KR20050071101A - Composition comprising the extract of chongkukjang or the apigenin isolated therefrom having blood glucose-lowering activity - Google Patents

Abstract

The present invention relates to a composition for the treatment and prevention of hypoglycemia or diabetes containing the cheonggukjang extract and apigenin isolated therefrom as an active ingredient. Cheonggukjang extract of the present invention and the apigenin isolated from it inhibits the activity of the glycolytic enzyme alpha-glucosidase (α-glucosidase), thereby inhibiting the absorption of glucose in the body shows an excellent effect of controlling post-prandial blood sugar The composition containing can be used for medicines for the treatment and prevention of hypoglycemia or diabetes, and dietary supplements.

Description

Composition comprising the extract of Chongkukjang or the apigenin isolated therefrom having blood glucose-lowering activity}

The present invention relates to a composition for the treatment and prevention of hypoglycemia or diabetes containing the cheonggukjang extract or apigenin isolated therefrom as an active ingredient.

Diabetes is ranked 6th among Korean deaths according to 1998 death statistics and 4th among deaths according to 2001 death statistics, and the mortality rate from diabetes has increased by more than 125% over the last decade (Korea National Statistical Office, Annual Report of on the Cause of Death Statistics , Statistics Korea, 2002). The rapid increase in the prevalence of diabetes has been attributed to changes in lifestyle, including diet and exercise.

Diabetes is a chronic disease in which glucose in the blood is higher than normal people due to lack of insulin, which causes glucose to be excreted in the urine. Normally, most of the food you eat is converted to glucose, and the blood carries it to the cells that make up your body. Insulin (hormones) is needed to move glucose into cells. Diabetes is caused by the inability of glucose to move into cells because insulin is not made or working properly. Glucose that was not transported into the cells remains in the blood, so the blood becomes a high blood sugar 'high blood sugar' state and is discharged into the urine.

Diabetes is characterized by two types, depending on whether insulin is produced. Insulin dependent diabetes mellitus (IDDM) is caused by a lack of secretion of insulin, a glucose-regulating hormone in the blood. It is called juvenile diabetes because it occurs in young people in their 20s. Type II diabetes (non-insulin dependent diabetes, NIDDM) occurs mainly after the age of 40, and occupies most of the diabetic patients in Korea. Unlike type I, it is called adult-type diabetes and the cause of the disease is not known yet, but it is known to be caused by genetic factors and environmental factors. The etiology of type II diabetes has been observed in both insulin secretion in pancreatic beta cells and defects in insulin action (insulin resistance) in target cells, although it is not yet clear which changes are of primary importance.

Three therapies are necessary for the treatment of diabetes: drug, diet, and exercise therapy. To date, no clear etiology and cure for diabetes have been established. An important treatment goal for diabetes is to control blood glucose as close to normal as possible, to prevent complications, and to minimize the risk of complications.

Oral hypoglycemic drugs currently used by some of the type 1 diabetic patients and most type 2 diabetic patients to control blood sugar may have side effects such as bloating, vomiting, abdominal pain, and hypoglycemia, which may limit their use. Hanefeld M., J. Diabetes Complications , 12 , pp228-237, 1998). Therefore, there is an urgent need to develop hypoglycemic agents from natural products.

Cheonggukjang is a fermented soybean food traditionally consumed in Korea. Soybean, the main ingredient of Cheonggukjang, has a low glycemic index, which is effective for diabetics, and contains bioactive substances such as protein, dietary fiber, isoflavones, and phenols. Known as a good food (David, JA, et al., Am. J. Clin. Nutr. , 34 , pp362-366, 1981; Adlercreutz, H., et al., W. Ann. Med. , 29 , pp95 -120 1997). Cheonggukjang has the potential to produce new bioactive substances in the fermentation process as well as various bioactive substances in soybeans and is expected to help health (Kim, OH, et al., Food Industry and Nutrition). ., 4, pp40-46 1999).

Meanwhile, Touchi , a traditional Chinese fermented soybean Water Extracts to Type 2 Diabetic Animal Models and Diabetics It has been reported to reduce blood sugar levels after long-term intake, suggesting the possibility that soybean fermented foods have hypoglycemic effects (Fujita, H., et al., T. Life Science , 70 , pp 219-227, 2001; Fujita, H., et al., Nutrition Research, 23 , pp713-722, 2003).

