Telomerase expression enhancer

Abstract

To provide a telomerase expression enhancer having an excellent telomerase expression enhancing effect.SOLUTION: A telomerase expression enhancer contains at least one material selected from a barley young leaf, a kudzu flower, Kaempferia parviflora, Lagerstroemia Speciosa Linn. or Pers., a Terminalia plant, an oyster, Citrus unshiu peel, and Matteuccia struthiopteris.SELECTED DRAWING: Figure 1

Description

Technical Field

[0001] The present invention relates to a telomerase expression enhancer containing natural materials.

Background Art

[0002] In recent years, against the backdrop of the growing trend towards health and safety, in the food and cosmetics industries, products using natural materials are preferred, and it is desired to make better use of the useful functions of such natural materials.

[0003] For example, young barley leaves are generally used as a raw material for health foods and green juices, and their functionality, such as the effect of improving bowel movements and the antihypertensive effect, has been widely studied (Patent Documents 1 to 3).

Prior Art Documents

Patent Documents

[0004]

Patent Document 1

Patent Document 2

Patent Document 3

Summary of the Invention

Problems to be Solved by the Invention

[0005] An object of the present invention is to provide a telomerase expression enhancer containing natural materials and having an excellent telomerase expression enhancing effect.

Means for Solving the Problems

[0006] As a result of exploring the functions of various natural materials, the present inventors have found that a specific natural material has an excellent telomerase expression enhancing effect, and have thus completed the present invention.

[0007] That is, the present invention is as follows. [1] A telomerase expression enhancer characterized by containing at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, Terminalia, oysters, tangerine peel, and Psoralea corylifolia. [2] The telomerase expression enhancer according to [1], which is an oral preparation. [3] The telomerase expression enhancer according to [1], which is a topical preparation.

Effects of the Invention

[0008] The telomerase expression enhancer of the present invention has an excellent telomerase expression enhancing effect.

Brief Description of the Drawings

[0009]

Figure 1

Modes for Carrying Out the Invention

[0010] The telomerase expression enhancer of the present invention is characterized by containing at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, Terminalia, oysters, tangerine peel, and Psoralea corylifolia. In the telomerase expression enhancer of the present invention, the material may contain one kind, two kinds, or three or more kinds. When two or more kinds are contained, it is preferable to combine those with high individual effects.

[0011] The telomerase expression enhancer of the present invention enhances the expression of telomerase (telomere synthase). Therefore, it can extend the telomeres of cells and prevent cell aging.

[0012] Hereinafter, each material contained in the telomerase expression enhancer of the present invention (hereinafter sometimes referred to as the material of the present invention) will be described.

[0013] [Pine bark] As the raw material pine tree bark used in the present invention, examples include Pinus Martima, Larix kaempferi, Pinus thunbergii, Pinus densiflora, Pinus parviflora, Pinus koraiensis, Pinus sylvestris, Pinus densata, Pinus luchuensis, Pinus taiwanensis, Pinus armandii, and Pinus bungeana. Among these, it is preferable to use Pinus Martima, which has a high telomerase expression enhancing effect.

[0014] In the present invention, pine tree bark can be processed and used. Examples of processed pine tree bark include chip-like substances, crushed products, squeezed juices, extracts, and their dried powders. Considering the formulatability, the pine tree bark used in the present invention is preferably a crushed product, squeezed juice, extract, or their dried powder because it is easy to apply. From the viewpoint of the telomerase expression enhancing effect, it is more preferably an extract or its dried powder. The processed pine tree bark may be manufactured by methods commonly known to those skilled in the art or may be those on the market. For example, the pine tree bark extract manufactured by Toyo Pharmaceutical Co., Ltd. can be used.

[0015] Examples of the extraction solvent used for obtaining the pine tree bark extract include water, organic solvents, and water-containing organic solvents (water-containing alcohols such as water-containing ethanol). Examples of organic solvents include methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, butane, acetone, hexane, cyclohexane, propylene glycol, water-containing ethanol, water-containing propylene glycol, ethyl methyl ketone, glycerin, methyl acetate, ethyl acetate, diethyl ether, dichloromethane, edible oils and fats, 1,1,1,2-tetrafluoroethane, and 1,1,2-trichloroethene. These water and organic solvents may be used alone or in combination. It is preferable to use water as the extraction solvent. Note that the temperature of the solvent during extraction is not limited as long as it is below the boiling point of the solvent used.

