JP7088779B2 - Sirtuin protein activation composition - Google Patents

Description

The present invention relates to a sirtuin protein activating composition and a food or drink or a pharmaceutical product using the same.

It has been elucidated that the sirtuin gene activated by calorie intake restriction has a function of extending the life span, and is attracting attention as a longevity gene. Research is also underway on methods for activating the sirtuin gene. For example, by ingesting resveratrol, which is a component contained in red wine, the sirtuin gene is activated and the life span of mice fed a high-calorie diet is extended. It has been reported that it can be extended (Non-Patent Document 1).

In recent years, in the fields of cosmetics and supplements, products claiming beauty or health promotion have been increasing, for example, "action on skin" (tension, luster, beautiful skin, etc.) and "action on joints such as knees" (improvement of joint pain). Etc.) are on the market in many products. In such fields, activation of the sirtuin gene is expected as a new appealing point, and sales of supplements containing resveratrol, which is known as a substance that activates the sirtuin gene, are increasing. Therefore, expectations for the development of materials that activate the sirtuin gene are increasing.

The life-prolonging effect of sirtuins is believed to be due to various actions brought about by increased sirtuin expression or activation of sirtuin proteins. Such actions include an action of suppressing physiological functional decline due to aging (Non-Patent Document 2), an action of recovering a disorder due to autoimmunity (Non-Patent Document 3), and cell proliferation caused by a defect in the telomea gene. Action to recover from dysfunction, recovery from disorders caused by neurodegenerative diseases (eg Parkinson's disease, Alzheimer's disease, Huntington's disease, muscular atrophic lateral sclerosis), recovery from hypofunction of neurotrophic factor genes in the brain , The action of improving learning and memory ability (Non-Patent Document 4) and the like are considered.

Alkylresorcinol is a compound contained in many plants and fungi such as wheat and rye. Alkylresorcinol is known as an inhibitor of intestinal carbohydrate hydrolase (Patent Document 1), and has been suggested to be involved in the nutritional inhibitory effect of rye (Non-Patent Document 5). More recently, it has become clear that alkylresorcinol directly activates sirtuin proteins and contributes to aging suppression (Patent Document 2 and Non-Patent Document 6).

Japanese Unexamined Patent Publication No. 5-170645 Japanese Unexamined Patent Publication No. 2013-249260

Baur et al, Nature, 444: 337-342 (2006) Colak Y et al, Med Sci Monit. May; 17 (5): HY5-9 (2011) Kong S et al, Immunol Cell Biol. Jan; 90 (1): 6-13 (2012) Carafa V et al, Front Pharmacol. 3: 4. Epub (2012) Ross et al, Nutrition Review, 62: 81-95 (2004) Imai et al, Scientific Reports, 7:43679 (2017)

However, there are compounds with different chain lengths and structures in alkylresorcinol, and the correlation between their structure and activation function has not yet been fully clarified. It has been reported that wheat and rye contain a large amount of alkylresorcinol, but it is not yet fully known in barley.

Therefore, the present invention is a novel composition having a sirtuin protein active ability, which comprises grains derived from a specific barley variety or a pulverized product or an extract thereof, and a sirtuin protein activating composition containing an alkylresorcinol having a specific alkyl chain length. It is an object of the present invention to provide a method for producing an alkylresorcinol using a product and a specific barley variety.

As a result of diligent research to solve the above problems, the present inventor has found that barley that suppresses starch synthase II is rich in alkylresorcinol and has excellent sirtuin protein activity and histone deacetylation ability. I found. In addition, they have found that alkylresorcinol having a specific alkyl chain length has excellent sirtuin protein activity and histone deacetylation ability, and have completed the present invention. That is, the present invention includes the following inventions.

[1] A sirtuin protein containing barley grains or crushed products or extracts thereof in which the expression level and / or activity of starch synthase II is suppressed, and the grains or crushed products or extracts thereof contain alkylresorcinol. Activation composition.
[2] A sirtuin protein activating composition comprising 5-pentacosyl resorcinol.
[3] Under the following measurement conditions, the fraction has a peak top column retention time of 22.1 to 22.5 minutes in the chromatogram, and ± 0. With respect to the peak top column retention time. A sirtuin protein activating composition comprising an alkylresorcinol contained in a fraction in the range of 4 minutes.
<Measurement condition>
Measuring method: Reversed phase high performance liquid chromatography column: GL science Inertsil ODS4 (filler particle diameter 5 μm, inner diameter 4.6 mm × height 250 mm)
Mobile phase (Liquid A): Water Mobile phase (Liquid B): Methanol elution gradient: 0 → 5 minutes (Liquid A: 11 → 8% by mass, Liquid B: 89 → 92% by mass) 5 → 30 minutes (Liquid A) : 8 → 0% by mass, Liquid B: 92 → 100% by mass)
Flow velocity: 1.0 mL / min Detection method: UV (275 nm)
Injection volume: 10 μL

