JP2019073476A - Agent for atp production promotion - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide an ATP production promoting agent which is safe even when taken for a long period of time, can be used as a food or drink, and has an excellent ATP (energy) production promoting action. SOLUTION: A barley young leaf or an ATP production promoting agent containing a specific component contained in the barley young leaf as an active ingredient, and a food or drink containing the ATP production promoting agent. Since such ATP production promoting agents and foods and drinks have an ATP production promoting action, they are caused by activation of cell functions such as cell proliferation, metabolism and repair, effects such as anti-aging, and a decrease in ATP production. Prevention or improvement (treatment) of the disease or condition (symptom) is expected. [Selection diagram] None

Description

  The present invention relates to an agent for promoting ATP production, and more specifically, an agent for promoting ATP production containing a specific ingredient contained in barley young leaves or young barley leaves, the agent for promoting ATP production comprising the agent for promoting ATP production The present invention relates to a food or drink for the prevention or amelioration (treatment) of a disease or condition (symptoms) caused by a decrease.

ATP (adenosine triphosphate) is widely distributed in cells of all living bodies, and plays an important role as an energy source of biological reactions in particular of eukaryotes. ATP functions as an important energy source, for example, in various in vivo reactions such as glucose metabolism, muscle contraction, active transport, biosynthesis and the like.
Therefore, not only during exercise but also at rest, the production of ATP is promoted, and by supplying the ATP to the cells, activation of cell functions such as cell proliferation, metabolism, repair and anti-aging (anti-aging) The effect of can be expected.

  Therefore, attention has been focused on components having an action of promoting ATP production derived from natural products which are highly safe and have no side effects even when taken for a long time, and various proposals have been made (see, for example, Patent Documents 1 to 5). However, substances that are more effective, safer, and safe to take on a daily basis are expected.

  On the other hand, young leaves of barley and the like, especially young barley leaves, have anti-ulcer activity, anti-hypercholesterol activity, anti-inflammatory activity, hypoglycemic activity, antimutagenic activity, antithrombotic activity, blood vessel protective activity, adipocyte differentiation inhibitory activity, melamine synthesis It has been reported that it has various physiological actions such as inhibitory action, osteoporosis preventive action, active oxygen scavenging enzyme (SOD) gene expression promoting action (see, for example, Non-Patent Document 1, Patent Documents 6 to 9). Also, various proposals have been made for the manufacturing method thereof (see, for example, Patent Documents 10 to 12). However, as far as the present inventors know, it has not been reported that barley young leaves have an ATP production promoting action.

JP, 2013-139394, A JP, 2011-184381, A JP, 2011-162504, A Unexamined-Japanese-Patent No. 2010-120908 JP, 2009-256272, A JP 2008-56580 A JP, 2011-51948, A JP, 2013-63940, A JP, 2015-140338, A JP, 2009-148213, A JP 2003-33151 A Japanese Patent Laid-Open No. 2002-65204

Kasahara Yoshihide, et al., "The physiologically functional substances of barley young leaf green juice powder," Journal of Japan Society for Food Chemistry and Technology Vol. 48, No. 10, pp. 712-725 Oct. 2001

  If ATP production can be promoted in cells, as described above, effects of activation of cell function and anti-aging can be expected. Therefore, the subject of the present invention is attributed to the reduction of ATP production including an ATP production promoter which has high safety, higher ATP production promoting effect, and can be taken safely over a long period of time, and the ATP production promoter. Food or drink for the prevention or amelioration (treatment) of a disease or condition (symptom) to be treated.

  As a result of intensive studies on the above problems, the present inventors have found that barley young leaves, which are safe even when taken for a long time, have an promoting action on ATP production, and certain components contained in barley young leaves promote ATP production. It has been found that it has an action, and the present invention has been completed. That is, the present invention is as follows.