[0016] The method for obtaining the pine bark extract is not particularly limited. For example, it includes a heating extraction method, a supercritical fluid extraction method, a liquid carbon dioxide batch method, a liquid carbon dioxide reflux method, a supercritical carbon dioxide reflux method, etc. Also, a plurality of extraction methods may be combined. By combining a plurality of extraction methods, it becomes possible to obtain pine bark extracts with various compositions.

[0017] The supercritical fluid extraction method is a method of performing extraction using a supercritical fluid, which is a fluid in a state exceeding the critical point (critical temperature, critical pressure) of the gas and liquid of a substance. As the supercritical fluid, carbon dioxide, ethylene, propane, nitrous oxide (laughing gas), etc. are used, but carbon dioxide is preferably used.

[0018] In the supercritical fluid extraction method, an extraction step of extracting the target component with the supercritical fluid and a separation step of separating the target component and the supercritical fluid are performed. In the separation step, any of extraction separation by pressure change, extraction separation by temperature change, and extraction separation using an adsorbent or absorbent may be performed.

[0019] Also, supercritical fluid extraction by the entrainer addition method may be performed. This method is a method of adding, for example, ethanol, propanol, n-hexane, acetone, toluene, other lower aliphatic alcohols, aliphatic hydrocarbons, aromatic hydrocarbons, and ketones to the extraction fluid at about 2 to 20 W / V%, and performing supercritical fluid extraction using this fluid, thereby dramatically increasing the solubility of the target extract such as OPC (Oligomeric proanthocyanidin) and catechins in the extraction solvent, or enhancing the selectivity of separation, and is a method for efficiently obtaining a pine bark extract.

[0020] The supercritical fluid extraction method has the advantages that it can be operated at a relatively low temperature, so it can be applied to substances that are altered or decomposed at high temperatures, there is no residue of the extraction fluid, and since the solvent can be recycled, the solvent removal process and the like can be omitted, and the process becomes simple.

[0021] It is preferable from the viewpoint of safety to purify the pine bark extract obtained by the above extraction by a column method or a batch method. Examples of the column method include purification methods using adsorptive carriers such as Diaion HP-20, Sephadex-LH20, and chitin.

[0022] The pine bark extract used in the present invention contains proanthocyanidins as one of the main components. Proanthocyanidins are a group of compounds composed of polycondensates with a degree of polymerization of 2 or more having flavan-3-ol and / or flavan-3,4-diol as structural units.

[0023] The pine bark extract used in the present invention preferably contains a polycondensate with a degree of polymerization of 2 or more as proanthocyanidins. In particular, proanthocyanidins containing a large amount of polycondensates with a low degree of polymerization are preferred. Examples of the polycondensate with a low degree of polymerization include polycondensates with a degree of polymerization of 2 to 30 (2 to 30 mers), preferably polycondensates with a degree of polymerization of 2 to 10 (2 to 10 mers), and more preferably polycondensates with a degree of polymerization of 2 to 4 (2 to 4 mers). In this specification, a polymer with a degree of polymerization of 2 to 4 is referred to as OPC (oligomeric proanthocyanidin). The pine bark extract used in the present invention preferably contains 20% by mass or more of OPC, more preferably 30% by mass or more, and even more preferably 40% by mass or more.

[0024] [Barley young leaves] Barley (Hordeum vulgare L.) is native to Central Asia and is an annual or biennial herb belonging to the Poaceae family. It is roughly classified into two-row barley, six-row barley, etc. according to the ear shape. The barley young leaves used in the present invention are not particularly limited as long as they are commonly available, and young leaves of any barley such as two-row barley and six-row barley may be used. Also, any variety may be used. The barley young leaves only need to contain young leaves of barley, and other parts such as stems may also be contained.