[4] A food or drink containing the composition according to any one of [1] to [3].
[5] A pharmaceutical product containing the composition according to any one of [1] to [3].
[6] An anti-aging agent comprising the composition according to [2] or [3].
[7] A method for producing an alkylresorcinol, which comprises a step of extracting an alkylresorcinol from barley in which the expression level and / or activity of starch synthase II is suppressed.
[8] The production method according to [7], wherein the alkyl chain length of the alkylresorcinol is C19, C21, C23 or C25.
[9] The production method according to [8], wherein the alkylresorcinol has an alkyl chain length of C25.
[10] De-histone containing barley grains or crushed products or extracts thereof in which the expression level and / or activity of starch synthase II is suppressed, and the grains or crushed products or extracts thereof contain alkylresorcinol. Acetylation promoting composition.
[11] A histone deacetylation promoting composition comprising 5-pentacosyl resorcinol.

[12] Under the following measurement conditions, the fraction has a peak top column retention time of 22.1 to 22.5 minutes in the chromatogram, and ± 0. With respect to the peak top column retention time. A histone deacetylation promoting composition comprising an alkylresorcinol contained in a fraction in the range of 4 minutes.
<Measurement condition>
Measuring method: Reversed phase high performance liquid chromatography column: GL science Inertsil ODS4 (filler particle diameter 5 μm, inner diameter 4.6 mm × height 250 mm)
Mobile phase (Liquid A): Water Mobile phase (Liquid B): Methanol elution gradient: 0 → 5 minutes (Liquid A: 11 → 8% by mass, Liquid B: 89 → 92% by mass) 5 → 30 minutes (Liquid A) : 8 → 0% by mass, Liquid B: 92 → 100% by mass)
Flow velocity: 1.0 mL / min Detection method: UV (275 nm)
Injection volume: 10 μL

According to the present invention, a composition relating to barley having a sirtuin protein activity ability and a histone deacetylation ability, and foods and drinks and pharmaceuticals using such a composition can be obtained. Further, among the alkylresorcinols known to have sirtuin protein activity, a composition containing an alkylresorcinol having a specific alkyl chain length of the present invention provides excellent sirtuin protein activity and histone deacetylation ability. Will be done. Further, by the production method using the specific barley of the present invention, alkylresorcinol can be obtained without relying on chemical synthesis.

It is a chromatogram showing the peak of alkylresorcinol of each chain length contained in SSII-suppressed barley. It is a graph which shows the alkylresorcinol content of each chain length of SSII suppressed barley and Shunrai. It is a graph which shows the histone deacetylation amount of SSII-suppressed barley and control. It is a graph which shows the histone deacetylation amount of the alkylresorcinol of each chain length.

The sirtuin protein activation composition of the present invention (hereinafter, also referred to as “composition of the present invention”) is a barley grain or a pulverized product or an extract thereof in which the expression level and / or activity of starch synthase II is suppressed. It is characterized in that the grain or a pulverized product or an extract thereof contains an alkylresorcinol.

In the present invention, the "sirtuin protein" means any of seven types of proteins from SIRT1 to SIRT7 constituting the sirtuin family, and is not particularly limited, but is particularly limited in terms of histone deacetylase activity. SIRT1 is preferred.

"Barley in which the expression level and / or activity of starch synthase II is suppressed" in the present invention is barley disclosed in Japanese Patent Publication No. 2004-512427 and Japanese Patent Publication No. 2013-500022 (the present specification). Then, it is also called "SSII-suppressed barley"). Specifically, SSII-suppressed barley is compared to the wild SSII gene by introducing a mutation or exogenous nucleic acid molecule in the endogenous starch synthase II gene that down-regulates the expression of the starch synthase II gene. A method for introducing a mutation that reduces the expression of the SSII gene, or a method for introducing a mutation that causes the expression of a starch synthase II polypeptide that has lower synthetic activity than the wild-type starch synthase II polypeptide in barley. Can be obtained using. As a commercial product, it can be obtained as a breed-improved barley barley max (registered trademark). Starch synthase IIa, starch synthase IIb and starch synthase IIc are known as isoforms of starch synthase II, but starch synthase IIa is suppressed from the viewpoint of phenotype of barley grain. It is preferable to be.

In the present invention, the expression level and / or activity of the starch synthase II of the barley is "suppressed", which means that the expression level and / or the enzyme activity of the starch synthase II is unmodified (wild type). It means a state in which the expression level of the protein and / or the enzyme activity in the granules is reduced to at least 40%, at least 60%, at least 75%, at least 90% or at least 95%.

