KR101949023B1 - A Food Composition for Weight Gain which has Improved Gastrointestinal Enzyme Ectivity and Gastrointestinal Protective Effect - Google Patents

Abstract

The present invention relates to a food composition for weight gain, which comprises a fruit extract and a mushroom extract, and the composition for weight gain comprising the extract of the fruit of the present invention and the mushroom extract of the present invention improves the activity of the digestive enzyme It has the effect of protecting the stomach, and can be useful for solving low body weight.

Description

[0001] The present invention relates to a food composition for enhancing digestive enzyme activity and having a gastrointestinal protective effect,

The present invention relates to a food composition for weight gain comprising fruit extract and shiitake mushroom extract.

Many modern people are threatened by obesity due to industrialization and changes in eating habits, but there are also many people who worry about underweight. People who are underweight have a weakened immune system and are susceptible to infectious diseases such as tuberculosis, hepatitis and cold, and increase the risk of anemia, osteoporosis, reproductive problems and malnutrition. Especially, low birth weight in childhood can cause growth disturbance, affects secondary puberty, and causes problems in bone maturity, which adversely affects key growth. In addition, since the mortality rate increases as the age increases, maintaining a normal weight corresponding to the physique is as important as the obesity problem.

To increase your weight, you should choose foods that contain calories and nutrients rather than high-calorie foods filled with sugar and saturated fats, and include foods that are full of protein such as lean meat, fish, egg whites, beans, tofu, and dairy products . Proteins are needed for bone health, immune system enhancement, and recovery from disease. However, since it takes a lot of time and effort for the general public to take these foods, and it is difficult to implement them, weight control foods are widely used.

Weight control foods are foods that contain nutrients such as vitamins or minerals and are used to control calories to replace some or all of the diet for those who need weight control. These weight control foods include weight loss foods and food supplements to help with weight gain. Weight gain supplements include protein supplements to increase muscle, protein and carbohydrate supplements to increase body weight and muscle together, and carbohydrate supplements to increase body weight.

However, these supplements can cause kidney disease in protein supplements because the specific nutrient ratio is too high, and carbohydrate supplements can cause side effects such as diarrhea, indigestion and abdominal distension. Therefore, there is a need for a food for weight gain which is safe and easy to use while having few side effects.

According to Herbal Gram, a quarterly publication published by the American Botanical Council (ABC), the natural supplements market is estimated to be worth more than $ 5.2 billion in 2010, up 3.3 percent from 2009, Herbal dietary supplements Sales are growing steadily, so we need to actively respond to this.

Accordingly, the inventors of the present invention have completed the present invention by confirming that the multifunctional food for controlling body weight using the local special products such as cucurbitan fruit extract and shiitake mushroom extract has not only a simple weight gain effect but also a digestive enzyme activity improvement and gastrointestinal protective effect.

Korean Patent Publication No. 10-2016-0068014

It is an object of the present invention to provide a food composition for weight gain comprising fruit extract and shiitake mushroom extract.

Another object of the present invention is to provide a functional food for improving digestive enzyme activity or a special nutritional food comprising a food composition for weight gain comprising a fruit extract and a mushroom extract.

It is still another object of the present invention to provide a functional food for gastrointestinal protection or a special nutrition food comprising a food composition for weight gain comprising a fruit extract and a mushroom extract.

It is another object of the present invention to provide an antioxidant food comprising a food composition for weight gain comprising a fruit extract and a mushroom extract.

Another object of the present invention is to provide a pharmaceutical composition for the treatment of inflammation, which comprises a food composition for weight gain comprising a fruit extract and a mushroom extract.

An aspect of the present invention provides a food composition for weight gain comprising a fruit extract and a mushroom extract.

The food composition of the present invention can be prepared by a method commonly used in the art, and the extract according to the present invention can be obtained as follows. Rinse fruit and shiitake mushrooms with water to remove foreign matter, then dry and shred in shade. Cucumber mushroom and shiitake mushroom can be used without restrictions such as grown or sold. The appropriate amount of solvent is added to the crushed coconut fruit and the powder of mushroom to be completely immersed. The mixed powder of cucurbit fruit and shiitake can be extracted using a usual extraction solvent. Preferably, (a) anhydrous or hydrated alcohol having 1 to 4 carbon atoms (e.g. methanol, ethanol, propanol, butanol, Propanol, iso-propanol and n-butanol), (b) a mixed solvent of the lower alcohol and water, (c) acetone, (d) ethyl acetate, (e) chloroform, (G) hexane, (h) diethyl ether, (i) butyl acetate or (j) water, and most preferably water. The extract may be impregnated or warmed at room temperature. As the extraction method, any one of the methods such as hot water extraction, alcohol extraction, cold extraction, reflux cooling, solvent extraction, steam distillation, ultrasonic extraction, elution and compression can be used. Preferably, It can be alcohol extraction. The extract may be filtered and concentrated to obtain a final extract. Preferably, the extract is lyophilized to obtain the fruit of the present invention and the mushroom extract of the present invention.

The food composition of the present invention can be prepared by adding raw materials and ingredients which are conventionally added in the art, and it is also possible to use various flavors or fragrances as well as ordinary food compositions, Natural carbohydrates and the like as additional components.

Such natural carbohydrates include monosaccharides such as glucose, fructose and the like; disaccharides such as maltose, sucrose and the like; And polysaccharides, for example, conventional sugars such as dextrin, cyclodextrin and the like, and sugar alcohols such as xylitol, sorbitol and erythritol. The flavoring agents may be natural flavoring agents (tau martin), stevia extracts (e.g., rebaudioside A and glycyrrhizin), and synthetic flavors (saccharine, aspartame, etc.).

The food composition of the present invention may be formulated into a food composition containing at least one pharmaceutically acceptable or pharmaceutically acceptable carrier in addition to the above-described effective ingredients. The form of the food composition may be a tablet, a capsule, a powder, a granule, a liquid, a ring, a liquid, a syrup, a juice, a suspension, an oil, or a drip agent. For example, for formulation into tablets or capsules, the active ingredient may be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, and the like.

The food composition of the present invention is a vitamin mixture consisting of vitamin A acetate, vitamin E, vitamin B1, vitamin B2, vitamin B6, vitamin B12, vitamin C, biotin, nicotinamide, folic acid, calcium pantothenate; And one or more minerals that can be conventionally added in the art, such as ferrous sulfate, zinc oxide, magnesium carbonate, potassium monophosphate, potassium diborate, potassium citrate, calcium carbonate, and magnesium chloride.

If necessary, suitable binders, lubricants, disintegrants and coloring agents may also be included as a mixture. Suitable binders include natural and synthetic gums such as starch, gelatin, glucose or beta-lactose, natural sugars such as corn sweeteners, acacia, tracker candles or sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, Sodium chloride, and the like. Disintegrants may include starch, methylcellulose, agar, bentonite, xanthan gum, and the like.

The food composition of the present invention formulated in the above-described manner can be used as a functional food or added to various foods. Foods to which the composition of the present invention can be added include functional foods such as nutritional supplements, weight control foods and health supplements, tea drinks, beverages, meat, chocolates, foods, confectionery, pizza, ramen noodles, other noodles, Alcoholic beverages, vitamin complexes, and the like.

In addition, the above-mentioned food composition may contain various kinds of nutrients, vitamins, minerals (electrolytes), synthetic flavors and flavors such as natural flavors, coloring agents and aging agents (cheese, chocolate, etc.) And salts thereof, alginic acid and salts thereof, organic acids, protective colloid thickening agents, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols and the like.

According to one embodiment of the present invention, the composition may further comprise a soybean protein isolate, fructose fructose powder, and casein sodium.

According to one embodiment of the present invention, the composition may be a food composition for weight gain, which further comprises cocoa powder, potassium chloride, cookie right-handed, silicon dioxide, honey powder and vitamin powder.