[0025] The form of young barley leaves is not particularly limited. For example, it may be in the state of the freshly harvested young leaves, or it may be something that has been further processed after harvesting the young leaves. Examples of the processing of young barley leaves include drying and powdering, slicing and drying and powdering, juicing and drying and powdering, extract extraction and drying and powdering, etc. The processing methods adopted in the industry can be applied to these. That is, examples of the processed products include chip-like substances, pulverized products, juice, extracts, etc. of young barley leaves, and examples of the pulverized products include powders, granules, etc. As a method of making a pulverized product, for example, a method of drying the leaves and / or stems of barley, coarsely pulverizing them, heating at 110°C or higher, and then finely pulverizing (see Japanese Patent Application Laid-Open No. 2003-033151), or a method of blanching the leaves and / or stems of barley, drying them, and then pulverizing them (see Japanese Patent Application Laid-Open No. 2002-065204), etc. can be mentioned.

[0026] The juice and extract of young barley leaves may be in a liquid state, but can also be used in the form of a paste or a dry powder. The juicing method of young barley leaves is not particularly limited, and the juicing methods commonly used by those skilled in the art can be applied. The extraction, separation, and synthesis methods from young barley leaves are not particularly restricted and can be appropriately selected according to the purpose. For example, methods such as adding extraction solvents commonly used by those skilled in the art, such as ethanol, water, aqueous ethanol, etc., and heating if necessary for extraction can be mentioned.

[0027] Considering the formulatability, the young barley leaves used in the present invention are preferably a pulverized product, a juice product, an extract, or a dry powder thereof because they are easy to apply. From the viewpoint of enhancing the telomerase expression effect, a pulverized product, a juice product, or a dry powder thereof is more preferable. The processed young barley leaf product may be manufactured by a method commonly known to those skilled in the art or may be something that is on the market.

[0028] [Flower of Kudzu Vine] Kudzu is a vine-like perennial herbaceous plant of the genus Pueraria in the legume family. As for the flowers of kudzu, those collected at any stage from the bud to the fully opened flower may be used, or those collected at each stage may be mixed and used. There are no particular restrictions on the type of kudzu, but examples include Pueraria thomsonii, Pueraria lobata, Pueraria thunbergiana, etc.

[0029] In the present invention, the harvested kudzu flowers can be processed and used. Examples of the processed kudzu flower products include chip-like substances, pulverized products, squeezed juices, extracts, and dried powders thereof. Considering the pharmaceutical properties, the kudzu flowers used in the present invention are preferably pulverized products, squeezed juices, extracts, or dried powders thereof because they are easy to apply. From the perspective of the effect of enhancing telomerase expression, extracts or their dried powders are more preferable. The processed kudzu flower products may be those manufactured by methods commonly known to those skilled in the art or those currently on the market.

[0030] The extract of kudzu flowers can be obtained by extraction using an appropriate solvent. Examples of the solvent include water; lower alcohols such as ethanol, methanol, isopropanol, and butanol; lower esters such as ethyl acetate and methyl acetate; acetone; and mixed solvents of these with water. The temperature of the extraction solvent can be appropriately set from room temperature to below the boiling point according to the solvent used. In the present invention, a mixed solvent of an organic solvent and water can be used. Examples of the mixed solvent include mixed solvents of ethanol, methanol, isopropanol, butanol, ethyl acetate, methyl acetate, acetone, and water. The temperature of the extraction solvent can be appropriately set from room temperature to below the boiling point according to the solvent used. In the present invention, hot water extracts are preferred from the perspective of the effect of enhancing telomerase expression.

[0031] [Black Ginger] The black ginger (Kaempferia parviflora) used in the present invention is not particularly limited as long as it is a plant known as a plant of the genus Boesenbergia in the Zingiberaceae family that grows naturally in Southeast Asia and the like. For example, parts such as its roots, stems, leaves, flowers, and branches can be used, but roots and stems are preferred.

[0032] In addition to the unprocessed state of the harvested black ginger, it may also be a processed product obtained by subjecting the harvested black ginger to predetermined processing after harvesting. Examples of the processed product include chip-like substances, pulverized products, squeezed juices, extracts, and dried powders thereof. Considering the pharmaceutical properties, the black ginger used in the present invention is preferably a pulverized product, squeezed juice, extract, or dried powder thereof because it is easy to apply. From the viewpoint of enhancing the telomerase expression effect, it is more preferably an extract or its dried powder. Also, from the viewpoints of ease of processing, storage, transportation, and versatility of the usage form, it is preferably a dried powder. The black ginger processed product may be manufactured by a method commonly known to those skilled in the art or may be a product currently on the market.