In the present invention, the "barley grain" means a barley grain with a hull after cultivation and harvesting, or a barley grain with the hull removed after milling. In the present invention, from the viewpoint of processability, barley grains in a state where the outer skin after milling is removed are preferable.

In the present invention, the "crushed grain" of barley refers to the solid substance in which the grain of barley is crushed, and preferably, the barley grain in a state where the hull after milling is removed and the bran is attached is used. It is crushed whole grain flour. The crushing may be performed manually using a mortar, a kitchen knife, a cutter knife, scissors, etc., but if a large amount of plants are to be processed in a short time, an apparatus is used. Such devices include, for example, mills, hammer grinders, mixers, blenders.

In the present invention, the "grain extract" of barley means a substance extracted and concentrated from barley grains so as to contain an alkylresorcinol component, and can be obtained by a method such as oil extraction or solvent extraction. For example, in one of the chemical separation operation methods, a liquid or solid raw material is brought into contact with a solvent, and the solvent-soluble component contained in the raw material is selectively separated from the solvent-insoluble or sparingly soluble component. Refers to the substance that has been used. Preferably, it is an extract extracted from whole grain flour obtained by crushing barley grains with bran. The extraction operation in the present invention is the above-mentioned general extraction operation, in which the raw material is a solid plant and a fat-soluble solvent is used as the solvent.

In the present invention, the pulverized product or extract obtained by the above pulverization or extraction is further subjected to one or more known techniques such as squeezing, concentration, solid-liquid separation, heat sterilization, and filtration sterilization, if necessary. It can also be processed in combination.

The SSII-suppressed barley contains alkylresorcinols having different chain lengths in a total content of 120 μg / g or more with respect to 1 g of barley grains in a state where the barley is milled by a known method and the outer skin is removed. Although the upper limit is not limited, it usually contains about 500 μg / g, and in some cases, about 450 μg / g. The combined content of the alkylresorcinols having an alkyl chain length of C17, C19, C21, C23 and C25 is usually 120 to 350 μg / g, preferably 150 to 320 μg / g, and more preferably 200 to 200. Contains 300 μg / g.

Since the SSII-suppressed barley in the present invention contains abundant alkylresorcinol as described above, it is also advantageous in the method for producing alkylresorcinol without relying on chemical synthesis. Therefore, the present invention provides a method for producing an alkylresorcinol, which comprises a step of extracting an alkylresorcinol from barley in which the expression level and / or activity of starch synthase II is suppressed. In particular, it is suitable for producing an alkylresorcinol having an alkyl chain length of C19, C21, C23 or C25. In addition, C25 alkylresorcinols are particularly suitable.

In the present invention, "alkylresorcinol" is referred to as the following formula I:

In the formula (I), R is a compound representing a saturated or unsaturated linear or branched chain alkyl group. The number of carbon atoms of the alkyl group represented by R is not limited, but is preferably 1 to 27, more preferably 3 to 27, still more preferably 5 to 27, and particularly preferably 5, 7, 9, 11 , 13, 15, 17, 19, 21, 23, 25 and 27. Preferably, R is a saturated or unsaturated linear alkyl group, more preferably a saturated linear alkyl group. Examples of saturated linear alkyl groups represented by R are methyl, n-propyl, n-pentyl, n-heptyl, n-nonade, n-undecyl, n-tridecylic, n-pentadecyl, n-heptadecyl, n. -Nonadecyl, n-henicosylic acid, n-tridecylic acid, n-pentacosyl, n-heptacosyl and the like can be mentioned.

The position and number of unsaturated bonds of the unsaturated linear alkyl group represented by R are not particularly limited. Examples of the unsaturated linear alkyl group include an alkyl group having an unsaturated bond at an arbitrary position on the carbon chain of the saturated linear alkyl group described above. Further, the position and number of branches of the saturated or unsaturated branched chain alkyl group represented by R, and the position and number of unsaturated bonds are not particularly limited.