Techniques related to Cheonggukjang include Patent Publication Nos. 2001-2980, 1999-70171, 1999-60113, 1993-11894, 1993-1812, 1991-7446, 1988-5879, Patent Publication Nos. 2001-18095, 1999-72468, 2001-25273, 2001-18094, 1997-64430, 1999-65689, 1990-12548 and 1998-8040 1994-10744, 1992-2675, 1993-2356 and 1996-16567. However, these technologies are all focused on eliminating off-flavor and improving physical properties.

Korean Patent Publication No. 2002-21178 relates to functional Cheonggukjang and its manufacturing method, and describes anti-cancer functional Cheonggukjang. Patent Publication No. 2003-59985 describes fermented milk for preventing adult diseases containing soybean fermented extract and its manufacturing method. And it discloses its serum cholesterol reduction and blood pressure lowering effect.

However, neither of these documents teaches or discloses that Cheonggukjang is used for hypoglycemic effects and diabetes.

Therefore, the researchers completed the present invention by confirming the excellent hypoglycemic effect of the cheonggukjang extract and apigenin separated from it while searching for the physiological activity of Cheonggukjang.

Accordingly, it is an object of the present invention to provide a composition for the treatment and prevention of hypoglycemia or diabetes containing the cheonggukjang extract and apigenin isolated therefrom as an active ingredient.

In order to achieve the above object, the present invention provides a pharmaceutical composition for treating and preventing hypoglycemia or diabetes mellitus containing a non-solvent soluble solvent extract or a polar solvent soluble extract as an active ingredient.

The chungkukjang non-polar solvent soluble extract comprises a non-polar solvent selected from hexane, dichloromethane, chloroform or ethyl acetate, preferably extracted with ethyl acetate, the chungkukjang polar solvent soluble extract is selected from acetone, butanol, ethanol, methanol or water Polar solvents, preferably those extracted with butanol.

The present invention also provides a composition for the treatment and prevention of hypoglycemia or diabetes containing apigenin isolated from the extract of Cheonggukjang as an active ingredient.

Hereinafter, the present invention will be described in detail.

Apigenin isolated from chungkukjang non-polar solvent soluble extract, polar solvent soluble extract and chunggukjang extract of the present invention can be obtained by preparing a crude extract to obtain a crude extract, and then fractionated into each solvent, as follows: To prepare the soybeans, the soybeans were washed and immersed in 3 to 5 times the volume of water for 12 to 24 hours, dehydrated for 10 minutes to 2 hours, and 100 to 150 ° C, preferably using a pressurized steamer. After steaming at 121 degreeC for 10 minutes-2 hours, Preferably it is 15 minutes, it cools so that temperature may be 50-70 degreeC. Cultured Bacillus subtilis ( Bacillus subtilis , SUN 816) The strain was inoculated at 1-3% by weight, preferably 2% by weight of soybean, fermented at 37 ° C. for 40 hours to 80 hours, preferably 72 hours, and additionally lyophilized to obtain Cheonggukjang. can do.

Cheonggukjang crude extract is a polar solvent of water and C ₁ to C ₄ lower alcohols such as methanol, ethanol, butanol, and the like, having a volume of water of about 2 to 20 times, preferably about 5 to 10 times the weight of the Cheonggukjang. Mixed solvent having a mixing ratio of 1: 0.1 to 1:10, preferably in methanol at about 10 to 100 ° C., preferably at about 15 to 40 ° C. for about 5 to 24 hours, preferably 8 to 15 hours Using an extraction method such as hot water extraction, cold needle extraction, reflux cooling extraction or ultrasonic extraction, preferably hot water extraction to recover the supernatant by vacuum filtration, and then repeating the above procedure 2 to 5 times. The obtained Chungcheongjang crude extract can be obtained by concentrating under reduced pressure, and the chungkukjang non-polar solvent soluble extract and polar solvent soluble extract can be prepared by suspending the above-mentioned Chungkukjang crude extract in water, followed by hexane and ethyl. Three Tate, butanol can be the order of the obtained extract soluble in soybean each solvent of the present invention using a solvent.