(1) An agent for promoting ATP production, which contains, as an active ingredient, a specific component contained in young barley leaves or young barley leaves.
(2) The agent for promoting ATP production according to (1), wherein the young barley leaves are squeezed powder or dried powder of young barley leaves.
(3) (1) or (2), wherein the specific component contained in young barley leaves contains at least one compound selected from the group consisting of the following compounds (a), (b), (c) and (d): The agent for promoting ATP production as described in 2.).
(A) Composition _formula: C _{33 H} 40 _{O 19,} molecular weight: 740, compound name: robinin (b) the composition _formula: C _{27 H} 30 _{O 15,} molecular weight: 594, compound name: saponarin (c) the composition _{formula: C 25} H ₃₇ O ₅ N ₂ , molecular weight: 445
(D) Compositional Formula: C ₂₁ H ₂₀ O ₁₀ , Molecular Weight: 432, Compound Name: Afzelin (4) At least one selected from the group consisting of the following compounds (a), (b), (c) and (d) An agent for promoting ATP production, comprising a compound of a species as an active ingredient.
(A) Composition _formula: C _{33 H} 40 _{O 19,} molecular weight: 740, compound name: robinin (b) the composition _formula: C _{27 H} 30 _{O 15,} molecular weight: 594, compound name: from saponarin (c) young barley Compound formula: C ₂₅ H ₃₇ O ₅ N ₂ , molecular weight: 445 compound (d) Composition formula: C ₂₁ H ₂₀ O ₁₀ , molecular weight: 432, compound name: afzelin (5) above (1) to (4) A food or beverage for preventing or ameliorating (treating) a disease or condition (symptoms) caused by a decrease in ATP production, comprising any of the agent for promoting ATP production according to any of the above.

  In addition, as another embodiment of the present invention, in a subject in need of preventing or ameliorating (treating) a disease or condition (symptom) caused by a decrease in ATP production, a specific component contained in barley young leaves or barley young leaves Or a disease or condition caused by a decrease in ATP production (a symptom comprising a step of administering at least one compound selected from the group consisting of the above compounds (a), (b), (c) and (d) Specific methods contained in young barley leaves or young barley leaves for use as a method for preventing or improving (treating) or for use as a preventive or ameliorating (treating) agent for diseases or conditions (symptoms) caused by decreased ATP production, or At least one compound selected from the group consisting of the above compounds (a), (b), (c) and (d), or the prevention or modification of a disease or condition (symptoms) caused by a decrease in ATP production A specific component contained in barley young leaves or barley young leaves for use in (treatment), or at least one compound selected from the group consisting of the above compounds (a), (b), (c) and (d), Specific components contained in barley young leaves or barley young leaves for producing an agent for the prevention or amelioration (treatment) of a disease or condition (symptoms) caused by decreased ATP production, or the above compounds (a), (b), (c) And the use of at least one compound selected from the group consisting of (d).

  According to the present invention, at least one compound selected from the group consisting of the above-mentioned compounds (a), (b), (c) and (d), or a specific component contained in barley young leaves or young barley leaves that is the active ingredient. Since it has an action of promoting intracellular ATP production, that is, an action of promoting energy production, effects such as activation of cell functions such as cell proliferation, metabolism and repair, and anti-aging can be expected. In addition, subjects who require the prevention or amelioration (treatment) of a disease or condition (symptoms) resulting from a decrease in ATP production are expected to prevent or ameliorate (treatment) such a disease or condition without side effects. Ru.

  The young barley leaves used in the present invention have no problems with safety and can be freely prepared in the form of food, drink or medicine. Therefore, healthy people as well as old people, sick people, post-ill people, etc. It can be taken over a period of time.

It is a figure which shows the fractionation flow of a squeeze liquid. It is a figure which shows the result of a cytotoxicity test (MTT assay). It is a figure which shows the ATP production amount of an ethyl acetate fraction (BLE) and an aqueous-layer fraction (BLW). It is a figure which shows the chromatogram by HPLC analysis of ethyl acetate fraction (BLE). It is a figure which shows the ATP production amount of each fraction obtained by HPLC. It is a figure which shows the analysis result by LC-MS / MS of a HPLC fraction (BLE-5). It is a figure which shows the analysis result by LC-MS / MS of a HPLC fraction (BLE-6). It is a figure which shows the analysis result by LC-MS / MS of a HPLC fraction (BLE-8). It is a figure which shows the analysis result by LC-MS / MS of a HPLC fraction (BLE-9).