Preferred examples of the alkylresorcinol of the above formula (I) include the following (the number after C represents the number of carbon chains in the alkyl group and the number after the colon represents the number of unsaturated bonds. In the book, each compound is also referred to as "CX (X is an integer) alkylresorcinol").
5-Pentyl resorcinol [olivetol or 1,3-dihydroxy-5-n-pentylbenzene (C5: 0)]
5-Heptylresorcinol [or 1,3-dihydroxy-5-n-heptylbenzene (C7: 0)]
5-Nonyl resorcinol [or 1,3-dihydroxy-5-n-nonylbenzene (C9: 0)]
5-Undecyl Resorcinol [or 1,3-Dihydroxy-5-n-Undecylbenzene (C11: 0)]
5-Tridecylic resorcinol [or 1,3-dihydroxy-5-n-tridecylic benzene (C13: 0)]
5-Pentadecylic Resorcinol [or 1,3-dihydroxy-5-n-pentadecylicbenzene (C15: 0)]
5-Heptadecylresorcinol [or 1,3-dihydroxy-5-n-heptadecylbenzene (C17: 0)]
5-Nonadecyl resorcinol [or 1,3-dihydroxy-5-n-nonadesylbenzene (C19: 0)]
5-Henikosyl resorcinol [or 1,3-dihydroxy-5-n-Henikosylbenzene (C21: 0)]
5-Tricosyl resorcinol [or 1,3-dihydroxy-5-n-tricosylbenzene (C23: 0)]
5-Pentacocil resorcinol [or 1,3-dihydroxy-5-n-pentacosylbenzene (C25: 0)]
5-Heptacosyl resorcinol [or 1,3-dihydroxy-5-n-heptacosylbenzene (C27: 0)]

The present invention also provides a sirtuin protein active composition containing 5-pentacosylresorcinol (C25) among the alkylresorcinols of the above formula (I). 5-Pentacosyl resorcinol has a very strong histone deacetylation activity, as shown in the examples below. The histone deacetylation activity caused by the alkylresorcinol of each chain length is sirtuin protein SIRT1 as demonstrated by JP2013-249260A and Imai et al, Scientific Reports, 7:43679 (2017). Is known to be caused by activation.

Further, in the present invention, under the following measurement conditions, the peak top column retention time in the chromatogram is a fraction between 22.1 and 22.5 minutes, and the peak top column retention time is relative to the peak top column retention time. Also provided is a sirtuin protein activating composition comprising an alkylresorcinol contained in a fraction in the ± 0.4 minute range:
<Measurement condition>
Measuring method: Reversed phase high performance liquid chromatography column: GL science Inertsil ODS4 (filler particle diameter 5 μm, inner diameter 4.6 mm × height 250 mm)
Mobile phase (Liquid A): Water Mobile phase (Liquid B): Methanol elution gradient: 0 → 5 minutes (Liquid A: 11 → 8% by mass, Liquid B: 89 → 92% by mass) 5 → 30 minutes (Liquid A) : 8 → 0% by mass, Liquid B: 92 → 100% by mass)
Flow velocity: 1.0 mL / min Detection method: UV (275 nm)
Injection volume: 10 μL
Injection sample: Extract obtained through the following steps (1) A step of adding 400 ml of ethanol to 20 g of powder obtained by subjecting barley grains from which the hull has been removed to a crusher and stirring at room temperature for 24 hours.
(2) A step of filtering the obtained dispersion with a filter paper to obtain an extract.
(3) A step of concentrating the extract under reduced pressure, adding 50 ml of hexane to the remaining residue, and filtering to obtain a filtrate.
(4) A step of concentrating the obtained filtrate again under reduced pressure and dissolving the residue in 20 ml of methanol to obtain an extract.

The alkylresorcinol of the above formula (I) can be extracted from a plant by a conventional method. Further, the alkylresorcinol of the above formula (I) can also be produced by a chemical synthesis method with reference to the pamphlet of International Publication No. WO2007 / 77770.

As a method for extracting the alkylresorcinol of the formula (I) from barley, a method of immersing barley or a fermented product thereof in an extraction solvent at room temperature or warmed under normal pressure or pressure with stirring as necessary, reflux extraction Law etc. can be mentioned. The plant may be cut, ground, squeezed, dried, or a combination thereof, if necessary, before being added to the extraction solvent.

Examples of the extraction solvent include lower alcohols such as methanol, ethanol, n-propanol, isopropanol and n-butanol, and alcohols which are liquid at room temperature such as polyhydric alcohols such as 1,3-butylene glycol, propylene glycol and glycerin. Ethers such as diethyl ether and propyl ether; esters such as butyl acetate and ethyl acetate; ketones such as acetone and ethyl methyl ketone; hexane; and organic solvents such as chloroform. These solvents may be used alone or in combination of two or more. Among the above solvents, alcohols that are liquid at room temperature, for example, lower alcohols having 1 to 4 carbon atoms are preferably used from the viewpoint of operability and the influence on the environment, and from the viewpoint of safety due to the residual solvent. It is more preferable to use ethanol.

