that the structure and function of the human body have a beneficial effect for health use such as controlling nutrients or physiological action. The food of the present invention can be prepared by a method commonly used in the art and can be prepared by adding raw materials and ingredients which are conventionally added in the art. In addition, the formulations of the food can also be produced without restrictions as long as they are formulations recognized as food. The composition for food of the present invention can be manufactured in various forms, and unlike general pharmaceuticals, it has the advantage that there is no side effect that may occur when a drug is used for a long period of time, and is excellent in portability, Can be ingested as an adjuvant to promote the effect of preventing or improving osteoporosis.

The health food refers to a food having an active health promotion or promotion effect compared with a general food, and a health supplement food refers to a food for health assistance. In some cases, the terms health functional foods, health foods, and health supplements are used.

Specifically, the health functional food is a food prepared by adding the extract of the present invention to food materials such as beverage, tea, spice, gum and confectionery, or by encapsulation, powdering, suspension or the like, But it has the advantage that there is no side effect that can occur when the food is used as a raw material and long-term use of the drug.

Since the food composition of the present invention can be routinely ingested, a high effect can be expected for prevention or improvement of osteoporosis, so that it can be very usefully used.

The food composition may further comprise a physiologically acceptable carrier. The type of carrier is not particularly limited, and any carrier conventionally used in the art can be used.

In addition, the food composition may contain additional components that are commonly used in food compositions and can improve odor, taste, visual appearance, and the like. For example, vitamins A, C, D, E, B1, B2, B6, B12, niacin, biotin, folate, panthotenic acid and the like. In addition, it may include minerals such as zinc (Zn), iron (Fe), calcium (Ca), chromium (Cr), magnesium (Mg), manganese (Mn), copper (Cu) It may also include amino acids such as lysine, tryptophan, cysteine, valine, and the like.

In addition, the food composition may further contain antiseptic agents (such as potassium sorbate, sodium benzoate, salicylic acid, sodium dehydroacetate), bactericides (Sodium nitrite), bleach (sodium sulfite), seasoning (sodium MSG glutamate, etc.), sweeteners (dicin, cyclamate, saccharin, etc.), coloring agents , Sodium, etc.), perfume (vanillin, lactones, etc.), swelling agents (alum, potassium hydrogen D-tartrate), emulsifiers, thickeners (foams), encapsulating agents, gum bases, foam inhibitors, solvents, And may include food additives. The additives may be selected and used in appropriate amounts depending on the type of food.

The extract of the present invention can be added intact or used together with other food or food ingredients, and can be suitably used according to conventional methods. The amount of the active ingredient to be mixed can be suitably determined according to its intended use (prevention, health or therapeutic treatment). Generally, the food composition of the present invention may be added in an amount of not more than 50 parts by weight, specifically not more than 20 parts by weight, based on the food or beverage, when the food or drink is prepared. However, in case of long-term ingestion for health and hygiene purposes, the active ingredient may be contained in an amount not exceeding the above range and there is no problem in terms of safety.

As an example of the food composition of the present invention, it can be used as a health beverage composition. In this case, various flavors or natural carbohydrates can be added as an additional ingredient like ordinary beverages. The above-mentioned natural carbohydrates include monosaccharides such as glucose and fructose; Disaccharides such as maltose, sucrose; Polysaccharides such as dextrin, cyclodextrin; Xylitol, sorbitol, erythritol, and the like. Sweeteners include natural sweeteners such as tau Martin and stevia extract; Synthetic sweetening agents such as saccharin and aspartame, and the like can be used. The ratio of the natural carbohydrate may be generally about 0.01 to 0.04 g, specifically about 0.02 to 0.03 g per 100 mL of the health beverage composition of the present invention.

In addition to the above, the health beverage composition may contain various nutrients, vitamins, electrolytes, flavors, colorants, pectic acid, salts of pectic acid, alginic acid, salts of alginic acid, organic acid, protective colloid thickener, pH adjuster, stabilizer, Alcohols or carbonating agents, and the like. It may also contain flesh for the production of natural fruit juices, fruit juice drinks, or vegetable drinks. These components may be used independently or in combination. The proportion of such additives is not critical, but is generally selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the health beverage composition of the present invention.