  The embodiment of the present invention will be described in detail below, but the following description is an example of the embodiment of the present invention, and the present invention is not limited to the following description. In addition, when using the expression "-" in this specification, it shall use as an expression containing the numerical value or physical-property value before and behind that.

  In a preferred embodiment of the present invention, the agent for promoting ATP production contains young barley leaves as an active ingredient, and the young barley leaves are barley harvested from the beginning of the tilling period to the beginning of the heading period (plant height about 20 to 40 cm) Young leaves (stems and leaves) are preferably used.

  Here, as barley, any species may be used so long as it belongs to the Hordeum vulgare species such as Nijo barley, Shijo barley, Six-row barley, naked barley etc., and any breed can be used, but Nijo barley, Six-row Barley and naked barley are preferred, and six-row barley is particularly preferred.

  More specifically, as varieties of Nijo barley (beer barley), for example, Aomi Nori, Aguri mochi, Asaka gold, Kinu Yutaka, Umi Yutaka, Kiri Nijo, Haruna Nijo, Amanagi Nijo, Fuji Nijo, Ishiku Shiraz, Misato Golden, Kawahonami, Kawa Mizuki, Kinuka Nijo, Komaki Nijo, Sakitama Nijo, Tone Nijo, Sachiho Golden, Satsuki Baare, Mihoru Gold, Sky Golden, Takachiho Golden, No Susoshi, Nijo Nijo, Nashiro Nijo, Nishino Gold , Nishinochikara, Nishino Hoshi, Haruka Nijo, Nirasaki Nijo, Nira Nijo, Harushi Shizuku, Satsuki Nijo, Hoshin, Mikamo Golden, Miho Golden, Myogi Nijo, Yashio Golden, Yachiho Golden, Kita Ikyou No. 41, Aya Stars, Ryofu and the like.

  As a variety of six-row barley, for example, asam wheat, kashamagor, minrimony, katrimigi, musashinomugi, drill wheat, sanadamugi, bald eel, sea eel, snail, suzurashi, sun rai, silky snow, シ ボ ジ ボ シ, す モ, ツ ゲ ン, ト リ オ オHamayutaka, Hayamiomumu, fiber snow, doubles, Miyuki barley etc. may be mentioned.

  As varieties of naked barley, for example, asparagus scorpion, scorpion hadaka, sanuki syllabary, snail snail, sanshu, ss ss, ss ss ss s, sss sss ss ss ss, sss sss sss sss ss ss sss s Mantenboshi), sembonghadaka etc. are mentioned.

  These varieties are nurtured by agricultural test stations, agricultural research centers, private companies, etc., and fruits and young leaves are used for food (including beverages). From these varieties, it is preferable to select and breed a variety suitable for the cultivation area, and to harvest young leaves (stems and leaves).

  In the present invention, as described above, the young barley leaf is a young leaf (stem / leaf) itself having a height of about 20 to 40 cm from the beginning of tillering to the initial period of heading, or any treated form thereof. Good.

  Examples of treated products of young leaves (stems and leaves) include, but are not limited to, the squeeze processed product, the crushed processed product, the shredded processed product, the dried processed product, the roasted processed product, the extracted processed product, the powdered processed product, the frozen processed product Examples thereof include processed products such as squeezed and dried powder processed products, dried processed products, crushed and filtered processed products (purees and juices), shredded and hot water extracted processed products, and combinations thereof.

  Among these, as a form usually used, powdered treated product (hereinafter sometimes referred to as "squeezed powder") of squeezed treated product (squeezed juice) of young leaf (stem and leaf), dried processed material of young leaf (separate) Hereinafter, it may be referred to as "dry powder").

  Here, the squeeze powder is obtained by, for example, harvesting (cutting off) young leaves of barley according to the method described in Patent Documents 6 to 10, etc. It can be prepared by mixing with an excipient and spray-drying. This squeeze powder can be suitably used as the agent for promoting ATP production of the present invention as it is or in combination with other excipients and the like.