[0033] The extract of black ginger is not particularly limited as long as it is a substance in which the components in black ginger are extracted. For example, it includes an extract obtained by extracting the components contained in black ginger with a solvent according to a conventional method, a diluted solution or concentrated solution thereof, or a dried product or powder thereof. For example, examples of the components contained in black ginger include polymethoxyflavonoids such as 5,7-dimethoxyflavone, but are not limited thereto.

[0034] Examples of the solvent used for obtaining the black ginger extract include water such as normal temperature water, warm water, and hot water; lower alcohols such as ethanol, methanol, isopropanol, and butanol; lower esters such as ethyl acetate and methyl acetate; acetone; and mixed solvents of these with water. Examples of the mixed solvent include an acetone / water (2 / 8 to 8 / 2, volume ratio) mixture and an ethanol / water (2 / 8 to 8 / 2, volume ratio) mixture.

[0035] The method for obtaining the black ginger extract is not particularly limited. For example, a solvent with a mass 2 to 20 times that of the black ginger is added thereto, and extraction is carried out under any conditions such as standing, shaking, stirring, refluxing, etc. in the range of 0°C to the reflux temperature of the solvent for several minutes to several tens of hours. After the extraction operation, it is preferable to perform a solid-liquid separation operation such as filtration or centrifugation to remove insoluble solids. By performing operations such as dilution or concentration as necessary, an extract can be obtained. Further, the same operations may be repeated for the insoluble matter for extraction, and the extract thus obtained may be combined with the previous extract for use. These extracts may be further purified and used by a purification method commonly used by those skilled in the art.

[0036] The method for obtaining a dried product from the extract is not particularly limited. For example, the extract or its concentrate may be subjected to a drying treatment commonly used by those skilled in the art such as spray drying, freeze drying, vacuum drying, fluidized drying, etc. Further, the dried product thus obtained can be powdered and used by a method known to those skilled in the art.

[0037] [Banaba] Banaba (Lagerstroemia Speciosa, Linn. or Pers.) belongs to the family Lythraceae of the order Myrtales, and is also commonly referred to as "Great Flower Crape Myrtle". It is a plant that widely grows in Southeast Asia such as the Philippines, India, Malaysia, southern China, and Australia.

[0038] In the present invention, the flower, leaf, bark, root or seed of banaba can be used, but from the viewpoint of the effect of enhancing telomerase expression, it is preferable to use the leaf. The banaba leaf may include the stem. Since the banaba leaf is easily corroded, it is preferably dried immediately after harvesting to obtain a dried product. The drying of the fresh leaf may be either natural drying or air drying, but preferably it is forcibly dried by air drying. Drying is carried out so that the moisture content becomes 20% by mass or less, preferably 10% by mass or less, whereby corrosion by microorganisms can be prevented and the components in the banaba leaf can be stably retained.

[0039] In the present invention, banana can be processed and used. Examples of banana processed products include chip-like substances, crushed products, squeezed juices, extracts, and their dry powders. Considering the formulatability, the banana used in the present invention is preferably a crushed product, squeezed juice, extract, or their dry powder because it is easy to apply. From the viewpoint of enhancing the telomerase expression effect, an extract or its dry powder is more preferable. The banana processed product may be manufactured by a method commonly known to those skilled in the art or may be a product commercially available.

[0040] The method for obtaining a crushed product of banana is not particularly limited. For example, after washing and drying using sunlight or a dryer, the processed product obtained as it is or cut into an appropriate shape and size can be crushed using a crushing device. As the crushing device, those commonly used can be widely used. For example, a crusher composed of a raw material hopper, crusher, classifier, product holder, etc. can be used.

[0041] The method for obtaining a squeezed juice of banana is not particularly limited. For example, a method of squeezing banana or its shredded pieces, a method of centrifuging or filtering the shredded pieces of banana, etc. can be mentioned. As an example of a specific method for producing a squeezed juice, a method of obtaining a squeezed juice by squeezing with mechanical crushing means such as a mixer or juicer and removing coarse solids by means such as sieving or filtering as necessary can be mentioned. The squeezed juice may be concentrated as necessary or may be processed by freeze-drying, hot air drying, spray drying, etc. to obtain a dry powder.