The organic solvent may also include a hydrous organic solvent containing an aqueous component. From the viewpoint of maintaining high extraction efficiency, it is desirable that the content of the aqueous component in the hydrous organic solvent is usually 50% by volume or less, preferably 30% by volume or less, and more preferably 20% by volume or less. Examples of the hydrous organic solvent include hydrous alcohols in which an aqueous component is further contained in alcohols that are liquid at room temperature as described above, and more preferably hydrous ethanol.

The temperature and time for extraction can be appropriately set depending on the type of extraction solvent used, extraction conditions, etc., but the extraction temperature is preferably 2 to 100 ° C. and the extraction time is preferably about 30 minutes to 60 hours. .. The amount of the extraction solvent may be preferably about 200 to 2000 parts by mass with respect to 100 parts by mass of the barley or its fermented product.

The mixture containing the extract and the residue is subjected to filtration or centrifugation as necessary, and the solid component is removed to obtain an extract containing the alkylresorcinol of the formula (I). The removed solid component can be subjected to an extraction operation using an extraction solvent again, and this operation may be repeated several times. Further, the obtained extract may be further purified by liquid chromatography or the like. The extract containing the alkylresorcinol of the formula (I) thus obtained can be concentrated, freeze-dried, hot-air dried, pulverized, powdered, classified, diluted, mixed with water, etc. as it is or as needed. After being treated, it can be used in the present invention.

The alkylresorcinol of the formula (I) has an action of activating the sirtuin protein SIRT1 and further animal as shown in JP2013-249260A and Imai et al, Scientific Reports, 7:43679 (2017). It has the effect of extending the lifespan of an individual. Therefore, the alkylresorcinol of the formula (I) can be used for sirtuin activation, life extension, aging suppression, prevention or amelioration of diseases and conditions that may occur with aging, and the like.

The present invention also provides an anti-aging agent comprising a sirtuin protein activating composition comprising an alkylresorcinol of formula (I), in particular 5-pentacosylresorcinol. The present invention also provides a life-prolonging agent containing an alkylresorcinol of the formula (I), particularly 5-pentacosylresorcinol, as an active ingredient. The present invention also provides an agent for preventing or ameliorating a disease or condition that may occur with aging, which comprises an alkylresorcinol of the formula (I), particularly 5-pentacosylresorcinol, as an active ingredient.

In the present invention, "aging" refers to a decrease in the physiological function of an individual animal that occurs with aging, for example, a decrease in sight or hearing, a decrease in motor ability, a decrease in homeostatic function, a decrease in immune function, a brain such as memory impairment. Examples include functional deterioration and functional deterioration of each organ in the living body.

In the present invention, "suppression of aging" means suppression of deterioration of the physiological function of an individual animal that occurs with aging, suppression of diseases and disorders caused by the deterioration of the physiological function, and suppression of death of an individual due to the progress of aging (that is, suppression of aging). It is a concept that includes (extension of life).

Further, in the present invention, the "disease or condition that may occur with aging" refers to various diseases or conditions caused by the above-mentioned deterioration of the physiological function of an individual animal that occurs with aging, such as disorders due to autoimmunity and defects in the telomea gene. Defects in cell proliferation caused by, disorders caused by neurodegenerative diseases (eg Parkinson's disease, Alzheimer's disease, Huntington's disease, muscular atrophic lateral sclerosis), decreased function of neurotrophic factor genes in the brain, learning, and decreased memory capacity And so on.

Since the composition of the present invention is derived from barley, which has abundant eating experience, there is little concern that it will have a harmful effect on the living body even if it is continuously ingested for a long period of time, and it has safety. Therefore, the composition of the present invention can be safely and continuously inoculated not only to healthy persons and adults but also to elderly people or sick persons who particularly need the present invention.

The composition of the present invention can also be used as a pharmaceutical product, food or drink or feed for humans or animals. The drug, food or drink or feed can be used for sirtuin activation, life extension, aging suppression, prevention or improvement of diseases and conditions that may occur with aging, and the like.

The pharmaceutical product may be a sirtuin activator, a life-prolonging agent, an aging inhibitor, a preventive or ameliorating agent for a disease or condition that may occur with aging, or the like, which contains the composition of the present invention. The dosage forms of the above-mentioned pharmaceutical products include, for example, oral preparations such as tablets, capsules, granules, powders, syrups, dry syrups, liquids and suspensions, and enteral preparations such as inhalants, transdermal preparations and suppositories. Examples thereof include parenteral preparations such as pharmaceuticals, infusions, and injections. Liquid preparations such as the above liquids and suspensions may be dissolved or suspended in water or other suitable media immediately before administration, and the above tablets and granules are coated on the surface by a well-known method. May be. Further, the injection may be a solution of sterile distilled water or sterile physiological saline containing a lysis aid, if necessary.