  In addition, the dried powder is shredded after harvesting (cutting) the young leaves of barley according to the method described in, for example, Patent Documents 11 and 12 and the like, drying treatment and crushing treatment, and blanching as necessary. The treatments can be combined and prepared. This dry powder can be suitably used as the agent for promoting ATP production of the present invention as it is or in combination with other excipients and the like.

  In the present invention, the young barley leaves or the specific components contained in the young barley leaves have an action of promoting intracellular ATP production, that is, an action of promoting energy production, as specifically shown in the examples described later. By using the preparation, effects such as activation of cell functions such as cell proliferation, metabolism, and repair and anti-aging can be expected. In addition, subjects who require the prevention or amelioration (treatment) of a disease or condition (symptoms) resulting from a decrease in ATP production are expected to prevent or ameliorate (treatment) such a disease or condition without side effects. Ru. Such diseases or conditions resulting from the decrease in ATP production include, for example, neurodegenerative diseases (Alzheimer's disease, Parkinson's disease etc.), chronic kidney disease, diabetes, heart disease, liver dysfunction etc. ("new ATP production Elucidation of mechanism and its application to the treatment of mitochondrial diseases-Therapeutic drug candidate MA-5 activates mitochondrial function-"Tohoku University 2017 Press Release (June 1, 2017) https: //www.tohoku. See ac.jp/japanese/2017/06/press20170601-01.html).

It has already been confirmed that various physiologically active ingredients are present in young barley leaves (see, for example, Non-Patent Document 1). According to the study of the present inventors, it was speculated that at least the following compounds (a) to (d) are active components of the above-mentioned ATP production promoting action.
(A) Composition _formula: C _{33 H} 40 _{O 19,} molecular weight: 740, compound name: robinin (b) the composition _formula: C _{27 H} 30 _{O 15,} molecular weight: 594, compound name: saponarin (c) the composition _{formula: C 25} H ₃₇ O ₅ N ₂ , molecular weight: 445
(D) Composition formula: C ₂₁ H ₂₀ O ₁₀ , molecular weight: 432, compound name: afzelin However, if the active ingredient of the agent for promoting ATP production in the present invention is derived from young barley leaves, these four types of It is not limited to the compound, and any component may be used.

  Here, robinin, saponarin (Saponarin) and afzelin (Afzelin) are all linked to flavonoids having a 3-hydroxyflavone (3-hydroxy-2-phenylchromen-4-one) skeleton by glycosidic linkage. Compounds, namely flavonol glycosides. Therefore, the agent for promoting ATP production in the present invention may have flavonol glycoside as an active ingredient.

  The flavonol glycoside is not limited to the above three compounds, and may be any flavonol glycoside as long as it has an action of promoting ATP production in one or more species. Specifically, for example, astragalin, azalein, hyperoside, isoquercitrin (isoquercitrin, isoquercitrin, isoquercitrin), kaempferitrin (kaempferitrin), in addition to the above three compounds. ), Quercitrin (quercitrin, quercitrin, quercitrin), rutin (rutin, rutoside, quercetin-3-rutinoside), amrensin (amurensin), icariin and the like.

  Therefore, the agent for promoting ATP production of the present invention comprises any compound selected from the group consisting of the above (a), (b), (c) and (d), and has an action of promoting ATP production. The above compounds and flavonol glycosides are not limited to those derived from young barley leaves, but may be derived from any plant. In addition, the agent for promoting ATP production of the present invention may contain an isolated compound or flavonol glycoside as an active ingredient, and any of these agents is included in the present invention. In the present specification, the above-mentioned compound and / or flavonol glycoside may be referred to as "specified component".