[0042] The method for obtaining an extract of banana is not particularly limited. For example, an extract obtained by extracting the components contained in banana leaves with a solvent according to a conventional method, its diluted solution or concentrated solution, or their dried products or powders can be mentioned.

[0043] Examples of the solvent used for extraction include water; lower alcohols such as ethanol, methanol, isopropanol, and butanol; lower esters such as ethyl acetate and methyl acetate; acetone; and mixed solvents of these with water. In the present invention, a mixed solvent of an organic solvent and water can be used. Examples of the mixed solvent include mixed solvents of ethanol, methanol, isopropanol, butanol, ethyl acetate, methyl acetate, acetone, and water. Preferably, an acetone / water (2 / 8 to 8 / 2, volume ratio) mixture, an ethanol / water (2 / 8 to 8 / 2, volume ratio) mixture, etc. can be used. The temperature of the extraction solvent can be appropriately set according to the solvent used, within the range from room temperature to below the boiling point.

[0044] Examples of the extraction method include adding a solvent in an amount of 2 to 20 times the mass, preferably 5 to 15 times the mass, more preferably 8 to 10 times the mass, relative to banaba, and performing extraction under any conditions such as standing, shaking, stirring, and reflux at a temperature preferably of 50°C or higher, more preferably in the range of about 50 to 85°C, for several minutes to several tens of hours, preferably about 30 minutes to 2 hours. When performing extraction at a high temperature, since an organic solvent (e.g., ethanol) may evaporate and the extraction efficiency may decrease, it is preferable to perform extraction by heating under reflux. After the extraction operation, it is preferable to perform a solid-liquid separation operation such as filtration or centrifugation to remove insoluble solids. By performing operations such as dilution and concentration as necessary, an extract can be obtained. Further, the same operation can be repeated for the insoluble matter for extraction, and the extract thereof may be combined with the previous extract for use. These extracts may be further purified by a purification method commonly used by those skilled in the art before use.

[0045] The method for obtaining a dried product from the extract is not particularly limited. For example, methods include subjecting the extract or its concentrate to drying treatments commonly used by those skilled in the art, such as spray drying, freeze drying, vacuum drying, fluidized drying, etc. Furthermore, the dried product thus obtained can be powdered and used by using methods known to those skilled in the art. For example, according to the method described in JP-A-2005-263650, a banaba extract can be obtained. According to this method, by performing (a) an extraction step with an aqueous ethanol solution, (b) a treatment step with activated carbon, and (c) a step of concentrating and recovering the precipitate on the leaves of banaba, a banaba extract with a high content of corosolic acid can be obtained.

[0046] [Terminalia] Terminalia is a broad-leaved tree of the genus Terminalia in the Combretaceae family. In the present invention, for example, Terminalia bellirica (belerica), Terminalia catappa, Terminalia tomentosa, Terminalia citrina, Terminalia phellocarpa, Terminalia copelandii, Terminalia brassi, Terminalia ivorensis, Terminalia superba, Terminalia arjuna, Terminalia chebula, etc. can be mentioned. Among these, Terminalia bellirica (belerica) and Terminalia chebula are preferred, and Terminalia belliricca (belerica) with a high telomerase expression enhancing effect is particularly preferred. Also, in the present invention, from the viewpoint of the telomerase expression enhancing effect, it is preferable to use the fruit of Terminalia.

[0047] In the present invention, the harvested Terminalia can be processed and used. Examples of processed Terminalia products include chip-like substances, crushed products, squeezed juices, extracts, and their dry powders. Considering the formulating properties, Terminalia used in the present invention is preferably a crushed product, squeezed juice, extract, or their dry powder because it is easy to apply. From the viewpoint of enhancing the telomerase expression effect, it is more preferably an extract or its dry powder. The processed Terminalia product may be produced by a method commonly known to those skilled in the art or may be a product currently on the market.

[0048] Examples of the extract of Terminalia include an extract obtained by extracting the components contained in Terminalia with a solvent according to a conventional method, a diluted solution or concentrated solution thereof, or a dried product or powder thereof.