In addition to the composition of the present invention, the above-mentioned pharmaceutical product may contain various pharmaceutically acceptable carriers such as excipients, stabilizers, other additives and the like, if necessary, or may be contained. In addition, other medicinal ingredients such as various vitamins, minerals, crude drugs and the like may be contained. The pharmaceutical product can be produced according to a conventional method by blending the above-mentioned carrier and other medicinal ingredients with the composition of the present invention.

The above-mentioned food or drink or feed contains the composition of the present invention, and may have effects such as activation of sirtuin, extension of life span, suppression of aging, prevention or improvement of diseases and conditions that may occur with aging, and the like. It may also be used as a health food, a functional food or drink, a food or drink for specified health use, a food or drink for the sick, a livestock, a race horse, a feed for an ornamental animal, a pet food or the like.

The forms of the food and drink and the feed are not particularly limited, and include all forms in which the composition of the present invention can be blended. For example, the form may be solid, semi-solid or liquid, or may include various forms such as tablets, chewable tablets, powders, capsules, granules, drinks, gels, syrups, and liquid foods for enteral feeding. ..

Specific examples of foods and drinks include green tea, tea beverages such as oolong tea and tea, coffee beverages, soft beverages, jelly beverages, sports beverages, dairy beverages, carbonated beverages, fruit juice beverages, lactic acid bacteria beverages, fermented dairy beverages, and powdered beverages. , Cocoa beverages, alcoholic beverages, purified water and other beverages, butter, jam, sprinkles, margarine and other spreads, mayonnaise, shortening, cream, dressings, breads, rice, noodles, pasta, miso soup, tofu, milk, Yogurt, soups or sauces, bakeries (bread, pie, cakes, cookies, biscuits, crackers, etc.), confectionery (biscuits and cookies, chocolate, candy, cakes, ice cream, chewing gum, tablets, etc.), nutritional supplements ( Supplements in the form of rounds, tablets, jellies or capsules, granola-like serials, granola-like snake bars, serial bars) and the like.

Since the feed can be used in almost the same composition and form as the food and drink, the description of the food and drink in the present specification can be applied to the feed as well.

The above-mentioned foods and drinks and feeds are added to the composition of the present invention by other food and drink materials used for producing foods and drinks and feeds, various nutrients, various vitamins, minerals, amino acids, various fats and oils, and various additives (taste components). , Sweeteners, acidulants such as organic acids, surfactants, pH adjusters, stabilizers, antioxidants, pigments, flavors, etc.) can be blended and produced according to a conventional method. Alternatively, the food or drink or feed according to the present invention can be produced by blending the composition of the present invention with the food or drink or feed that is normally eaten.

The content of the composition of the present invention in the above-mentioned pharmaceutical products, foods and drinks or feeds may be any amount as long as it can obtain desired effects such as sirtuin activating effect, life extension effect and aging suppressing effect. It can be appropriately changed depending on the form of the product, the species of the individual to be administered or ingested, symptoms, age, sex and the like.

Hereinafter, the present invention will be specifically described with reference to Examples and Comparative Examples, but this does not limit the scope of the present invention.

[Test Example 1] Method for Extracting Alkylresorcinol from Barley and Method for Analysis 100 g of barley grains were pulverized into powder. To 20 g of the obtained powder, 400 ml of ethanol was added, and the mixture was stirred at room temperature for 24 hours. The obtained dispersion was filtered through a filter paper to obtain an extract. Subsequently, the extract was concentrated under reduced pressure, 50 ml of hexane was added to the remaining residue, and the mixture was filtered to obtain a filtrate. This was concentrated again under reduced pressure, and the residue was dissolved in 20 ml of methanol to obtain an extract. This was analyzed by reverse phase high performance liquid chromatography under the following conditions.
Measuring method: Reversed phase high performance liquid chromatography Sample: Barley extract Apply amount: 10 μL
Flow velocity: 1 mL / min Mobile phase (Liquid A): Water mobile phase (Liquid B): Methanol elution gradient: 0 → 5 min (Liquid A: 11 → 8% by mass, Liquid B: 89 → 92% by mass), 5 → 30 minutes (Liquid A: 8 → 0% by mass, Liquid B: 92 → 100% by mass)
Detection wavelength: 275nm
Column: GL science Inertsil ODS4 (filler particle size 5 μm, inner diameter 4.6 mm x height 250 mm)
Silica gel: 3 series high-purity spherical silica gel (surface area: 450 m ² / g, pore diameter: 100 angstrom, pore volume: 1.05 mL / g, chemical bond group: octadecyl group, end cap treatment: yes, carbon content: 11% )
The quantification was calculated based on the calibration curve of the peak area and the concentration of olivetol (C5) prepared in advance by the peak area of the chromatogram at the wavelength of 275 nm. Peak identification was performed by column retention time and UV spectrum, and the structure was identified by mass spectrum.