  In addition, the agent for promoting ATP production of the present invention is a specific component contained in young barley leaves or young barley leaves in a subject who is required to prevent or ameliorate (treat) a disease or condition (symptoms) caused by a decrease in ATP production. A method for preventing or ameliorating (treating) a disease or condition (symptom) caused by a decrease in ATP production, comprising the step of administering Barley young leaf or barley young leaf for use in the prevention or amelioration (treatment) of a disease or condition (symptom) caused by a decrease in ATP production or a specific component contained in barley young leaf or young barley leaf for use as a treatment) agent Are included in barley young leaves or barley young leaves for manufacturing a specific component contained in the compound or a preventive or ameliorating (treating) agent for a disease or condition (symptom) caused by a decrease in ATP production The use of the constant component is expected.

  In the present specification, “prevention or amelioration” of a disease or condition is used in a meaning including regulation of disease or condition, delay of progression, alleviation, prevention of onset, prevention of recurrence, suppression and the like.

  In the present invention, the specific components contained in young barley leaves or young barley leaves have high affinity with water, can be dissolved or suspended, and have excellent miscibility with components that can be usually added to foods and beverages and pharmaceuticals. ing. In addition, barley young leaf is a component that has already been used for many years as a food and drink, and the specific component is a component contained in barley young leaf and is excellent in safety. Therefore, an agent for promoting ATP production, which contains as an active ingredient a specific ingredient contained in this young barley leaf or young barley leaf, can be suitably taken in daily eating habits and taken without undue relief.

  The agent for promoting ATP production according to the present invention contains a specific component contained in young barley leaves or young barley leaves as an active ingredient for promoting intracellular ATP production, which has a limited application of “use for promoting ATP production”. It is an agent, and can be used alone as a food or drink or medicine (preparation). Moreover, it can be made to be contained in various compositions, such as food-drinks and a pharmaceutical, as an active ingredient of ATP production promotion. Thereby, the food-drinks composition or pharmaceutical composition for ATP production promotion can be obtained. The obtained food-drinks composition or pharmaceutical composition for promoting ATP production can be effectively used for diseases or conditions (symptoms) caused by the above-mentioned decrease in ATP production.

  The agent for promoting ATP production in the present invention is not particularly limited as long as it is a form that can be ingested, such as a solution, a suspension, an emulsion, a powder, a solid molding, etc. Specifically, for example, in addition to powders, capsules, tablets, granules, powders, forms used in ordinary foods and medicines such as beverages and foods can be mentioned. Further, as the form of the preparation, powders, capsules, tablets, granules, drinks and the like which can be easily adjusted in intake can be suitably exemplified.

  In the present invention, specific components contained in young barley leaves or young barley leaves may be used as they are, or may be used to produce a desired preparation by a conventionally known conventional method. For example, it can be mixed with various kinds of additives permitted for preparation of a preparation, and molded as a composition. Additives may be added as long as the effects of the present invention are not impaired. For example, in addition to crude drugs, vitamins, minerals, etc., excipients, surfactants, coating agents, oils and fats, stable Agents, preservatives, wetting agents, emulsifiers, lubricants, sweeteners, colorants, flavors, buffers, antioxidants, pH adjusters, acidulants such as citric acid, malic acid, gluconic acid, etc. .

  Examples of the above-mentioned excipients include lactose, starch, cellulose, maltitol, dextrin and the like. Examples of the surfactant include glycerin fatty acid ester, sucrose fatty acid ester and the like. Examples of the coating agent include gelatin, pullulan, shellac, zein and the like. As said fats and oils, a wheat germ oil, rice germ oil, safflower oil etc. can be mentioned, for example. As said wax, a beeswax, a rice bran wax, a carnauba wax etc. can be mentioned, for example. As the above-mentioned sweetener, sucrose, glucose, fructose, stevia, saccharin, sucralose and the like can be mentioned.

  Examples of the above crude drug include ginseng, American ginseng, Chinese ginseng, Ginseng, Reishi, propolis, Agaricus, blueberries, ginkgo biloba and extracts thereof and the like. Examples of the vitamin include oil-soluble vitamins such as vitamin D and K, and water-soluble vitamins such as vitamins B1, B2, B6, B12, C, niacin, pantothenic acid, folic acid, and biotin.