According to one embodiment of the present invention, 15 to 60 parts by weight of isolated soybean protein, 5 to 20 parts by weight of crystalline fructose powder, 4 to 16 parts by weight of cocoa powder, 3.5 to 14 parts by weight of coconut fruit extract, 2 to 8 parts by weight of a mushroom extract, 5 to 20 parts by weight of casein sodium, 1.5 to 6 parts by weight of a cookie powder, 1 to 4 parts by weight of a creamy powder, 0.5 to 4 parts by weight of silicon dioxide, And 2 to 8 parts by weight of a vitamin powder.

Another aspect of the present invention provides a functional food for improving digestive enzyme activity, or a special nutrition food, comprising a food composition for weight gain comprising a fruit extract and a mushroom extract.

The improvement of digestive enzyme activity means that the digestive enzyme secreted by the ingestion of the food shows optimal activity according to the respective secretory organs and functions thereof or that the food composition of the present invention shows digestive enzyme- Which means that the degradation and absorption of nutrients is smooth.

The functional food or the special nutritional food of the present invention is a food or a food having a biological control function such as prevention and treatment of a disease, defense of a living body, immunity, recovery of a disease after a disease, suppression of aging, a special such as a child, infirm, sick, obese, Refers to foods intended to provide nutrients to a specific subject in need of nutritional care or to supplement nutrients to food ingredients for the purpose of substituting for food or to supplement nutrients that may be lacking in daily diets, It means harmless to human body.

In order to use the food composition of the present invention, which comprises the extract of Lycoris chejuensis and shiitake mushroom, as a functional food or a special nutrition food for improving digestive enzyme activity as described above, it may be prepared by various methods known in the field of food science or pharmacy And may be prepared in any food form that can be ingested orally by mixing with itself or with a pharmaceutically acceptable carrier, excipient, diluent or the like.

The functional food or special nutrition food of the present invention may further include food additive which is conventionally added in food manufacture and is pharmaceutically acceptable.

Another aspect of the present invention provides a functional food for gastrointestinal protection or a special nutrition food comprising a food composition for weight gain comprising a fruit extract and a mushroom extract.

The gastrointestinal protection of the present invention means to facilitate digestion of food, decomposition of nutrients, absorption of nutrients or defecation by alleviating gastrointestinal diseases. Gastrointestinal disorders are used in a broad sense to include all diseases and conditions that are accompanied by inflammation or damage to the gastrointestinal mucosa, or that may cause damage, irrespective of its etiology and mechanism. The gastrointestinal disorder may be selected from the group consisting of chronic gastritis such as acute gastritis, chronic active gastritis, chronic atrophic gastritis, duodenitis, gastric ulcer, duodenal ulcer, gastric perforation, gastric hemorrhage, digestive disorders associated with nonsteroidal antiinflammatory drug (NSAID) Streptococcal pylori infection, Zollinger-Ellison syndrome (ZES), Werner's syndrome, and systemic mastocytosis are the most common complications of gastroesophageal reflux disease (GERD) (reflux esophagitis), acute upper gastrointestinal bleeding, stress ulcerations, And erosion of the gastric mucosa due to the diseases, bleeding, redness, swelling and constipation or diarrhea.

Yet another aspect of the present invention provides an antioxidant food comprising a food composition for weight gain comprising a fruit extract and a mushroom extract.

The antioxidant food provided by the present invention means a food which can be used to prevent, treat or ameliorate a variety of diseases caused by excessive active oxygen, or by the active oxygen or, consequently, by causing excessive active oxygen . The diseases associated with active oxygen include, but are not limited to, degenerative neurological diseases including myocardial infarction, angina pectoris, coronary artery disease, ischemic heart disease, arteriosclerosis, Lou Gehrig's disease, Parkinson's disease, Alzheimer's disease, proximal axillary sclerosis and Huntington's disease May include various diseases such as cardiovascular diseases including ischemic brain diseases including stroke, diabetes, gastrointestinal diseases including gastritis and gastric cancer, aging, cancer, leukemia, aging, rheumatoid arthritis, hepatitis, atopic dermatitis, Preferably a gastrointestinal disorder caused by active oxygen.

The antioxidant food may be a functional food for preventing or ameliorating diseases caused by active oxygen. A processed food which can prevent or improve a disease caused by active oxygen can be produced by a method commonly used in the art. As a specific example, such processed foods include confectionery, drinks, liquor, fermented foods, canned foods, processed milk products, processed foods, noodles, and mushrooms. The sweets may be biscuits, pies, cakes, breads, candies, jellies, gums, cereals (including dinnerware items such as cereal flakes) or nutritional supplement products. The beverage can be drinking water, carbonated beverage, functional ionic beverage, juice (for example, apple, pear, grape, aloe, citrus, peach, carrot, tomato juice, etc.) or sikhye. The mainstream may be sake, whiskey, shochu, beer, liquor or fruit wine. The fermented food may be soy sauce, miso or kochujang. Canned products can be canned products (eg, tuna, mackerel, saury, canned fish), canned products (beef, pork, chicken, canned turkey, etc.) or agricultural products (corn, peach, canned pineapple etc). Milk processed foods can be cheese, butter or yogurt. Meat processed foods are pork cutlet, pork cutlet, chicken cutlet, sausage. Sweet and sour pork, nuggets, nuggets, and the like. Sealed noodles, and the like. In addition, the composition may be a mucilage product such as alginic acid mucilage, blue mushroom mushroom, acorn mucilage, omucum mushroom, mushroom mushroom, mushroom mushroom, mecum mushroom, and mushroom mushroom.

Yet another aspect of the present invention provides a pharmaceutical composition for the treatment of inflammation comprising a food composition for weight gain comprising a fruit extract and a shiitake mushroom extract.

When the composition of the present invention is prepared from a pharmaceutical composition, the pharmaceutical composition of the present invention includes a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier to be contained in the pharmaceutical composition of the present invention is one which is usually used in the present invention and includes lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia rubber, calcium phosphate, , Calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil, But is not limited thereto. The pharmaceutical composition of the present invention may further contain a lubricant, a wetting agent, a sweetening agent, a flavoring agent, an emulsifying agent, a suspending agent, a preservative, etc., in addition to the above components. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington ' s Pharmaceutical Sciences (22th ed., 2013).

The pharmaceutical composition of the present invention can be administered orally or parenterally, and is preferably administered orally.

The appropriate dosage of the pharmaceutical composition of the present invention may vary depending on such factors as formulation method, administration method, age, body weight, sex, pathological condition, food, administration time, route of administration, excretion rate, . A preferred dosage of the pharmaceutical composition of the present invention is within the range of 0.001-1000 mg / kg on an adult basis.

The pharmaceutical composition of the present invention may be formulated into a unit dosage form by using a pharmaceutically acceptable carrier and / or excipient according to a method which can be easily carried out by those having ordinary skill in the art to which the present invention belongs. Or by intrusion into a multi-dose container. The formulations may be in the form of solutions, suspensions, syrups or emulsions in oils or aqueous media, or in the form of excipients, powders, powders, granules, tablets or capsules, and may additionally contain dispersing or stabilizing agents.

According to the food composition for weight gain of the present invention comprising the extract of Fructus fruit and the extract of shiitake mushroom of the present invention, it has an effect of improving digestive enzyme activity and protecting stomach, and can be usefully used for solving low body weight.