The food composition of the present invention may be contained at various weight percentages as long as it can exhibit the effect of preventing or improving osteoporosis. Specifically, the food composition of the present invention may contain 0.00001 to 100% by weight or 0.01 to 80% by weight, But are not limited thereto.

Another aspect of the present invention provides a feed composition for preventing or ameliorating osteoporosis, which comprises a soybean germinated embryo extract or a fraction thereof.

The term "feed " of the present invention means any natural or artificial diet, single meal, or the like ingredients for feeding, ingesting, digesting, or suitable for the animal.

The feed may comprise a feed additive or supplementary feed.

The kind of the feed is not particularly limited, and feeds conventionally used in the art can be used. Non-limiting examples of such feeds include vegetable feeds such as cereals, muscle roots, food processing busines logistics, algae, fibers, pharmaceutical buses, oils, fats, pastes or grain by-products; Animal feeds such as proteins, inorganic substances, fats, oils, fats, oils, monocellular proteins, animal plankton or foods. These may be used alone or in combination of two or more.

The germinated germinated embryo extract of the present invention improves the proliferative and differentiating ability of osteoblasts and increases the bone density, bone volume and number of osteocytes, and thus can be effectively used for preventing or treating osteoporosis.

FIG. 1 is a graph showing the toxicity of an extract of soybean germinated embryo to osteoblasts. DMSO (dimethyl sulfoxide) - treated osteoblasts were used as controls and soybean germinated embryo extracts were treated at 100, 250 or 500 ㎍ / mL.
Fig. 2 is a graph showing the effect of soybean germinated embryo extract on osteoblast differentiation ability. DMSO - treated osteoblasts were used as controls and soybean germinated embryo extracts were treated at a concentration of 100 or 500 ㎍ / mL.
FIG. 3 is a graph showing the effect of soybean germinated embryo extract on bone density. Mice (menopausal group) that induced osteoporosis by surgically removing ovaries were used as a negative control group, and the mice were administered with 0.1 mg / kg of a germinated germ extract, which was used as a soybean embryo group.
4 is a graph showing the effect of the soybean germinated embryo extract on bone thickness. Mice (menopausal group) that induced osteoporosis by surgically removing ovaries were used as a negative control group, and the mice were administered with 0.1 mg / kg of a germinated germ extract, which was used as a soybean embryo group.
Fig. 5 is a graph showing the effect of soybean germinated embryo extract on bone cell number. Mice (menopausal group) that induced osteoporosis by surgically removing ovaries were used as a negative control group, and the mice were administered with 0.1 mg / kg of a germinated germ extract, which was used as a soybean embryo group.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the following examples are intended to illustrate the present invention, but the scope of the present invention is not limited thereto.

Production Example 1. Preparation of soybean germinated embryo extract

In order to prepare the soybean germinated embryo extract, the soybean germinated embryo prepared according to the previous study of the present inventor was used. Specifically, the soybean embryo was immersed in running water and germinated. After about 24 hours, the soybean germinated embryo was harvested, lyophilized and powdered to obtain a soybean germinated embryo powder. The resulting soybean germinated embryo powder was extracted by adding a fermented syrup, followed by filtration to obtain a liquid component, and the liquid component was freeze-dried to obtain a soybean germinated embryo extract.

Then, the soybean germinated embryo extract was applied to an UPLC (Ultra Performance Liquid Chromatography) QTOF-mass spectrometer to select major compounds.

As a result, it was confirmed that a large number of isoflavone compounds existed in the extract. Among them, compounds represented by the following formulas (1) and (2) were found to be novel isoflavone compounds. In addition, it was confirmed that a large number of soy sa saponins were present in the extract, and the soy sa saponins were summarized in Table 1 below.

[Chemical Formula 1]

(2)

Types of soya saponin contained in soybean germinated embryo extract compound
number Soya saponin compound name One Diacetyl line 2 Triacetyl soyasaponin Ab 3 Diacetyl linea papaina 4 Soyasaponin Ab 5 Soyasaponin Ae 6 Soyasaponin Af 7 Soyasaponin Aa 8 Soyasaponin Ac 9 Soyasaponin Ag 10 Soyasaponin Name 11 Soyasaponin Ah 12 Soyasaponin Ba 13 Soyasaponin Bc 14 Soyasaponin Bb 15 Soyasaponin Bd 16 Soyasaponin Be 17 Soyasaponin βg 18 Soyasaponin beta a 19 Soyasaponin γg