  Further, according to the present invention, there is provided a food or drink or a pharmaceutical comprising an agent for promoting ATP production, which comprises as an active ingredient a specific ingredient contained in young barley leaves or young barley leaves. Here, "comprising" may include a physiologically acceptable carrier according to the desired product form, and also means the case of containing other auxiliary components that can be used in combination.

  In the present invention, the food and drink are not pharmaceuticals, and are not particularly limited as long as they can be taken orally as described above. Specifically, for example, instant foods such as instant noodles, retort foods, canned foods, microwave foods, instant soups and miso soups, freeze-dried foods, etc .; soft drinks, fruit juices, vegetable drinks, soy milk drinks, coffee drinks, tea Beverages such as beverages, powdered beverages, concentrated beverages, alcoholic beverages, etc .; Bread products such as pasta, noodles, cake mix, fried chicken powder, bread crumbs, etc .; chewing, caramel, chewing gum, chocolate, cookies, biscuits, cakes, pies, Sweets such as snacks, crackers, Japanese sweets, dessert sweets; sauces, tomato processed seasonings, flavored seasonings, cooking mixes, sauces, dressings, soups, seasonings such as curry and stew; processed fats and oils, Fats and oils such as butter, margarine, mayonnaise; Milk drinks, yogurts, lactic acid bacteria drinks, ice creams, creams, etc. Dairy products; processed fish products such as processed egg products, fish meat ham and sausages, fish paste products, etc .; processed livestock products such as meat meat ham and sausages; processed agricultural products such as canned agricultural products, jams and marmalades, pickles, boiled beans and cereals; Food etc. may be mentioned.

  Foods and beverages include health foods (eg, functional foods, nutraceuticals, health supplements, fortified foods, nutraceuticals, supplements, etc.), health food (eg, food for specified health, nutrition food, Diseases or conditions caused by increased lipid absorption or decreased ATP production, as well as functional labeling foods etc., special-purpose foods (eg, food for sick persons, infant formula, etc.) Also included are food and drink classifications with an indication of risk reduction, prevention or amelioration (symptoms).

  According to another aspect of the present invention, there is provided a food and drink comprising an agent for promoting ATP production, wherein the function of preventing or ameliorating a disease or condition that can be prevented or ameliorated by promoting ATP production in cells is displayed Food and drink can be provided.

  In the present invention, the pharmaceutical preparation is prepared as an oral preparation or a parenteral preparation according to a conventional method using an additive which is acceptable for formulation. In the case of an oral preparation, as described above, it is not particularly limited as long as it is an orally ingestible form. In the case of parenteral preparations, they can be in the form of injections or suppositories. From the viewpoint of simplicity, the preparation is preferably an oral preparation. Acceptable additives for formulation include the same as those described above.

  In the present invention, the content of the active ingredient in the agent for promoting ATP production is difficult to define uniformly depending on the type and form of the preparation, the purpose of prevention or improvement, etc. However, for example, per adult (body weight 60 kg) It is usually about 15 mg to about 15 g, preferably about 50 mg to about 9 g, as squeezed powder of barley young leaf, and as about 50 mg to about 15 g, preferably about 100 mg to about 9 g as dried powder of barley young leaf. Furthermore, in order to be able to ingest (take) the necessary daily intake of the active ingredient, the content in the preparation may be set appropriately in consideration of the daily intake. In addition, the content of the specific component may be an amount that is included in the squeezed powder or dried powder of the young barley leaves.

  Hereinafter, the present invention will be more specifically described by way of examples, but the technical scope of the present invention is not limited to these examples.

[Example 1] Production of barley young leaf squeezed solution According to the method described in JP-A-2009-148213 (Patent Document 10), 10 kg of harvested barley young leaf (stem and leaf) was squeezed to obtain 8 kg of squeezed solution .

[Example 2] Functional evaluation of barley young leaf squeezing fluid 2-1. Introduction Long chain fatty acids are important fuels for energy production during skeletal muscle contraction. Saturation of palmitic acid has been shown to induce lipotoxicity, such as decreased ATP in muscle cells. Unsaturated fatty acids such as oleic acid have been reported to abolish palmitic acid-induced lipotoxicity by converting palmitic acid in fat droplets and to increase mitochondrial fatty acid oxidation (Reference [Reference [ 1) and [2]).