FIG. 1 is a graph showing DPPH free radical scavenging activity (FIG. 1A) and IC50 analysis (FIG. 1B) according to the concentration of mixed extract of shiitake mushroom and cilantro powder.
Fig. 2 shows the effect of the concentration of the extracts of mushroom water extract (Fig. 2A), ethanolic extract extract (Fig. 2B), cucumber fruit juice extract (Fig. 2C) and cucumber fruit juice extract DPPH free radical scavenging activity.
FIG. 3 is a graph showing hydroxyl radical scavenging activity depending on the concentrations of the mushroom hot water extract (FIG. 3A), the mushroom spirulina extract (FIG. 3B), the cucumber mushroom hot water extract (FIG. 3C) and the cucumber mushroom extract .
FIG. 4 is a graph showing the Fe ^{2 +} chelating assay according to the concentrations of the mushroom hot water extract (FIG. 4A), the shiitake mushroom extract (FIG. 4B), the cucumber mushroom hot water extract (FIG. 4C) to be.
FIG. 5 is a graph showing nitrite scavenging activity according to the concentration of hot water extract of shiitake mushroom in pH 1.2 (FIG. 5A), pH 3.0 (FIG. 5B) and pH 5.0 (FIG. 5C).
FIG. 6 is a graph showing the nitrite scavenging activity according to the concentration of the extract of shiitake mushroom in pH 1.2 (FIG. 6A), pH 3.0 (FIG. 6B) and pH 5.0 (FIG. 6C).
FIG. 7 is a graph showing the nitrite scavenging activity according to the concentration of hot water extract of Codonopsis lanceolata at pH 1.2 (FIG. 7A), pH 3.0 (FIG. 7B) and pH 5.0 (FIG. 7C)
FIG. 8 is a graph showing nitrite scavenging activity according to the concentration of the extract of Fructus vulgaris L. in the pH 1.2 (FIG. 8A), pH 3.0 (FIG. 8B) and pH 5.0 (FIG. 8C)
FIG. 9 is a graph showing the OD ₅₅₀ absorbance (FIG. 9A) and the α-Amylase-like activity (FIG. 9B) depending on the concentration of a mixture of a mushroom hot water extract, 1% starch and 0.1% 3,5-dinitrosalicylic acid (DNS).
10 is a graph showing the OD ₅₅₀ absorbance (FIG. 10A) and the α-Amylase-like activity (FIG. 10B) depending on the concentration of the mushroom extract of the mushroom, 1% starch and 0.1% DNS mixture.
11 is a graph showing the OD ₅₅₀ absorbance (FIG. 11A) and the α-Amylase-like activity (FIG. 11B) depending on the concentration of the hot water extract of Cilantom fruit, 1% starch and 0.1% DNS.
12 is a graph showing the OD ₅₅₀ absorbance (FIG. 12A) and the α-Amylase-like activity (FIG. 12B) according to the concentration of the extract of Cilantomaceae fruit, 1% starch and 0.1% DNS.
FIG. 13 is a graph showing OD ₄₀₅ absorbance (FIG. 13A) and α-Glucosidase-like activity (FIG. 13B) depending on the concentration of a mixture of 3 m ρ-nitrophenyl-α-D-glucopyranoside (ρNPG) and shiitake hot water extract.
14 is a graph showing the OD ₄₀₅ absorbance (FIG. 14A) and the? -Glucosidase-like activity (FIG. 14B) depending on the concentrations of the mushroom extract and the 3 mM ρNPG mixture.
15 is a graph showing the OD ₄₀₅ absorbance (FIG. 15A) and the? -Glucosidase-like activity (FIG. 15B) depending on the concentration of the hot water extract and the 3 mM ρNPG mixture.
FIG. 16 is a graph showing OD ₄₀₅ absorbance (FIG. 16A) and? -Glucosidase-like activity (FIG. 16B) depending on the concentration of the extract of cilantomaceae fruit and 3 mM ρNPG mixture.
FIG. 17 is a graph showing the survival rate and cytotoxicity of RAW264.7 cells (FIG. 17A), RBL-2H3 cells (FIG. 17B) and AGS cells (FIG. 17C) according to the concentration of mushroom extract.
FIG. 18 is a graph showing the survival rate and cytotoxicity of RAW264.7 cells (FIG. 18A), RBL-2H3 cells (FIG. 18B) and AGS cells (FIG. 18C)
FIG. 19 is a graph showing inhibitory effect of NO production on RAW 264.7 cells according to the concentrations of mushroom extract (FIG. 19A), cilantro fruit extract (FIG. 19B), and mushroom and cucumber mixed extract (FIG. 19C).
FIG. 20 is a graph showing the inhibitory effect of β-hexosaminidase secretion according to the concentration of mushroom extract (FIG. 20A), cilantro fruit extract (FIG. 20B), and shiitake mushroom mixed extract (FIG. 20C) in RBL-2H3 cells.
FIG. 21 is a graph showing the effect of inhibiting histamine secretion according to the concentration of mushroom extract (FIG. 21A), cilantro fruit extract (FIG. 21B), and shiitake mushroom mixed extract (FIG. 21C) in RBL-2H3 cells.
FIG. 22 is a graph showing the effect of promoting the secretion of prostaglandin E2 (PGE2) according to the concentration of mushroom extract (FIG. 22A), cilantro fruit extract (FIG. 22B) and shiitake mushroom to be.
23 is a graph showing the inhibitory effect of TNF-α secretion on the RAW 264.7 cells according to the concentration of mushroom extract (FIG. 23A), cucumber fruit extract (FIG. 23B), and shiitake mushroom mixed extract (FIG. 23C).
24 is a graph showing the inhibitory effect of TNF-a on secretion of mushroom extract (Fig. 24A), cilantro fruit extract (Fig. 24B), and shiitake mushroom mixed extract (Fig. 24C) in RBL-2H3 cells.
25 is a graph showing the mean body weight (FIG. 25A), the body weight gain (FIG. 25B) and the body weight gain (FIG. 25C) of the control group and the experimental group in an animal model experiment for analyzing the weight control effect of the nutritional shake.
FIG. 26 is a graph showing average daily gain (FIG. 26A), average daily intake (FIG. 26B), and dietary efficiency (FIG. 26C) in the control group and the experimental group in an animal model experiment for analysis of weight control effect of nutritional shake.
27 is a graph showing the total length (Fig. 27A), the electric field increase rate (Fig. 27B) and the electric field increase rate (Fig. 27C) of the control group versus the control group and the control group in the animal model experiment for analyzing the weight control effect of the nutrient shake.
FIG. 28 is a graph showing the content of the muscle protein in the fuzzy muscle of the control group and the experimental group in the animal model experiment for the analysis of the weight control effect of the nutritional shake.
29 is a graph showing the backfat thickness (FIG. 29A) and the mean visceral fat weight (FIG. 29B) of the control and experimental groups in an animal model experiment for analyzing the weight control effect of the nutritional shake.
Figure 30 shows the total cholesterol levels (Figure 30A), triglyceride levels (Figure 30B), HDL levels (Figure 30C), LDL levels (Figure 30D), and triglyceride levels of control and experimental groups in animal model experiments for nutritional shake- VLDL < / RTI > values (Figure 30E).
FIG. 31 shows the results of an animal model experiment for analysis of weight control effect of a nutritional shake. FIG. 31A shows the arterial hardening index (FIG. 31A), arterial hardening correlation coefficient (FIG. 31B), heart risk index (FIG. 31C) and coronary artery index 31D.

Hereinafter, one or more embodiments will be described in more detail by way of examples. However, these embodiments are intended to illustrate one or more embodiments, and the scope of the present invention is not limited to these embodiments.

Example  1. For weight gain Nutritional shake  Produce

Nutritional shakes for weight gain were prepared as follows at the blending ratios shown in Table 1 below.

15 parts by weight of crystalline fructose powder, 5 parts by weight of vitamin powder, 3 parts by weight of honey powder, 3 parts by weight of cookie right powder, 2 parts by weight of creamy flavor powder, 1 part by weight of potassium chloride and 1 part by weight of silicon dioxide, lt; RTI ID = 0.0 > (mesh) < / RTI > Next, 37 parts by weight of isolated soybean protein, 11 parts by weight of sodium caseinate, 9 parts by weight of cocoa powder, 8 parts by weight of cocoa powder and 5 parts by weight of mushroom powder were added to the homogenized mixture, Korea) for 60 minutes to finally prepare a nutrition shake.