Example  1. Beans Germination embryo  Evaluation of cytotoxicity of extracts

To confirm the safety of the soybean germinated embryo extract obtained according to Preparation Example 1, the cytotoxicity of the extract to osteoblasts was evaluated. Specifically, the osteogenic cell line C3H10T1 / 2 was counted at a density of 2 × 10 ² cells / well in a 96-well plate, and the cells were treated with soybean germinated embryo extract at a concentration of 100, 250 and 500 μg / mL for 24 hours or 72 hours Lt; / RTI > After the completion of the incubation, the water-insoluble formazan formed by the addition of the MTT solution was dissolved in a dimethyl sulfoxide solution, and the optical density (OD value) of the dissolved formazan at 595 nm was measured. The relative cell viability (%) was assessed by measurement.

As a result, as shown in FIG. 1, the soybean germinated embryo extract did not show cytotoxicity even at the concentrations of 100, 250, and 500 ㎍ / mL, and it was confirmed that the osteoblast proliferation performance was rather improved at the concentration of 500 ㎍ / mL Respectively. As a result, the soybean germinated embryo extract of the present invention is a safe substance free from harm to human body, and promotes bone formation by activating the proliferation of osteoblasts.

Example  2. Soybean for osteoblast differentiation Germination embryo  Analysis of the effect of extracts

In order to confirm the bone formation promoting effect of the soybean germinated embryo, the effect of the extract on the osteoblast differentiation ability was analyzed.

Specifically, osteoblast C3H10T1 / 2 (Korean Cell Line Bank, Korea) was inoculated into DMEM supplemented with 10% FBS (fetal bovine serum) and 1% antibiotics (penicillin 100 U / ml and streptomycin 0.1 mg / (Dulbecco's modified eagle medium) by using the medium was incubated in the 96-well plate in 37 _{℃, 5% CO 2 (95%} air) conditions. The cells were seeded at a density of 3 × 10 ² cells / well, and the cells were treated with soybean germinated embryo extracts at a concentration of 100 and 500 μg / mL for 72 hours. After completion of cultivation, MTT solution was added to evaluate the relative cell division ability by measuring the degree of formation of water-insoluble formazan.

As a result, as shown in FIG. 2, it was confirmed that the test group treated with 100 and 500 ㎍ / mL of soybean germinated embryo extracts showed a stronger staining than that of the control group. Especially, 500 ㎍ / mL of soybean germinated embryo extract was treated The osteoblast differentiation degree was further improved. Thus, it can be seen that the soybean germinated embryo extract of the present invention has a great effect on the bone formation by differentiating osteoblasts.

Example  3. Soybean Germination embryo  Evaluation of efficacy of extracts for osteoporosis treatment

Example  3-1. Beans for bone density Germination embryo  Effect of extract

Through Examples 1 and 2, it was confirmed that the soybean germinated embryo extract of the present invention improved osteoblast proliferation and differentiation ability. In order to evaluate the therapeutic effect of the extract on osteoporosis, the effect of soybean germinated embryo extract on osteoporosis induced osteoporosis was analyzed.

Specifically, the ovaries were removed from the ovaries by using 8-week-old female C57BL / 6 mice. For ovariectomy, a 1-2 cm skin incision was made in the lower middle region of the abdomen and forceps were applied to the ovaries (ovary) and oviduct (oviduct). One week after ovariectomy, randomized grouping was divided into randomized groupings using body weight measurement. Sonde was orally administered with 0.1, 1 and 5 mg / kg of soybean germinated embryo extracts six times a week.

After the administration, the femur was removed and the surrounding muscle was removed and fixed with 10% formalin. The bone mineral density was measured by μCT analysis and the treatment effect on osteoporosis was evaluated by calculating the volume of cancellous bone with respect to the total area of the bone at distal femur.

As a result, as shown in FIG. 3, in the test group administered with 0.1 mg / kg of the extract, compared to the negative control group in which osteoporosis was caused by menopause, the bone volume density of the cancellous bone of the distal femur (%)) Was increased.

Example  3-2. Beans for bone thickness Germination embryo  Effect of extract

To evaluate the therapeutic effect of soybean germinated embryo extracts on osteoporosis, we analyzed the effect of soybean germinated embryo extracts on the bone volume of osteoporosis - induced animal models by surgically removing ovaries.