In addition, the apigenin isolated from the extract of Chunggukjang can be obtained by comparing and analyzing the activity of each of the solvent-soluble extracts of Cheonggukjang, and separating and purifying the most active ethyl acetate soluble extract in the following manner. The ethyl acetate soluble fraction is subjected to silica gel column chromatography, wherein silica gel (Kiesel gel Art. 7734, Merck Co. Germany) is used as a stationary phase, and chloroform: acetone is mixed as a developing solvent. After evaporation with a solvent, 4 ml per minute was eluted, and 150 ml were fractionated. Then, TLC (Thin Layer Chromatography, Thin Layer Chromatography) was developed, and fractions having similar _Rf values were collected, divided into 6 groups, and evaporated to dryness. TLC is performed again for the six fractions to identify the active substance, where the stationary phase Dropwise to the fractions on a silica gel plate (Kiesel gel 60 F ₂₅₄₎ for TLC to, and as a developing solvent chloroform: acetone (2: 1) was developed with a mixed solvent and color-developing the color-developing reagent, and then the alpha-glucosidase inhibition Fractions with high activity were collected and purified by silica gel column chromatography again until they appeared as single spots to separate three spots, and then each alpha glucosidase inhibitory activity was applied to the three spots. Apigenin can be separated and obtained by measuring the molecular structure of spot 1, which is the most active spot by irradiation, by using a nuclear magnetic resonance apparatus, and identifying the structure.

The present invention provides a pharmaceutical composition for the prevention and treatment of hypoglycemia or diabetes containing apigenin isolated from the chungkukjang nonpolar solvent soluble extract or the polar solvent soluble extract and the cheonggukjang extract obtained by the above method.

The effects of inhibiting the alpha glucosidase activity of the cheonggukjang extract and apigenin of the present invention to inhibit the glycosylase activity, it was confirmed that exhibits an effect of inhibiting the activity.

The present invention provides a pharmaceutical composition for the prevention or treatment of hypoglycemia or diabetes containing the apigenin isolated from the chungkukjang nonpolar solvent soluble extract or the polar solvent soluble extract and the cheonggukjang extract obtained by the above method.

The pharmaceutical composition for the prevention and treatment of hypoglycemia or diabetes of the present invention comprises 0.1 to 80% by weight of apigenin separated from the Cheonggukjang nonpolar solvent soluble extract or the polar solvent soluble extract and the Cheonggukjang extract based on the total weight of the composition.

The pharmaceutical composition for preventing and treating hypoglycemic or diabetic of the present invention may further include appropriate carriers, excipients and diluents commonly used in the preparation of pharmaceutical compositions.

In addition, the pharmaceutical composition of the present invention may further include synergism by further including a known substance or a new substance used to prevent and treat diabetes by inhibiting glucose absorption in the human body in addition to the extract of Cheonggukjang or apigenin isolated therefrom as necessary. Can be exercised.

Compositions comprising apigenin isolated from the chungkukjang nonpolar solvent soluble extract or the polar solvent soluble extract and the cheonggukjang extract according to the present invention, respectively, powders, granules, tablets, capsules, suspensions, emulsions, syrups, etc. Oral formulations, external preparations, suppositories and sterile injectable solutions can be formulated and used, and the carriers, excipients and diluents that may be included in the composition 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 include at least one excipient such as starch in the composition comprising the Cheonggukjang extract and apigenin isolated therefrom. , Calcium carbonate (calcium carbonate), sucrose (sucrose) or lactose (lactose), gelatin and the like are mixed and prepared. In addition to simple excipients, lubricants such as magnesium styrate and talc are also used. Liquid preparations for oral use may include various excipients, such as wetting agents, sweeteners, fragrances, preservatives, etc., in addition to water and liquid paraffin, which are commonly used to include suspensions, solutions, emulsions, and syrups. have. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, 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 pharmaceutical dosage forms of apigenin isolated from the Chungkookjang Nonpolar Solvent Soluble Extract or the Polar Solvent Soluble Extract and Cheonggukjang Extract of the present invention can be used alone or in combination with other pharmaceutically active fractions as well as in a suitable collection.

The amount of apigenin isolated from the chungkukjang nonpolar solvent soluble extract or the polar solvent soluble extract and the cheonggukjang extract of the present invention may vary depending on the age, sex and weight of the patient, but generally in an amount of 0.01 to 1000 mg / kg, preferably May be administered in an amount of 0.1 to 100 mg / kg divided once to several times daily. The dosage may also be increased or decreased depending on the patient's condition, including the route of administration, extent of disease, sex, weight, age, complications, and the like. In addition, the patient should be provided to the patient in consideration of the patient's factors such as the patient's lifestyle and pharmacological factors. Therefore, the above dosage does not limit the scope of the present invention in any aspect.