  In order to evaluate lipid metabolism and function in mitochondria in myocytes, in vitro palmitic acid-induced reduction of ATP in rat L6 myotubes was performed in vitro for functional screening of extracts from natural products (young barley leaves) It carried out as a model.

References
[1] Yuzefovych L .; Wilson G .; Rachek L. Different effects vs. palmitate on mitochondrial function, apoptosis, and insulin signaling in L6 skeletal muscle cells: role of oxidative stress. Am J Physiol Endocrin Metab. 2010, , E1096-1105.
[2] Henique C .; Mansouri A .; Fumey G .; Lenoir V .; Girard J .; Bouillaud F .; Prip-Buus C .; Cohen I. Increased Mitochondrial fatty acid oxidation is sufficient to protect skeletal muscle cells from a palmate J-Biol Chem. 2010, 285, 36818-36827.

2-2. Myoblast Culture The rat L6 myoblast culture was cultured according to the procedure in high glucose DMEM (Dulbecco's modified Eagle medium) containing 10% FBS. For differentiation into myotubes, when myoblasts reached confluency, they were cultured in medium for differentiation (high glucose DMEM containing 3% horse serum and insulin at 50 ng / ml concentration). The culture medium for differentiation was frequently replaced. Myotubes were differentiated for 6 to 8 days and then used for various studies.

2-3. Cytotoxicity test (MTT assay)
MTT assay was performed as a cytotoxicity test. The MTT solution was added to the differentiation medium at 0.5 mg / ml and incubated for 30 minutes. The converted formazan blue crystals were dissolved in acidic isopropanol (isopropanol containing 0.04 M hydrochloric acid), and the absorbance at 570 nm was measured.

2-4. ATP Assay ATP assay was measured using a molecular probe ATP detection kit or its equivalent. Myotubes were first washed with 0.5 ml of ice cold PBS. The cells were then lysed with 0.5 ml of ice cold cell lysate (0.5% NP-40). After centrifuging at 12,000 g and 4 ° C. for 10 minutes to precipitate insoluble material, 5 μl of each was dispensed, 50 μl of the reaction solution was added, and the mixture was incubated at room temperature for 20 minutes. The luminescence intensity of bioluminescence was measured and quantified from the standard curve of the ATP standard solution. All measurements were repeated at least twice.

2-5. Fractionation of barley young leaf extract (squeezed solution) 3.5 liters of barley young leaf extract (squeezed solution) was filtered by suction filtration, and the same amount of ethyl acetate was added to the filtrate to carry out liquid-liquid distribution. Recovered 10.4 g of solid from the ethyl acetate layer (hereinafter referred to as "BLE"), 39.4 g from the aqueous layer (hereinafter referred to as "BLW"), and 16.3 g of solid from the intermediate layer (hereinafter referred to as "BLG"). (Figure 1).

2-6. MTT Assay Result The results of measuring the cytotoxicity of BLE and BLW by MTT assay are shown in FIG. 2 (FIG. 2 (A), FIG. 2 (B)). No cytotoxicity was observed in any of the fractions. In addition, the result of FIG. 2 is an average value of 2 examples.

2-7. ATP production amount The results of ATP assay of BLE and BLW are shown in FIG. 3 (FIG. 3 (A), FIG. 3 (B)). As apparent from FIG. 3, the amount of ATP produced by BLE and BLW showed a concentration-dependent increase. In addition, the result of FIG. 3 is an average value of 4 examples. Moreover, in FIG. 3, "*" and "**" respectively show "p <0.05" and "p <0.01" in Student's t-test (Student's t-test).