(Example 1)
Using barley grains with bran obtained by milling Australian bare barley (Healthy Grain, barley variety Burley Max®; equivalent to "SSII-suppressed barley" herein) and removing the hulls. The chromatogram obtained by obtaining the extract according to Test Example 1 and chromatographically analyzed is shown in FIG. 1, and the result of each content is shown in FIG.

From the chromatogram of FIG. 1, it was revealed that alkylresorcinols of C17, C19, C21, C23 and C25 having no unsaturated bond in the alkyl chain were detected, and the proportion of C25 was particularly high. In addition, peaks of unknown detailed structure having the same UV spectrum as alkylresorcinol were simultaneously detected at a column retention time of around 22.3 minutes (between 22.1 and 22.5 minutes) (in the present specification, of this peak fraction). Alkylresorcinol is referred to as "RT: 22.3"). The amount of each of these alkylresorcinol components is 5.2 μg / barley 1 g for C17, 48.7 μg / barley 1 g for C19, 40.2 μg / barley 1 g for C21, 33.8 μg / barley 1 g for C23, and 121.6 μg / g for C25. It was 1 g of barley, and the amount of alkylresorcinol (RT: 22.3) of unknown detailed structure was 165.6 μg / 1 g of barley. The total amount of alkylresorcinols of C17, C19, C21, C23 and C25 other than RT: 22.3 is 249.4 μg / barley 1 g, and the total amount of alkylresorcinols containing RT: 22.3 is 415.0 μg / barley 1 g. Met.

(Comparative Example 1)
FIG. 2 shows the results of quantifying each alkylresorcinol component by chromatographic analysis of the extract extracted according to Test Example 1 using barley grains with bran obtained by milling a general barley variety Shunrai and removing the outer skin. C17 was 3.9 μg / barley 1 g, C19 was 11.0 μg / barley 1 g, C21 was 12.4 μg / barley 1 g, C23 was 13.0 μg / barley 1 g, and C25 was 58.2 μg / barley 1 g. The total amount of alkylresorcinols of these C17, C19, C21, C23 and C25 was 98.5 μg / 1 g of barley, and the total amount of alkylresorcinol and the content of each chain length were higher than those of SSII-suppressed barley of Example 1. Both were few.

[Test Example 2] Analysis of the amount of histone deacetylation in cells Using the extract obtained in Test Example 1 above, the histone deacetylation in the human monocytic cell line THP-1 was carried out by the following method. The sirtuin activity was evaluated by measuring the degree.

<Preparation of extract containing alkylresorcinol>
A barley extract and an alkylresorcinol of each chain length were prepared according to Test Example 1, and a dry solid product dissolved in 20 ml of ethanol was used as an extract in the same manner. For olivetol, which is an alkylresorcinol of C5, a reagent manufactured by SIGMA-ALDRICH was used.

<Extraction of total histones>
Human monocyte-like cell line THP-1 was cultured in RPMI1640 medium containing 10% FBS, penicillin, streptomycin, and mercaptoethanol. The cells were dispensed into a 24-well plate at a concentration of 5 × 10 ⁵ / well, and an ethanol solution of the extract was prepared so that the total component of alkylresorcinol in the barley extract was equivalent to 3 μg / ml, and in the case of alkylresorcinol of each chain length. An ethanol solution of the extract was prepared so as to correspond to 10 μg / ml. After adding this extract to the plate, it was cultured at 37 ° C. for 22 hours. For the control, a level at which no extract was added was used. Histone extraction was performed using the EpiQuik ^TM Total Histone Extraction Kit based on the following method recommended by Epigentek.
(1) The cells were centrifuged at 1000 rpm at 4 ° C. for 5 minutes, and the supernatant was removed.
(2) Suspended in 1 mL of 1 × Pre-Lysis Buffer and shaken on ice for 10 minutes.
(3) After centrifugation at 3000 rpm, 4 ° C. for 5 minutes, the supernatant was removed.
(4) The precipitate was suspended in 3 times the amount of Lysis Buffer and allowed to stand on ice for 30 minutes.
(5) After centrifugation at 12000 rpm, 4 ° C. for 5 minutes, the supernatant was transferred to a plastic tube.
(6) A Balance-DTT Buffer 0.3 times the amount of the supernatant was added, and the mixture was stored at -30 ° C.