Nutritional shake composition content Weight portion Isolated soy protein 37.0% 37 Crystalline fructose powder 15.0% 15 Cocoa powder 9.0% 9 Cucumber powder extract powder 8.0% 8 Shiitake mushroom powder 5.0% 5 Casein sodium (milk) 11.0% 11 Potassium chloride 1.0% One Cookies also powdered right 3.0% 3 Cream-flavored powder 2.0% 2 Silicon dioxide 1.0% One Honey powder 3.0% 3 Vitamin powder 5.0% 5 sum 100% 100

Example  2. CJ  Active substance analysis of fruit extract

≪ 2-1 >

In order to analyze the active substances of fruit extracts of Cajunpara, cryopreserved fruit freeze - dried powder samples were prepared and analyzed by the Jangheung - gun Mushroom Industry Research Institute. For the analysis, the HPLC equipment was the Agilent Technologies 1200 series and the column was Agilent ZORBAX Eclipse XDB (4.6 X 150 mm, 5-micron). The specific conditions for the analysis are shown in the table below.

Analysis item Analysis condition Column Temperature 40 ° C / 30 ° C / 30 ° C Wavelength UV 350 nm / UV 355 nm / UV 210 nm Solvent 0.05% Phosphoric acid: Acetonitrile: Methanol: Water (70: 12: 12: 6) /
2.5% Acetic acid: Acetonitrile (55: 45) /
_{25mM KH 2 PO 4 (pH 2.5} ) Flow rate 1.0 ml / min Injection volume 20 쨉 l

<2-2> CJ  Active substance of fruit extract

As a result of analyzing the active substance of the fruit extract of Cucurbitaceae according to the above Example 2-1, 2.19 ± 0.16 mg of cetin, 7.69 ± 0.41 mg of rutin and 1.47 ± 0.13 mg of gallic acid were contained Respectively.

Example  3. Evaluation of antioxidant efficacy

<3-1> Evaluation method of antioxidant efficacy

3-1-1: Preparation of extract

Composite materials of shiitake mushroom and cucumber mushroom were prepared by mixing and extracting shiitake mushroom and cucumber mushroom 1: 1.

Shiitake mushroom and cilantrofruit were prepared by extracting with water or ethanolic extract as follows.

In the hot water extraction method, the fruit juice or shiitake mushrooms were treated with sterilized water for 5 hours at 100 ° C for 3 hours, and the extract solution was freeze-dried. The dried weight of the extract solution was measured by using phosphate buffered saline (PBS) Was suspended and used as a sample.

In the alcohol extraction method, the fruit juice or shiitake mushroom was immersed in triple solvent for 72 hours at room temperature and dark condition, and then the solution was freeze-dried using a hot distiller to measure the dry weight. The phosphate buffered saline (PBS) and a solvent to prepare a sample.

3-1-2: DPPH  radical Scatters  analysis

20, 50, 100, 200 and 500 μg of free radical scavenging activity of 2,2-diphenyl-1-picrylhydrazyl (DPPH) (1: 1) extract prepared from shiitake mushroom and cucumber mushroom (1: 1) prepared at a concentration of 50 μg and the absorbance value was lowered.

The samples were prepared at a concentration of 1 mg / mL using methanol and the DPPH reagent was prepared by dissolving in methanol at a concentration of 0.1 mM in the absence of light. 100 μl of sample and 0.5 ml of DPPH reagent were added and incubated for 20 minutes. The absorbance was measured at 517 nm using a microplate reader (Molecular Devices, Sunnyvale, Calif., USA). As a negative control, methanol was used instead of the sample. As a positive control, ascorbic acid (1 mg / ml) was added instead of the sample, and the experiment was carried out under the same conditions. The DPPH radical scavenging activity was calculated by the following equation (1): DPPH inhibitory activity (%).

3-1-3: Hydroxyl radical Scatters  analysis

 Hydroxyl radicals generated by Fenton's reagent were reacted with 5, 10, 20, and 50 ㎍ of hot water or shiitake extract of shiitake mushroom or cucurbit fruit of Example 3-1-1, respectively, The scavenging ability was measured.

To 0.15 ml of a sample prepared at a concentration of 1 mg / ml using methanol was added 0.35 ml of buffer, 3 mM deoxyribose, 0.1 mM ascorbic acid, 0.1 mM EDTA, 0.1 mM FeCl ₃ , 1 mM H ₂ O ₂ Solution, stirred well, and reacted at 37 ° C for 1 hour. After the reaction was completed, 2% TCA solution and 1% TBA solution were mixed well, reacted at 100 ° C for 20 minutes, cooled to room temperature and centrifuged. The supernatant was taken and absorbance was measured at 532 nm using a spectrophotometer. For comparative analysis, vehicle (phosphate buffer) was used as a negative control, ascorbic acid was used as a positive control, and the radical scavenging activity (%) was calculated by the following equation (2).

3-1-4: Fe ^{2 +} Chelating chelating activity assay

In the oxidative reaction induction process by the Fe ^{2 +} -ferrozine complex, 5, 10, 20, and 50 μg of the hot water or shiitake extract of the shiitake mushroom or the cucurbit fruit of Example 3-1-1 were reacted respectively and the absorbance value was low The antioxidative effect was measured by using the extract.

60 μl of 2 mM FeCl ₂ and 700 μl of distilled water were mixed with 40 μl of the sample prepared at a concentration of 1 mg / ml using methanol and reacted at room temperature for about 10 minutes. After completion of the reaction after about 5 minutes of reaction at room temperature for by the addition of 5mM ferrozine deriving a Fe ^{2 +} complex -ferrozine using a UV / Visible spectrophotometer (Berkman, USA ) was measured at 562nm. To compare the negative control we were treated with vehicle (Tris-HCl buffer), the positive control group were analyzed for Fe ^{2 +} chelating activity by treatment of ascorbic acid (ascorbic acid). The chelating activity (%) was calculated by the following formula (3).

3-1-5: Analysis of total polyphenol and flavonoid content

In order to analyze the total polyphenol content, the antioxidative effect was measured by reacting the shiitake mushroom or the cucurbit fruit of Example 3-1-1 with 5, 10, 20 or 50 μg of hot water or alcoholic beverage.

100 μl of a sample prepared at a concentration of 1 mg / ml and methanol was added to 100 μl of a Folin-Denis reagent (Fluka, Buchs, Switzerland), and the mixture was reacted at room temperature for 3 minutes. After 3 minutes, 100 μl of 10% sodium carbonate solution was added and mixed. After 1 hour of reaction, the supernatant was taken and absorbance was measured at 760 nm using a microplate reader (Molecular Devices, Sunnyvale, CA, USA). Total polyphenol content was calculated using a standard calibration curve of 10 μg / ml, 20 μg / ml, 30 μg / ml, 40 μg / ml and 50 μg / ml of gallic acid dissolved in methanol.

In order to analyze the total flavonoid content, 1 ml of diethylene glycol was added to 100 쨉 l of a sample prepared by dissolving hot water or alcoholic beverage of Example 3-1-1 or methanol extract in a concentration of 1 mg / ml, Respectively. After mixing, 100 μl of 1N NaOH was added, mixed well, and reacted in a water bath at 37 ° C for 1 hour. After 1 hour, the absorbance was measured at 420 nm using a microplate reader (Molecular Devices, Sunnyvale, Calif., USA). Naringin was used as a reference material, and a standard calibration curve of 10 μg / ml, 20 μg / ml, 30 μg / ml, 40 μg / ml and 50 μg / ml was prepared and the total flavonoid content was converted.

3-1-6: Reducing power active analysis

In the oxidative reaction induction process by FeCl ₃ , 5, 10, 20 and 50 μg of the hot water of the shiitake mushroom or the cucurbit fruit of Example 3-1-1 were reacted with each of 5, 10, 20 and 50 μg respectively, , and ascorbic acid.