2-8. HPLC analysis of BLE First, the fraction was advanced from BLE under the following conditions, and fractionated into 26 fractions by HPLC. The results are shown in FIG.
HPLC conditions Column: CAPCELL PAK C18, 250 mm × 10 mm, 5 μm
Gradient: 0 to 3 minutes, methanol: water = 35: 65
After 100% methanol by 30 minutes, hold for 15 minutes
Conditioning for 5 minutes with methanol: water = 35: 65 Detection wavelength: 210 nm
Flow rate: 4 ml / min

2-9. ATP production amount of HPLC fraction Of the obtained 26 fractions, the ATP production amount up to 3 to 20 fractions was evaluated. The results are shown in FIG. The BLE in FIG. 5 is the sum of the fractions other than the fraction fractionated in FIG. 4 as other fractions. As is clear from FIG. 5, an increase in the amount of ATP production was confirmed in the BLE-3, 5, 6, 8, 9, 10, 11, 12, 13, 14, 15 fractions. In particular, BLE-5, 6, 13, 14, 15 showed an increase of 50% or more.

The component concentrations (μg / ml) and the number of examples (n) of the fractions in FIGS. 5 (A) to (D) are as follows.
(A) 25 μg / ml, n = 3
(B) 25 μg / ml, n = 4
(C) 25 μg / ml, n = 4
(D) 20 μg / ml, n = 4
Moreover, in FIG. 5, "*" and "**" respectively show the significant difference in Student's t test (Student's t-test), "p <0.05" and "p <0.01".

[Example 3] Analysis of active ingredient 3-1. LC-MS / MS analysis of BLE-5 The analysis of BLE-5 by LC-MS / MS showed that robinin (kaempferol-3-O-robinoside-7-O) was obtained from the molecular weight and fragment ion obtained. -rhamnoside) was estimated to be. The analysis results are shown in FIG.

3-2. LC-MS / MS Analysis of BLE-6 The BLE-6 was analyzed by LC-MS / MS. As a result, it was estimated from the obtained molecular weight and fragment ion to be saponarin. The analysis results are shown in FIG.

3-3. LC-MS / MS Analysis of BLE-8 The analysis of BLE-8 by LC-MS / MS was performed, but the obtained molecular weight and fragment ion could not estimate the chemical structure of the component. The analysis results are shown in FIG.

3-4. LC-MS / MS analysis of BLE-9 The analysis of BLE-9 by LC-MS / MS showed that it was afzelin (afemperin; 3-emperol 3-rhamnoside) from the obtained molecular weight and fragment ion The The analysis results are shown in FIG.

Claims (5)

  An agent for promoting ATP production, which contains as an active ingredient a specific ingredient contained in young barley leaves or young barley leaves.   The agent for promoting ATP production according to claim 1, wherein young barley leaves are squeezed powder or dried powder of young barley leaves. The ATP according to claim 1 or 2, wherein the specific component contained in young barley leaves comprises at least one compound selected from the group consisting of the following compounds (a), (b), (c) and (d): Production promoting agent.
(A) Composition _formula: C _{33 H} 40 _{O 19,} molecular weight: 740, compound name: robinin (b) the composition _formula: C _{27 H} 30 _{O 15,} molecular weight: 594, compound name: saponarin (c) the composition _{formula: C 25} H ₃₇ O ₅ N ₂ , molecular weight: 445
(D) Composition formula: C ₂₁ H ₂₀ O ₁₀ , molecular weight: 432, compound name: afzelin An agent for promoting ATP production, which comprises as an active ingredient at least one compound selected from the group consisting of the following compounds (a), (b), (c) and (d):
(A) Composition _formula: C _{33 H} 40 _{O 19,} molecular weight: 740, compound name: robinin (b) the composition _formula: C _{27 H} 30 _{O 15,} molecular weight: 594, compound name: from saponarin (c) young barley Compound formula: C ₂₅ H ₃₇ O ₅ N ₂ , molecular weight: 445 compound (d) Composition formula: C ₂₁ H ₂₀ O ₁₀ , molecular weight: 432, compound name: afzelin A food or drink for the prevention or amelioration (treatment) of a disease or condition (symptoms) caused by a decrease in ATP production, comprising the agent for promoting ATP production according to any one of claims 1 to 4.