<Quantification of histones>
The histone was quantified using the TaKaRa BCA Protein Assay Kit based on the following method recommended by Takara Bio Inc.
(1) A BSA standard solution (50, 40, 30, 20, 10, 5 μg / ml) was prepared.
(2) 100 μL of BSA standard solution or 15 μL of histone extract and 85 μL of deionized water were added to each well.
(3) 100 μL of Working Solution was added, mixed and reacted at 60 ° C. for 60 minutes.
(4) The absorbance at 570 nm was measured using a spectrophotometer.
(5) A calibration curve was prepared from the measurement results of the BSA standard solution, and the amount of protein contained in each sample was calculated.
(6) The protein amount of the sample stock solution was determined by multiplying the sample result by a dilution ratio (100/15).

<Quantification of the amount of histone acetylation>
The EpiQuik ^TM Total Histone H3 Acetylation Detection Fast Kit (Colorimetric) was used to quantify the amount of acetylation, and the method was based on the following method recommended by Epigentek.
(1) 50 μL of Antibody Buffer was added to each well.
(2) Standards (100, 50, 25, 12, 6, 3, 1.5 ng / μl) were prepared.
(3) 50 to 200 ng of histone extract sample or standard was added to each well, and the mixture was reacted at room temperature for 2 hours.
(4) Each well was washed 3 times with 150 μL Wash Buffer.
(5) 50 μL of Detection Antibody diluted with Antibody buffer was added, and the mixture was reacted with an orbital shaker at room temperature of 100 rpm for 60 minutes.
(6) Each well was washed 6 times with 150 μL Wash Buffer.
(7) 100 μL of Color Developer was added and the color was developed at room temperature for 2 minutes.
(8) After changing to a medium blue color, 50 μL of Stop Solution was added to stop the reaction.
(9) The absorbance at 450 nm was measured using a spectrophotometer, and the amount of acetylation was calculated from the standard calibration curve.

When the extract of SSII-suppressed barley containing alkylresorcinol obtained according to the above procedure was analyzed according to the above Test Example 2, as shown in FIG. 3, the cells to which the extract of SSII-suppressed barley was added had a histone. It was confirmed that the amount of acetylated histone decreased and deacetylation was progressing. Further, as shown in FIG. 4, the amount of acetylated histone in histone also decreased in the cells to which the extract containing the alkylresorcinol containing the alkyl chain length C25 and the column retention time of about 22.3 minutes was added, and the alkyl chain length was reduced. Compared with olivetol of C5, alkylresorcinol of C25 was 1.8 times, and alkylresorcinol of RT: 22.3 was 1.7 times, and it was confirmed that deacetylation was progressing, which was excellent. It was found to have histone deacetylation ability.

Using olivetol having an alkyl chain length of C5, an extract was obtained according to Test Example 2 above, and the amount of histone deacetylation was analyzed. As shown in FIG. 4, olivetol having an alkyl chain length of C5 had a smaller amount of histones acetylated than the control, and thus deacetylation was progressing, but the alkyl chain length C25 and the column retention time 22. Deacetylation did not proceed when compared to the extract containing alkylresorcinol around 3 minutes.

Claims (8)

A barley grain or a crushed product or extract thereof in which the expression level and / or activity of starch synthase II is suppressed is contained, and the grain or crushed product or extract thereof contains 5- pentacosyl resorcinol. -Composition for activating sirtuin protein. Under the following measurement conditions, the fraction has a peak top column retention time of 22.1 to 22.5 minutes in the chromatogram, and is ± 0.4 minutes with respect to the peak top column retention time. The composition for activating a sirtuin protein according to claim 1 , which comprises an alkylresorcinol contained in a range of fractions.
<Measurement condition>
Measuring method: Reversed phase high performance liquid chromatography column: GL science Inertsil ODS4 (filler particle diameter 5 μm, inner diameter 4.6 mm × height 250 mm)
Mobile phase (Liquid A): Water Mobile phase (Liquid B): Methanol elution gradient: 0 → 5 minutes (Liquid A: 11 → 8% by mass, Liquid B: 89 → 92% by mass) 5 → 30 minutes (Liquid A) : 8 → 0% by mass, Liquid B: 92 → 100% by mass)
Flow velocity: 1.0 mL / min Detection method: UV (275 nm)
Injection volume: 10 μL A food or drink for activating sirtuin protein , which comprises the composition according to claim 1 or 2 . A sirtuin protein activating drug comprising the composition according to claim 1 or 2 . An anti-aging agent comprising the composition according to claim 1 or 2 . A method for producing an alkylresorcinol, which comprises a step of extracting an alkylresorcinol from barley in which the expression level and / or activity of the starch synthase II is suppressed. The production method according to claim 6 , wherein the alkylresorcinol has an alkyl chain length of C19, C21, C23 or C25. The production method according to claim 7 , wherein the alkylresorcinol has an alkyl chain length of C25.

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