500 μl of 0.2 M sodium phosphate buffer (pH 6.6) and 500 μl of 1% potassium ferricyanide were added to the samples prepared at 1 mg / ml and the appropriate concentration using methanol, and the mixture was reacted at 50 ° C for 20 minutes. 10% trichloroacetic acid 2.5 Ml, and the above reaction solution was centrifuged at 650 rpm for 10 minutes. 500 μl of the supernatant was added to 500 μl of distilled water and 100 μl of 1% ferric chloride, and the absorbance of the reaction solution was measured at 700 nm.

3-1-7: Nitrite Scatters  analysis

The antioxidative effect of nitrite was measured by using 5, 10, 20 and 50 μg of each of the hot water or alcoholic beverages of the mushroom or cucumber mushroom of Example 3-1-1 and lowering the absorbance value.

To 50 μl of 1 mM NaNO ₂ solution, add 50 μl of sample prepared with 1 mg / ml and appropriate concentration using methanol, add 300 μl of 0.1 N HCl solution (pH 1.2) to the final volume of the reaction solution to 0.4 ml And reacted at 37 캜 for 1 hour. After the first reaction, 2 ml of 2% acetic acid solution and 0.2 ml of Griess reagent were added, mixed well and reacted at room temperature for 15 minutes. Absorbance was measured at 520 nm using a microplate reader to calculate the amount of remaining nitrite. Experiments were conducted under the same conditions except that distilled water was used as a control, distilled water was used instead of Griess reagent as a negative control, and ascorbic acid (1 mg / ml) was used as a positive control. For comparative analysis, vehicle (PBS) was treated with negative control group and nitrite scavenging ability was treated with ascorbic acid in positive control group. The nitrite scavenging activity was calculated by the following equation (4).

<3-2> DPPH  radical Scatters  Identification of antioxidative effect by analysis

To confirm the antioxidative effect of the shiitake mushroom, the cucumber mushroom, and the composite sample, and the antioxidative effect of the mushroom and the fruit extract of the cucumber mushroom, the scavenging ability of DPPH radical was analyzed by performing the above Example 3-1-2.

As a result, DPPH radical scavenging activity was shown in proportion to the concentration of the sample (FIG. 1A) and the IC50 value of the composite sample was calculated to be 79.481 μg (FIG. 1B).

The DPPH free radical scavenging activity was determined by measuring the DPPH free radicals (Fig. 2A), the shiitake mushroom extract (Fig. 2B), the shiitake mushroom extract (Fig. 2C) Lt; / RTI &gt; activity.

<3-3> Hydroxyl radical Scatters  Identification of antioxidative effect by analysis

In order to confirm the antioxidative effect of the extract of Shiitake mushroom and Curcuma longa, the activity of hydroxyl radical scavenging activity was analyzed by performing the above Example 3-1-3.

As a result, the hydroxyl radical scavenging activity (Fig. 3A), shiitake mushroom extract (Fig. 3B), cucumber mushroom hydrothermal extract (Fig. 3C) and cucumber mushroom extract Respectively.

<3-4> Fe ^{2 +}  Identification of antioxidative effects by chelating activity assay

In order to confirm the antioxidative effect of the shiitake mushroom and the cucurbitan fruit extract, the Fe ^{2 +} chelating activity was analyzed by performing the above Example 3-1-4.

As a result, it was found that Fe ^{2 +} -ferrozine (Fig. 4B), which is proportional to the concentration of the extract, was obtained from the extracts of mushroom hot water extract (Fig. 4A), shiitake mushroom extract complex inhibited the oxidative reaction. Especially, at 50 ㎍ concentration of the extracts of Cryptomeria japonica, the inhibition rate was higher than that of positive control ascorbic acid (AA).

<3-5> Determination of antioxidative effect by total polyphenol and total flavonoid content analysis

In order to confirm the antioxidative effect of the extract of Shiitake mushroom and Cucumber mushroom, the total polyphenol and total flavonoid contents were analyzed by performing the above Example 3-1-5.

The polyphenol contents and flavonoid contents of the mushroom hot water extract, shiitake mushroom extract, cucumber mucilage hot water extract and cucumber mushroom extract were found in all the extracts as shown in Tables 3 and 4, respectively.

Shiitake mushrooms Cucumber fruit Hot water extraction Alcohol extraction Hot water extraction Alcohol extraction Polyphenol content (mg / g) 12.28 ± 0.93 17.15 + - 1.23 26.31 + - 1.85 31.65 ± 2.14

Shiitake mushrooms Cucumber fruit Hot water extraction Alcohol extraction Hot water extraction Alcohol extraction Flavonoid content (mg / g) 1.73 ± 0.31 2.13 ± 0.27 4.13 ± 0.39 7.73 + - 0.42

<3-6> Identification of antioxidant effect by reducing power efficacy analysis

In order to confirm the antioxidative effect of the extract of Shiitake mushroom and Curcuma longa, the total polyphenol and total flavonoid contents were analyzed by performing the above Example 3-1-6.

As a result, Reducing power efficacy of hydrothermal extract of Shiitake mushroom, Shiitake mushroom extract, Shiitake mushroom extract and Cucumber powdery mushroom extract was confirmed in all the extracts as shown in Table 5 below.

Shiitake mushrooms Cucumber fruit Hot water extraction Alcohol extraction Hot water extraction Alcohol extraction Flavonoid content (mg / g) 4.21 + - 0.35 4.89 ± 0.52 11.57 ± 1.03 13.21 + 1.36

<3-7> Nitrite Scatters  Identification of antioxidative effect by analysis

In order to confirm the antioxidative effect of the extract of Shiitake mushroom and Curcuma longa, the nitrite scavenging activity of the Example 3-1-7 was analyzed.

(Fig. 5A, 5B and 5C), shiitake mushroom extract (Figs. 6A, 6B and 6C), cilantom fruit hot water extract (Figs. 7A, 7B and 7C) at pH 1.2, pH 3.0 and pH 5.0 ) And the extracts of Corynebacterium spp. (FIGS. 8A, 8B and 8C) showed the nitrite scavenging activity in proportion to the concentration of the extract.

Example  4: Evaluation of fat and glycosylase activity

<4-1> Evaluation method of fat and digestive enzymatic activity

4-1-1: Preparation of sample

Samples of hot water or alcoholic beverages of shiitake mushrooms or cucurbit fruit were extracted in the same manner as in Example 3-1-1 and prepared at 1, 2, 5 and 10 μg concentrations.

4-1-2: Analysis of α-Amylase activity

Amylase-derived α-amylase enzyme solution (5 units / ml, in 50mM potassium phosphate buffer) was mixed with the hot water of the shiitake mushroom of the Example 4-1-1, 1% starch solution was added to the solution, and the mixture was reacted at room temperature for 5 minutes. After the reaction, 0.1% 3,5-dinitrosalicylic acid (DNS) solution was added, and the mixture was boiled at 100 ° C for 5 minutes. After cooling, distilled water (DW) was added to the reaction solution. The reaction mixture was stirred with a microplate reader (Molecular Devices, Sunnyvale, Calif. And the activity was evaluated. For comparison, the negative control was not treated with 1% starch solution and the positive control was treated with 10 mU of α-Amylase in 1% starch solution.

4-1-3: α- Glucosidase  activation

 rat intestinal acetone powder was mixed with 1 ml of 50 mM potassium phosphate buffer previously reacted at 4 ° C, and the mixture was centrifuged at 4 ° C and 4,000 rpm for 10 minutes. The intermediate layer was used as an enzyme solution of α-glucosidase.

50 μl of the α-glucosidase enzyme solution and 100 μl of 3 mM ρ-nitrophenyl-α-D-glucopyranoside (ρNPG) were mixed with the hot water or the hot water sample of the shiitake mushroom or cranberry fruit of Example 4-1-1 at 37 ° C. After reacting for 60 minutes, the activity was evaluated by measuring the absorbance at 405 nm with a microplate reader (Molecular Devices, Sunnyvale, Calif., USA). For comparative analysis, negative control was treated with 3 mM ρNPG substrate solution, and positive control was treated with 10 mU of α-Glucosidase in 3 mM ρNPG substrate solution.