Specifically, the extract was administered to an osteoporosis-inducing animal model according to the method of Example 3-1. After the end of the administration, the separated femur was analyzed using μCT. Approximately 800 images were obtained from μCT images, and the sections were reconstructed using NRecon scanning software. Using a data viewer, the bone density was calculated by the weight of the cancellous bone per cm ³ by setting the region to be 3.5 mm away from the growth plate in each region of the femur.

As a result, as shown in FIG. 4, it was confirmed that the bone thickness of cancellous bone of distal femoral distal femur was increased in the experimental group to which 0.1 mg / kg of the extract was administered compared to the negative control group in which osteoporosis was induced by menopause.

Example  3-3. Soy beans for bone area Germination embryo  Effect of extract

To evaluate the therapeutic effect of soybean germinated embryo extracts on osteoporosis, we analyzed the effect of soybean germinated embryo extracts on the number of osteoblasts in osteoporosis - induced animal models by ovariectomy.

Specifically, the extract was administered to an osteoporosis-inducing animal model according to the method of Example 3-1. After the end of the administration, the femur was removed and peripheral muscle was removed, fixed with paraffin through 10% formalin fixation and EDTA demineralization, and the block was sectioned at 4 μm. After performing hematoxylin and eosin (HE) staining, the area of cancellous bone was measured using an image analysis program for the total area of distal femoral epiphysis observed at a magnification of 100 × under the microscope. Respectively.

As a result, as shown in FIG. 5, it was confirmed that the bone area of distal femoral tibia was increased in the experimental group to which 0.1 mg / kg of the extract was administered compared to the negative control group in which osteoporosis was caused by menopause.

From the above results, it can be seen that the soybean germinated embryo extract of the present invention has an effect of treating osteoporosis by increasing bone density, volume and area.

From the above description, it will be understood by those skilled in the art that the present invention may be embodied in other specific forms without departing from the spirit or essential characteristics thereof. In this regard, it should be understood that the above-described embodiments are to be considered in all respects as illustrative and not restrictive. The scope of the present invention should be construed as being included in the scope of the present invention without departing from the scope of the present invention as defined by the appended claims.

Claims (13)

A pharmaceutical composition for preventing or treating osteoporosis, comprising a soybean germinated embryo extract or a fraction thereof.
The composition according to claim 1, wherein the soybean germinated embryo is obtained by germination of an embryo isolated from soybean.
The composition according to claim 1, wherein the extract is obtained by extracting the soybean germinated embryo with water, a lower alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof.
The composition according to claim 1, wherein the fraction is obtained by fractionating the extract with water, a lower alcohol having 1 to 4 carbon atoms, a non-polar solvent, or a mixed solvent thereof.
The composition of claim 1, wherein the extract comprises isoflavone or soyasaponin.
6. The composition according to claim 5, wherein the isoflavone is selected from the following formula (1) or (2).
[Chemical Formula 1]

(2)

6. The method according to claim 5, wherein the soy saff saponin is selected from the group consisting of Triacetyl soyasaponin Ab, Diacetyl soyasaponin Aa, Soyasaponin Ab, Soyasaponin Ae, Soyasaponin Af, Soyasaponin Aa, Soyasaponin Ac, Soyasaponin Ag, Soyasaponin Ad, Soyasaponin Ah, Soyasaponin Ba, Soyasaponin Bc, Soyasaponin Bb , Soyasaponin Bd, Soyasaponin Be, Soyasaponin? G, Soyasaponin ?, and Soyasaponin? G.
The composition according to claim 1, wherein the extract enhances the proliferative and differentiating ability of osteoblasts.
2. The composition of claim 1, wherein the extract increases bone density, bone volume, and bone area.
2. The composition of claim 1, wherein the composition further comprises a pharmaceutically acceptable carrier, excipient or diluent.
11. A method for preventing or treating osteoporosis, comprising administering the composition of any one of claims 1 to 10 to a subject other than a human.
A food composition for preventing or improving osteoporosis, comprising a soybean germinated embryo extract or a fraction thereof.
A feed composition for preventing or ameliorating osteoporosis, comprising a soybean germinated embryo extract or a fraction thereof.

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