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

The present invention provides a health functional food comprising the Cheonggukjang non-polar solvent soluble extract or polar solvent soluble extract and food acceptable food additives exhibiting a preventive effect of hypoglycemia or diabetes. In addition, the present invention provides a health functional food comprising apigenin and a food acceptable food supplement additive isolated from the Cheonggukjang extract exhibiting a preventive effect of hypoglycemia or diabetes.

The health functional food of the present invention is preferably in the form of a pharmaceutical or health beverage such as powders, granules, tablets, capsules, pills, suspensions, emulsions, syrups, aerosols. Examples of the food to which the functional Cheonggukjang can be added include various foods, beverages, gums, teas, vitamin complexes, and health functional foods.

It may also be added to foods or beverages for the purpose of lowering blood sugar or preventing diabetes.

At this time, the amount of the Chunggukjang extract in the food or beverage and the apigenin separated therefrom is generally 0.1 to 80% by weight, preferably 1 to 50% by weight of the total food weight of the health functional food composition of the present invention. It can be added to, and may be added to the health beverage composition in a ratio of 1 to 30 g, preferably 3 to 10 g based on 100 ml.

The health beverage composition of the present invention has no particular limitation on the liquid component except for containing the above-mentioned Cheonggukjang as an essential ingredient in the indicated ratio, and may contain various flavors or natural carbohydrates, etc. as additional ingredients, like ordinary drinks. Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And conventional sugars such as polysaccharides such as dextrin, cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those mentioned above, natural flavoring agents (tauumatin, stevia Cheonggukjang (for example, rebaudioside A, glycyrrhizin, etc.), and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. The proportion of said natural carbohydrate 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, the composition of the present invention includes various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, coloring and neutralizing agents (such as cheese and chocolate), pectic acid and salts thereof, alginic acid and its Salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated beverages, and the like. The compositions of the present invention may also contain pulp for the production of natural fruit juices and fruit juice beverages 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 from the range of 0 to about 20 parts by weight per 100 parts by weight of the composition of the present invention.

Hereinafter, the present invention will be described in detail by way of examples.

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

Reference example. Cheonggukjang Ingredient Analysis

General components of Chungkookjang were analyzed by AOAC method. Moisture, crude protein, crude fat, crude ash and total dietary fiber content were measured. Moisture was analyzed by atmospheric pressure drying, crude fat by Soxhlet extraction, crude protein by semi-micro Kjeldhal method, and crude ash by incineration (Shin Hyo-sun. Food analysis (Theory and Experiment)) , Shinkwang Publishing Co., pp69-87, 1992). The total dietary fiber content was measured by AOAC method developed and modified by Prosky et al. (Prosky, L., et al., Journal-Association of Official Analytical Chemists , 71 , pp1017-1023, 1988).

Example 1. Preparation of Cheonggukjang

The soybeans used in the experiment were domestic soybeans. After washing the beans, soaked in water for 18 hours, soaked and drained and autoclave for 15 minutes at 121 ℃ using a pressure cooker, and then cooled to 60 ℃. The 2.4% nutrient agar medium is sterilized at 121 ° C. for 15 minutes and then placed in a petri-dish to solidify, followed by Bacillus subtillus , SUN 816) was plated for 4 minutes and stored for 24 hours in an incubator. 0.8% liquid medium was prepared and sterilized in a pressurized steamer at 121 ° C. for 15 minutes, cooled to 60 ° C., and the Bacillus subtilis, which had been plated for 4 minutes in a petri dish, was transferred to a liquid medium using platinum teeth. Incubated for hours. Bacillus subtilis cultured in a liquid medium was inoculated at 2% of the weight of soybean, fermented at 37 ° C. for 72 hours in an incubator, and the Chungkookjang of the present invention was obtained. . In addition, it was additionally lyophilized and powdered to determine its content.

The general contents of Cheonggukjang were 55.7% of moisture, 19.7% of protein, 8.3% of crude fat, 2.4% of crude ash, and 7.0% of total fiber. The lyophilized Cheonggukjang contained 8.6% water, 44.5% protein, 18.7% fat, 5.4% ash, 15.8% total fiber and 7.0% carbohydrate.