4-1-4: Lipolytic  activity assay

(10 mM) of P-nitrophenyl butyrate (PNPB), P-nitrophenyl palmitate (PNPP) and P-nitrophenyl decanoate (PNPD) Lipolytic activity assay.

The substrate solution and the sample of the mushroom of the Example 4-1-1 or the hot water of the fruit of the cedar mulberry or the sample of the alcoholic beverage were reacted at 60 ° C for 15 minutes and the p-nitrophenol (PNP) was measured at 405 nm for 10 minutes to obtain Vmax / min). Microplate reader (Molecular Devices, Sunnyvale, CA, USA) was used for the measurement. For comparative analysis, negative control group treated only vehicle and positive control group treated lipase.

<4-2> Confirming the activity of α-Amylase activity

In order to confirm the effect of the α-Amylase-like activity of the extract of Shiitake mushroom and Cajonpong fruit, the activity of the saccharification activity was analyzed by carrying out Example 4-1-2.

The OD ₅₅₀ absorbance of the extracts and concentrations was measured and analyzed to find that the extracts of mushroom hot water (Figs. 9A and 9B), mushroom juice extract (Figs. 10A and 10B), cucumber fruit juice hot water extract (Figs. 11A and 11B) The alcohol extracts (Figs. 12A and 12B) showed the digestive activity in proportion to the concentration of the extract, except for the mushroom hot water extract.

&Lt; 4-3 > Glucosidase  Identification of the digestive activity by activity assay

In order to confirm the effect of? -Glucosidase-like activity of the extract of Shiitake mushroom and Cucumber mushroom, the activity of saccharifying activity was analyzed by carrying out Example 4-1-3.

The extracts were treated with 3 mM ρNPG substrate solution for 60 minutes to determine the OD ₄₀₅ absorbance according to the concentration. As a result, the extracts of mushroom hot water (Figs. 13A and 13B), shiitake mushroom extract (Figs. 14A and 14B) 15A and 15B) and Corynebacterium sp. Extract (Figs. 16A and 16B) showed the digestive activity in proportion to the concentration of the extract. Especially, α-Glucosidase-like activity was the highest in fruit juice extract.

<4-4> Lipolytic  Activity assay to identify lipid digestive activity

In order to confirm the effect of the lipid digestion activity of the extract of Shiitake mushroom and Lycoris spp., The activity of the saccharide digesting activity was analyzed by carrying out Example 4-1-4.

As a result, the hot water or alcoholic beverages of shiitake mushroom or cucumber mushroom were tested for P-nitrophenyl butyrate (PNPB), P-nitrophenyl palmitate (PNPP) and P-nitrophenyl decanoate (PNPD) Lipolytic activity assay showed lipogenic activity in proportion to the concentration of the extract (Tables 6 to 9).

Synthetic substrates (mU / min) Shiitake mushroom hot water extract Lipase 1 g 2 g 3 g 5 g PNPB 0 ± 0 0.03 ± 0.01 0.08 ± 0.01 0.24 ± 0.05 PNPP 0 ± 0 0 ± 0 0.05 ± 0.02 0.32 ± 0.03 PNPD 0 ± 0 0.04 0.02 0.11 + - 0.01 0.31 + 0.04

Synthetic substrates (mU / min) Shiitake mushroom extract Lipase 1 g 2 g 3 g 5 g PNPB 0 ± 0 0 ± 0 0.06 ± 0.01 0.24 ± 0.05 PNPP 0 ± 0 0 ± 0 0.03 ± 0.01 0.32 ± 0.03 PNPD 0 ± 0 0.02 ± 0.01 0.07 ± 0.02 0.31 + 0.04

Synthetic substrates (mU / min) Cucumber fruit fruit extract Lipase 1 g 2 g 3 g 5 g PNPB 0.09 0.02 0.13 + - 0.01 0.19 + 0.04 0.24 ± 0.05 PNPP 0.06 ± 0.01 0.17 + 0.03 0.24 ± 0.05 0.32 ± 0.03 PNPD 0 ± 0 0 ± 0 0.05 ± 0.01 0.31 + 0.04

Synthetic substrates (mU / min) Cucumber fruit juice extract Lipase 1 g 2 g 3 g 5 g PNPB 0.14 + 0.02 0.18 ± 0.05 0.31 + 0.04 0.24 ± 0.05 PNPP 0.10 0.02 0.22 + 0.03 0.25 0.06 0.32 ± 0.03 PNPD 0.03 ± 0.01 0.05 ± 0.01 0.09 0.02 0.31 + 0.04

Example  5: Analysis of gastrointestinal protection effect

<5-1> Analysis of gastrointestinal protection effect

5-1-1: Cell culture

RAW264.7 cells, RBL-2H3 cells and AGS cells were cultured in DMEM medium containing 10% FBS and 1X antibiotic-antimycotic at 37 ° C and 5% CO ₂ . When the cells grew to about 70 to 80% of the culture flask area, they were subcultured using trypsin-EDTA.

5-1-2: Cytotoxicity measurement

Cell viability was measured by 3- (4,5-dimethylthiazole-2-yl) -2,5-diphenyl-tetrazoliumbromide (MTT) reduction method. RAW264.7 cells, RBL-2H3 cells, and AGS cells of Example 5-1-1 were divided into 96-well plates and cultured for 12 hours. Then, shiitake mushroom or cilantrofruit extract was added to 0, 1, 5 10, 50, 100, 200, 300 and 500 占 퐂 / ml, and cultured for 24 hours. Subsequently, MTT solution (final concentration: 0.5 mg / ml) was added and incubated at 37 ° C for additional 4 hours to reduce the MTT and the medium was removed so that the generated formazan did not fall off the medium. 100 μl of DMSO was added, and the mixture was mixed for 10 minutes. Then, the absorbance at 540 nm was measured to analyze the cell survival rate.

5-1-3: Measurement of NO production

The RAW 264.7 cells of Example 5-1-1 were divided into 24-well plates and cultured for 12 hours. Then, the mushroom extract, cucumber fruit extract or their mixed extracts were added to 0, 50, 100, 150, / Ml, followed by treatment with LPS (1 占 퐂 / ml) and culturing for 20 hours. 100 μl of the cell culture solution and 100 μl of Griess reagent were mixed and reacted at room temperature for 10 minutes. Absorbance was measured at 550 nm using an ELISA reader, and a standard curve was prepared with sodium nitrate to calculate the NO content.

5-1-4: β- hexosaminidase  Secretion inhibiting activity

Using the DMEM medium containing dinitropheny-immunoglobulin E (DNP-IgE) at a concentration of 0.5 μg / ml, RBL-2H3 cells of Example 5-1-1 were inoculated into a 24-well plate at 2 × 10 ⁵ cells / well And then cultured in a 5% CO ₂ incubator at 37 ° C for 12 hours. Each cell Siraganian buffer (119mM NaCl, 5mM KCl , 5.6mM glucose, 0.4mM MgCl 2, 25mM PIPES, 1mM CaCl 2, 0.1% BSA and pH 7.2) and washed twice with and _{then, 37 ℃, 5% CO 2} incubator The cells were treated with shiitake mushroom extract, cucumber powdery extract or mixed extract thereof at the concentrations of 0, 50, 100, 150, and 200 μg / ml, respectively, at 37 ° C and 5% CO ₂ incubator for 30 min. DNP-bovine serum albumin (DNP-BSA) was added to a final concentration of 100 ng / ml, incubated at 37 ° C in a 5% CO ₂ incubator for 2 hours, and incubated in an ice bath for 10 minutes. 40 μl of the supernatant was transferred to a 96-well plate, and 40 μl of substrate buffer (2 mM 4-p-nitrophenyl-N-acetyl-β-D-glucosaminide, 0.05 M sodium citrate, pH 4.5) , And 200 μl of stop solution (0.1 M Na ₂ CO ₃ / NaHCO ₃ , pH 10.5) was added to each well to terminate the reaction. Absorbance was measured at 405 nm.

5-1-5: Histamine secretion inhibitory activity

The cultured RBL-2H3 cells of Example 5-1-1 were treated with DNP-BSA to induce histamine release, and the extracts of mushroom, cilantro, or their mixture were administered at 0, 50, 100, 150 and 200 μg / ml, and incubated at 37 ° C in a 5% CO ₂ incubator for 30 minutes. After centrifugation at 2000 rpm for 5 minutes at 4 ° C, the supernatant was used as a sample.

25 μl of the above sample was added to a 1.5 ml tube, and 22.5 μl of 0.1 N HCl and 42.5 μl of 60% HClO were added, and the mixture was centrifuged, and 40 μl of the supernatant was added to 25 μl of 5N NaOH, 500 μl of n-butanol, The mixture was placed in a tube, shaken, and centrifuged (2000 rpm, 10 minutes). Butanol layer was added to a fresh 1.5 ml tube, 150 0.1 of 0.1 N HCl solution and 0.5 ml of n-heptane were added, and the mixture was shaken and centrifuged. To 100 수 of the water layer obtained, 200 1 of 1N NaOH solution and 5 0.1 of 0.1% ο-phthaldialdehyde solution were added, and the mixture was reacted at 37 캜 for 3 minutes. Then, 10 3 of 3N HCl solution was added, mixed and left for 2 minutes, and the absorbance was measured at 450 nm (excitation) and 450 nm (emission).

5-1-6: Measurement of Prostaglandin E2

The concentration of AGS cells in Example 5-1-1 was adjusted to 1 × 10 ⁴ / well, and the cells were treated with shiitake mushroom extract, cucumber extract or their mixed extracts at 0, 100 and 150 μg / ml for 1 hour Were preincubated. Indomethacin (500 μM) was used as a control and quantified using the PGE2 assay kit.

5-1-7: Measurement of cytokine

The RAW264.7 cells and RBL-2H3 cells of Example 5-1-1 cultured with 1 μg / ml of LPS and 0, 100 and 150 μg / ml of mushroom extract, And TNF-a, a secreted cytokine, was quantified.

100 μl of the primary antibody of TNF-α to be quantified is pre-coated on a 96-well plate which is coated beforehand, and then it is overnight at 4 ° C. The next day, the cells were washed three times with 0.5% tween 20 wash solution for 5 minutes each. Then, 100 μl of the sample to be measured and the standard solution were added and reacted at room temperature for 2 hours. After washing, HRP-conjugated secondary antibody (100 μl / well) was added thereto, followed by reaction at room temperature for 2 hours. After washing again, avidin / biotin was added to measure the absorbance. Standard curves were obtained from the absorbance of the standard and the amount of TNF-? Was determined from the absorbance of each sample.

<5-2> Cell survival analysis by cytotoxicity measurement

The cell viability was analyzed by performing the above Example 5-1-2 to confirm cytotoxicity of the shiitake mushroom or the cucumber mushroom fruit extract.

As a result, in the RAW 264.7 cells, the RBL-2H3 cells and the AGS cells, the mushroom extract (Figs. 17A, 17B and 17C) and the cilantro fruit extract (Figs. 18A, 18B and 18C) It does not appear to have any effect.

<5-3> Determination of inhibitory effect of NO formation by measurement of NO production amount

In order to confirm the NO production amount of the shiitake mushroom or the cucumber mushroom extract, the inhibitory effect of NO production was analyzed by carrying out Example 5-1-3.

As a result, the NO inhibitory effect was increased as the concentration of the extract increased, and the inhibitory effect of the mushroom extract was more excellent than that of the fruit extract and mixed extract (Fig. 19A, 19B and 19C).

&Lt; 5-4 > hexosaminidase  Identification of anti-inflammatory effects by secretion inhibition activity

In order to confirm the effect of inhibiting β-hexosaminidase secretion activity of the extract of Shiitake mushroom or Cjidjipan fruit, anti-inflammatory effect was analyzed by performing the above Example 5-1-4.

As a result, the inhibitory effect of β-hexosaminidase secretion was increased as the concentration of the extract increased, and the inhibitory effect of the fruit extract was better than that of the mushroom extract and mixed extract (FIGS. 20A, 20B and 20C).

<5-5> Antiinflammatory effect according to histamine secretion inhibition activity

In order to confirm the effect of inhibiting the histamine secretion of the extract of Shiitake mushroom or Cjidjipon fruit, the anti-inflammatory effect was analyzed by performing the above Example 5-1-5.

As a result, the inhibitory effect of histamine was increased as the concentration of the extract increased (Figs. 21A, 21B, and 21C).

<5-6> Determination of anti-inflammatory effect by measurement of Prostaglandin E2 (PGE2)

In order to confirm the effect of promoting the secretion of PGE2 in shiitake mushroom or cucumber mushroom fruit extract, the anti-inflammatory effect was analyzed by carrying out Examples 5-1-6.

As a result, as the concentration of the extract increased, the promoting effect of the PGE2 secretion was increased, and the promoting effect of the mixed extract was superior to that of the mushroom extract and cucumber mushroom extract (FIGS. 22A, 22B and 22C).

<5-7> Identification of anti-inflammatory effects by cytokine measurement

The anti-inflammatory effect was analyzed by performing the above Example 5-1-7 in order to confirm the TNF-α secretion inhibitory activity effect, which is cytokine of the extract of Shiitake mushroom or Curcuma longa.

As a result, in RAW264.7 cells (Fig. 23A, 23B and 23C) and RBL-2H3 cells (Fig. 24A, 24B and 24C), secretion of TNF-a was inhibited as the concentration of extract increased.

Example  6: Nutritional shake  Analysis of weight control effect

<6-1> Nutritional shake  Method of analysis of weight control effect

6-1-1: Design of animal models

Six week old ICR male rats were used and the experimental group was divided into four groups as follows. Powdered nutritional shake samples were used and treated as follows.

- Group 1 (control group): fed with normal diet and saline.

Group 2 (Experimental Group 1): The nutritional shake of Example 1 was orally administered in powdered form (100 mg / kg / day) for 50 days.

Group 3 (Experimental Group 2): The nutritional shake of Example 1 was orally administered in powdered form (200 mg / kg / day) for 50 days.

- Group 4 (experimental group 4): A 50-day oral administration of a comparative sample (200 mg / kg / day of the developed nutritional shake powder)

6-1-2: Daily Average weight gain (Average Daily Gain, ADG ) analysis

The body weights of the experimental animals were measured in the animal breeding room for 50 days after the adaptation period, and the general feed intake was measured for 50 days.

The body weight of the control and experimental groups was measured at the same time until the end of the dieting, and the average daily gain was analyzed.

6-1-3: Average Daily Feed Intake, ADFI ) analysis

Dietary intake of control and experimental groups was measured at the same time till the end of the dietary cycle and the average daily intake was analyzed.

6-1-4: Dietary efficiency (G / F) analysis

 Based on the results of the body weight and the intake of the control group and the experimental group measured until the end of the dieting, the efficiency was calculated by the following equation (5).

6-1-5: Analysis of growth rate and growth rate

The growth rate and body weight gain of the control and experimental groups were calculated by the following equations (6) and (7).

6-1-6: Protein analysis of muscle tissue

After the end of the diet, the thighs and muscle tissues extracted from the control and experimental groups were disrupted, lysed by adding lysis buffer, and quantified by BCA kit.

6-1-7: Back room  Thickness and visceral fat weight analysis

After the end of the meal, the thickness of the back room and the weight of visceral fat extracted from the control and experimental groups were measured.