Example 2. Preparation of Cheonggukjang crude extract

30 kg of methanol was added to 3 kg of the Cheonggukjang powder prepared in Example 1, followed by extraction three times at 12 ° C. for 12 hours, followed by filtration under reduced pressure (Watman, USA), followed by filtration extract. After removing methanol using a silver vacuum rotary concentrator, 70 g of Chunggukjang methanol extracts were obtained as an extracted residue.

Example 3. Preparation of Cheonggukjang hexane soluble extract

70 g of the chungkukjang crude extract obtained in Example 2 was dissolved by adding hexane: methanol extract: water in a ratio of 10: 1: 9 and then shaking it in a fractional filter to extract it at about 25 ° C. for 12 hours. Repeatedly obtaining the water-soluble fraction and the hexane-soluble fraction layer, separated only from the components dissolved in the hexane-soluble fraction layer and filtered under reduced pressure to obtain 31.5 g of hexane-soluble extract was used as a sample for screening α-glucosidase inhibitory activity.

Example 4. Preparation of Cheonggukjang Ethyl Acetate Soluble Extract

To the Cheonggukjang water-soluble fraction obtained in Example 3, ethylacetate: water-soluble fraction: water was added at a ratio of 10: 1: 9 to dissolve, followed by shaking in a fractional filter to extract for 12 hours at about 25 ° C. Repeatedly obtaining the water-soluble fraction and the ethyl acetate soluble fraction layer, separating only the components dissolved in the ethyl acetate soluble fraction layer and filtration under reduced pressure to obtain 8 g of ethyl acetate extract to search for α-glucosidase inhibitory activity and isolate the active substance. Used as a sample for.

Example 5. Preparation of Chunggukjang Butanol Soluble Extract

The butanol: water soluble fraction: water was added to the Cheonggukjang water-soluble fraction obtained in Example 4 in a ratio of 10: 1: 9 to dissolve, followed by shaking three times to extract the mixture for about 12 hours at 25 ° C. To obtain the water-soluble fraction and butanol soluble fraction layer, butane was separated by filtration only the components dissolved in the soluble fraction layer to obtain 14.1 g of butanol extract was used as a sample for screening α-glucosidase inhibitory activity.

Example 6 Preparation of Cheonggukjang Water Soluble Extract

In Example 5, the remaining Cheonggukjang water-soluble extract except for the Cheonggukjang butanol soluble fraction layer was separated and filtered under reduced pressure to obtain 16.3 g of the Cheonggukjang water-soluble extract, which was used as a sample for screening α-glucosidase inhibitory activity.

Example 6. Isolation of Apigenin from Cheonggukjang Ethyl Acetate Soluble Extract

6-1. Perform primary chromatography, etc. of Cheonggukjang ethyl acetate soluble extract

After comparatively examining the α-glucosidase inhibitory activity of each of the solvent-soluble extracts of Cheonggukjang, about 8 g of the ethyl acetate soluble fraction of Example 4, which was found to have the highest activity, was subjected to silica gel column chromatography. ) Was performed. At this time, silica gel (Kiesel gel Art. 7734, Merck Co. Germany) was used as the stationary phase, and chloroform: acetone was mixed at 13 concentrations (v / v) as a developing solvent, and each solvent was eluted at 4 ml per minute. 150 ml of sugar was then fractionated and developed on TLC (Thin Layer Chromatography, Thin Layer Chromatography) to collect fractions with similar _Rf values, divided into 6 groups, and evaporated to dryness.

6-2. TLC, etc., performed on the Cheonggukjang chromatography fractions

TLC was performed on the six fractions obtained in Example 6-1 to identify the active substance. At this time, the fraction was deposited on a TLC silica gel plate (Kiesel gel 60 F ₂₅₄ ) as a stationary phase, and developed using a mixed solvent of chloroform: acetone (2: 1) as a developing solvent. Positioning of the active ingredient was carried out using a UV detector (DU ^R 650, Beckman, Germany), and was developed using amialdehyde (anisaldehyde, Sigma, USA) as a coloring reagent. Then, the fractions with high alpha glucosidase inhibitory activity were collected and purified by silica gel column chromatography again until they appeared as a single spot, and as a result, three spots were separated (see FIG. 1).

6-3. Identification of substances exhibiting alpha glucosidase inhibitory activity

The three spots of Example 6-2, each of the alpha glucosidase inhibitory activity was investigated (see Experimental Example 2 below), and then the molecular structure of the highest activity spot was measured using a nuclear magnetic resonance apparatus. Was identified. ¹ H-NMR and ¹³ C-NMR used in this experiment were DMSO- d ₆ as a solvent by JEFT's FT-NMR, and TMS (tetramethylsilane) was used as an internal standard.

As a result of measuring the alpha glucosidase inhibitory activity of the isolated spots, the inhibitory activity of spot 1 was the highest (57.3%), and the structure of the active ingredient was determined using ¹ H-NMR and ¹³ C-NMR. , Apigenin (see Figs. 2a and 2b).

Experimental Example 1. Alpha-glucosidase Inhibitory Activity of Soluble Extracts from Cheonggukjang

In order to measure the alpha-glucosidase inhibitory activity of each solvent soluble extract of Chunggukjang, samples of Cheonggukjang hexane soluble extract of Example 3, ethyl acetate soluble extract of Example 4, butanol soluble extract of Example 5 and water soluble extract of The experiment was carried out as follows.

The alpha-glucosidase inhibitory activity of each solvent soluble extract was measured according to the method of Watanabe et al. (Watanabe, J., et. Al., Biosci Biotechi Biochem 61, pp177-178 1997). The extract of each solvent fraction used in this experiment was dissolved in DMSO (dimethylsulfoxide) or distilled water at a concentration of 5 mg / mL, and 50 μl of alpha-glucosidase (Sigma, USA; 0.7 U / mL) and each solvent were used. 10 μl of the fractional extract sample was added to a well of a 96-well plate, and then OD ₄₀₅ was measured by a microplate reader (Model 550, Biorad, USA). After 5 minutes of substrate solution (5 mM para- nitrophenyl-α- D-glucopyranoside in 0.1 M phosphate buffer, pH 7.0) was added to 50 ㎕ and reaction at room temperature for 5 minutes, the OD ₄₀₅ The enzyme inhibitory activity was calculated from the change in absorbance.

As a result, as shown in Table 1, it was found that the alpha-glucosidase inhibitory activity was the highest at 33.3% in Cheonggukjang ethyl acetate soluble extract.

Soluble Extract of Solvent Inhibitory Activity (%) Hexane Soluble Extract (Example 3) - Ethyl Acetate Soluble Extract (Example 4) 33.3 Butanol Soluble Extract (Example 5) 16.0 Water Soluble Extract (Example 6) 9.1 Akabos (control, Bayer-Korea) 47.7

Experimental Example 2 Alpha-glucosidase Inhibitory Activity of Fractions Isolated from Cheonggukjang

Alpha-glucosidase inhibitory activity of the three spots (spot 1, spot 2 and spot 3) isolated in Example 6-2 was measured in the same manner as in Experimental Example 1.

As a result, as shown in Table 2 below, the inhibitory activity of spot 1 was the highest at 57.3%.

Inhibitory Activity (%) Spot 1 57.3 Spot 2 - Spot 3 - Akabos 47.7

Examples of the formulation of the composition are described below, but are not intended to limit the present invention but to explain in detail only.

Formulation Example 1 Preparation of Injection

Cheonggukjang Ethyl Acetate Soluble Extract of Example 4 ... 100 mg

Sodium metabisulfite ............ 3.0 mg

Methylparaben ...... 0.8 mg

Propylparaben ...................................... 0.1 mg

Sterile Distilled Water for Injection ...

The above ingredients are mixed and made into 2 ml by a conventional method, and then filled into 2 ml ampoules and sterilized to prepare an injection.

Formulation Example 2 Preparation of Tablet

Cheonggukjang ethyl acetate soluble extract of Example 4 ... 200 mg

Lactose ......................................... 100 mg

Starch ......................................... 100 mg

Magnesium Stearate ...............

The above components are mixed and tableted according to a conventional method for producing tablets to produce tablets.

Formulation Example 3 Preparation of Capsule

Fraction containing apigenin as a main component of Example 6-2 ... 100 mg

Lactose 50 mg

Starch ......................................... 50 mg

Talc ........................................ 2 mg

Magnesium Stearate ...............

Capsules are prepared by mixing the above ingredients and filling into gelatin capsules according to a conventional method for preparing capsules.

Formulation Example 4 Preparation of Liquid

Fraction containing apigenin as a main component of Example 6-2.

Sugar ........................................ 20 g

Isomerized sugar ......................................... 20 g

Lemon scent ..........................................