6-1-8: Biochemical analysis in serum

After fasting for 12 hours after the end of the dietary habituation of the experimental animals, fasting blood was collected from the tail after inserting the rat into the fixed body without anesthesia, and blood was collected by sacrifice and cardiac blood collection. The collected blood was centrifuged at 3,000 rpm and 4 ° C for 15 minutes to separate the serum and stored at 70 ° C until the biochemical analysis of the serum. Total cholesterol (TC), high density lipoprotein (HDL), triglyceride (TG) and total protein in rats were measured using hematology analyzers. The contents of low density lipoprotein (LDL) and very low density lipoprotein (VLDL) were calculated by the Friedewald equation.

(HDL-cholesterol, HDL-C), triglyceride and low-density lipoprotein cholesterol (LDL-C) and ultra-low density lipoprotein cholesterol VLDL-cholesterol) are derived from the following equations (8) to (11).

<6-2> Confirmation of weight increase

In order to analyze the weight gain effect of the nutritional shake of Example 1, the body weights of the control group and the experimental group were measured.

The body weight gain rate was analyzed on the basis of the average body weight of each group before and after the dietary and sample intake (FIG. 25A). As a result, the weight gain rate of the experimental group 2 (69%)> experimental group 1 (62%)> experimental group 3 (52% ) Weight gain (FIG. 25B).

The body weight gain rate (FIG. 25C) based on the average body weight of the control group after the diet and the sample intake was analyzed to find that the weight gain rate of the test group 2 (12.5%)> test group 1 (7.6%)> test group 3 It looked.

<6-3> Daily Average weight gain  Analysis and analysis of average daily intake Dietary hygiene Check rate

In order to analyze the weight gain effect of the nutritional shake of Example 1, the daily average weight gain, daily average intake and dietary efficiency were analyzed by performing Examples 6-1-2 to 6-1-4.

As a result, the average daily gain was in the order of experimental group 2> experimental group 1> experimental group 3> control group (FIG. 26A). That is, 26.56% of the experimental group 1, 39.58% of the experimental group 2 and 4.69% of the experimental group 3 were higher than the average daily gain of the control group.

In addition, the average daily intake was in the order of experimental group 1> control group> experimental group 3> experimental group 2 (FIG. 26B). That is, compared with the average daily intake of the control group, the experimental group 1 was 1.17% higher, while the experimental group 2 was 2.80% and the experimental group 3 was 0.77% lower.

The dietary efficiency was calculated from daily average weight gain and average daily intake of the control group and each experimental group, and experimental group 2> experimental group 1> experimental group 3> control group (FIG. 26C). That is, the nutritional shake of Example 1 shows excellent dietary efficiency.

<6-4> Determination of Growth Effect by Field Measurement and Field Growth Rate Analysis

In order to analyze the weight gain effect of the nutritional shake of Example 1, the total length of the experimental animals was measured at the beginning and the end of the experiment to analyze the growth rate of the field.

As a result, the total growth of the experimental group 1 and the experimental group 3 was lower than that of the control group, but the experimental group 2 showed a higher total field growth than the control group (FIG. 27A).

(15%)> Experimental group 1 (14%)> Experimental group 3 (13%) Experimental group 2 (17%) Experimental group 1 %).

As a result of the analysis of the growth rate of the field based on the average length of the control group after the intake of the diets and samples, the field growth rate of the experimental group 1 was -1.02% and the growth rate of the control group 2 The growth rate of the field was 1.79% and the growth rate of the field 3 was -1.98% (Fig. 27C). That is, the nutrient shake of Example 1 exhibits an excellent field growth effect.

<6-5> Fuji Muscle protein  Confirmation of weight gain effect by analysis

In order to confirm the weight gain effect of the nutritional shake of Example 1, the muscle protein of the fuzzy muscle was analyzed by performing the Example 6-1-6.

As a result, muscle protein of Fuji muscle was analyzed to be 8.54% in Experiment 1, 10.73% in Experiment 2, and 2.19% in Experiment 3, compared with the control (Fig. 28).

<6-6> Back room  And visceral fat analysis

In order to confirm the effect of weight gain of the nutritional shake of Example 1, the above Example 6-1-7 was performed to analyze backbone and visceral fat.

As a result, the mean isokinetic thickness was 4.73% lower in the experimental group 1 than in the control group, but 13.51% in the experimental group 2 and 3.38% higher in the experimental group 3 (FIG. 29A).

In addition, the mean visceral fat weight was 7.23% in the experimental group 1, 10.84% in the experimental group 2, and 6.02% in the experimental group 3 compared to the control group (FIG. 29B).

<6-7> Improvement of metabolism of cholesterol in the body according to biochemical analysis in blood and maintenance of numerical value of risk indicators related to cardiovascular disease

In order to examine whether the nutritional shake of Example 1 improved the metabolism of cholesterol in the body and whether the risk index related to cardiovascular disease changes, biochemical analysis in blood was carried out in Example 6-1-8.

As a result, the total cholesterol level was not significantly changed in the experimental group 1, but slightly increased in the experimental group 2 and the experimental group 3 as compared with the control group (Fig. 30A). The triglyceride levels were slightly increased in experimental group 1 and experimental group 2 compared with the control group, and slightly increased in experimental group 3 (Fig. 30B). HDL levels were similar in Experiment 1 and Experiment 3 compared to the control, but slightly increased in Experiment 2 (Figure 30C). LDL levels were slightly increased in experimental group 1 and experimental group 2 compared with the control group, and increased significantly in experimental group 3 (FIG. 30D). VLDL levels were similar in experimental group 1 and experimental group 2 compared to the control group, but slightly increased in experimental group 3 (Fig. 30E). Taken together, the ingestion of the nutritional shake of Example 1 above could slightly increase the triglyceride and VLDL levels, but did not show a significant increase in LDL and total cholesterol, and HDL levels at high (200 mg / kg) And may help to metabolize and improve cholesterol in the body.

 Further, analysis of atherosclerosis index, arteriosclerosis correlation coefficient, cardiac risk index, and coronary artery index change by biochemical analysis in the blood revealed that the ingestion of the nutritional shake of Example 1 can slightly increase the arteriosclerosis index (FIG. 31A), the arterial hardening correlation coefficient (FIG. 31B), the cardiac risk index (FIG. 31C), and the coronary artery index (FIG. 31D) The ingestion of the nutritional shake of Example 1 can be expected to provide a weight control effect without changing the value of the cardiovascular disease risk index.

The present invention has been described with reference to the preferred embodiments. It will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims. Therefore, the disclosed embodiments should be considered in an illustrative rather than a restrictive sense. The scope of the present invention is defined by the appended claims rather than by the foregoing description, and all differences within the scope of equivalents thereof should be construed as being included in the present invention.

Claims (8)

A food composition for improving blood cholesterol and for increasing body weight, comprising a cranberry fruit extract and a mushroom extract,
The composition was prepared by mixing 37 parts by weight of isolated soybean protein, 15 parts by weight of crystalline fructose powder, 9 parts by weight of cocoa powder, 8 parts by weight of fruit extract powder, 5 parts by weight of mushroom powder, 11 parts by weight of casein sodium, 3 parts by weight of dough flavor powder, 2 parts by weight of cream-flavored powder, 1 part by weight of silicon dioxide, 3 parts by weight of a honey powder, and 5 parts by weight of a vitamin powder,
The above-mentioned fruit extract powder and shiitake mushroom powder,
(a) dipping fruit and mushroom in a three-fold volume of ethanol solvent, and allowing to stand at room temperature for 72 hours under dark condition;
(b) preparing an extract by ethanolic extraction using a hot distiller; And
(c) lyophilizing the extract
&Lt; / RTI &gt;
A food composition for improving blood cholesterol and for increasing body weight, comprising a fruit extract and a mushroom extract.
delete delete delete A functional food for improving digestive enzyme activity comprising the food composition of claim 1. A nutritional food for improving digestive enzymatic activity comprising the food composition of claim 1. delete delete

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