Purified water was added to adjust the total volume to 1000 ml. According to the conventional method for preparing a liquid, the above components are mixed, and then filled into a brown bottle and sterilized to prepare a liquid.

Formulation Example 5 Preparation of Healthy Food

Cheonggukjang ethyl acetate soluble extract of Example 4 .. 1000 mg

Vitamin Blend ...

Vitamin A Acetate ............... 70 μg

Vitamin E ......................................... 1.0 mg

Vitamin B1 .................................. 0.13 mg

Vitamin B2 ........................... 0.15 mg

Vitamin B6 ......................................... 0.5 mg

Vitamin B12 ......................... 0.2 μg

Vitamin C ......................................... 10 mg

Biotin ......................................... 10 μg

Nicotinic Acid Amide ... 1.7 mg

Folic acid ......................................... 50 μg

Calcium Pantothenate ......... 0.5 mg

Mineral mixture ........................

Ferrous Sulfate ............... 1.75 mg

Zinc Oxide ............... 0.82 mg

Magnesium Carbonate ... 25.3 mg

Potassium monophosphate ......................................... 15 mg

Dicalcium Phosphate Diluent ... 55 mg

Potassium Citrate ......................................... 90 mg

Calcium Carbonate ... 100 mg

Magnesium Chloride ......................................... 24.8 mg

Although the composition ratio of the above-mentioned vitamin and mineral mixtures is mixed with a component suitable for a health food in a preferred embodiment, the compounding ratio may be arbitrarily modified, and the above ingredients are mixed according to a conventional health food manufacturing method. The granules may be prepared and used for preparing a health food composition according to a conventional method.

Formulation Example 6 Preparation of Healthy Drink

Cheonggukjang ethyl acetate soluble extract of Example 4 .. 1000 mg

Citric Acid ......................................... 1000 mg

Oligosaccharide ......................................... 100 g

Plum concentrate ........................................ 2 g

Taurine ......................................... 1 g

Purified water is added ................... 900 ml

After mixing the above components in accordance with the conventional healthy beverage production method, and stirred and heated at 85 ℃ for about 1 hour, the resulting solution is filtered and obtained in a sterilized 2 L container, sealed sterilized and stored in the refrigerator and then Used to prepare the healthy beverage composition of the invention.

Although the composition ratio is mixed with a relatively suitable component for a preferred beverage in a preferred embodiment, the compounding ratio may be arbitrarily modified according to regional and ethnic preferences such as demand hierarchy, demand country, and usage.

Cheonggukjang extract of the present invention and apigenin isolated therefrom have an effect of inhibiting the activity of the glycolytic enzyme alpha-glucosidase excellently, so that the composition containing the drug and health function for the treatment and prevention of hypoglycemia or diabetes Can be used for food.

1 is a TLC result diagram of a fraction obtained by performing chromatography on Cheonggukjang ethyl acetate soluble extract,

Figure 2a is a diagram of the ¹ H-NMR spectrum of the spot 1 separated from the Chunggukjang extract, Figure 2b is a diagram of the ³ C-NMR spectrum of the spot 1 separated from the Cheonggukjang extract.

Claims (8)

Cheonggukjang Non-polar solvent soluble extract or polar solvent soluble extract containing a pharmaceutical composition for the treatment and prevention of hypoglycemia or diabetes. The pharmaceutical composition of claim 1, wherein the chungkukjang nonpolar solvent soluble extract is extracted with ethyl acetate. The pharmaceutical composition of claim 1, wherein the chungkukjang polar solvent soluble extract is extracted with butanol. A pharmaceutical composition for the treatment and prevention of hypoglycemia or diabetes containing apigenin isolated from the extract of Cheonggukjang as an active ingredient. The pharmaceutical composition according to claim 1 or 4, comprising 0.01 to 80% by weight of the Cheonggukjang nonpolar solvent soluble extract of claim 1, the polar solvent soluble extract, or the apigenin of claim 4 based on the total weight of the composition. A health functional food containing the Cheonggukjang nonpolar solvent soluble extract or polar solvent soluble extract of claim 1 and a food supplement acceptable food additive which exhibit a prophylactic effect of hypoglycemia or diabetes. A dietary supplement containing apigenin and food-acceptable food supplements that have a hypoglycemic or diabetic effect. The health functional food of Claim 6 or 7 which is a health drink.