[0049] Examples of the solvent used for extraction include water; lower alcohols such as ethanol, methanol, isopropanol, and butanol; lower esters such as ethyl acetate and methyl acetate; acetone; and mixed solvents of these with water. In the present invention, a mixed solvent of an organic solvent and water can be used. Examples of the mixed solvent include mixed solvents of ethanol, methanol, isopropanol, butanol, ethyl acetate, methyl acetate, acetone, and water. Preferably, an acetone / water (2 / 8 to 8 / 2, volume ratio) mixture, an ethanol / water (2 / 8 to 8 / 2, volume ratio) mixture, etc. are mentioned. In the present invention, from the viewpoint of enhancing the telomerase expression effect, it is preferable to use water, ethanol, methanol, or a mixed solvent thereof. The temperature of the extraction solvent can be appropriately set from room temperature to below the boiling point according to the solvent used.

[0050] [Oyster (Crassostrea gigas)] The oyster (Crassostrea gigas) used in the present invention is not particularly limited as long as it belongs to the family Ostreidae, and examples include oysters belonging to the genus Magallana and oysters belonging to the genus Ostrea. In the present invention, from the viewpoint of enhancing the telomerase expression effect, it is preferable to use oyster meat (Crassostrea gigas meat).

[0051] In the present invention, oyster meat can be processed and used. Examples of oyster processed products include crushed products, pulverized products, squeezed juices, extracts (including enzymatically treated products), and dried powders thereof. Considering the formulating properties, the oyster used in the present invention is preferably a pulverized product, squeezed juice, extract, or dried powder thereof because it is easy to apply. From the viewpoint of enhancing the telomerase expression effect, an extract or its dried powder is more preferable. The extraction method using a solvent is the same as that for Terminalia. Examples of the enzymatic treatment include hydrolyzing oyster meat with an enzyme. The type of the hydrolyzing enzyme is not limited as long as it is an enzyme that hydrolyzes food materials (proteins).

[0052] These extraction treatments and enzymatic treatments with solvents can be carried out by heating and / or pressurizing as necessary. In order to obtain more active ingredients, it is preferable to pre-pulverize oyster meat before the extraction treatment and enzymatic treatment. For example, the pulverized oyster meat is extracted with water at about 40 to 90°C for about 1 to 5 hours, the oyster meat residue is removed, and then concentrated. If necessary, spray drying, freeze drying, etc. are carried out to obtain an extraction concentrate (oyster extract) in a liquid or powder form. Also, the pulverized oyster meat is hydrolyzed by allowing an enzyme to act in water at about 40 to 80°C, the enzyme is inactivated, and then unnecessary substances such as oyster meat residue are removed, concentrated, and if necessary, spray drying, freeze drying, etc. are carried out to obtain an extraction concentrate (oyster extract) in a liquid or powder form.

[0053] The extract of the present invention may be the extraction solution itself, but is preferably a concentrated product in a liquid, semi-solid, or solid form obtained by concentrating this. For example, commercially available products containing oyster extract can be used.

[0054] [Dried Tangerine Peel] In the present invention, "Chen Pi" means the pericarp of citrus fruits. Generally, in China, the dried pericarp of ripe mandarin oranges is used as Chen Pi, and in Japan, the dried pericarp of ripe Satsuma oranges is used. However, in the present invention, it is not limited to these, and any citrus fruit pericarp may be used. The citrus fruits may be domestic or foreign. In the present invention, from the viewpoint of the effect of enhancing telomerase expression, for example, Amami, a kind of citrus fruit developed in Ehime Prefecture, is preferable.

[0055] In the present invention, Chen Pi can be processed and used. Examples of processed Chen Pi products include chip-shaped substances, crushed products, squeezed juices, extracts, and dried powders thereof. Considering the formulatability, the Chen Pi used in the present invention is preferably a crushed product, squeezed juice, extract, or dried powder thereof because it is easy to apply. From the viewpoint of the effect of enhancing telomerase expression, an extract or its dried powder is more preferable. The processed Chen Pi product may be manufactured by a method commonly known to those skilled in the art or may be a product commercially available on the market.

[0056] The extract of Chen Pi can be obtained by extraction using an appropriate solvent. The extraction solvent, extraction method, and purification method can be carried out in the same manner as in the case of pine bark.

[0057] [Japanese butterbur] Japanese butterbur is a perennial fern of the family Aspleniaceae or Blechnaceae. The young shoots of Japanese butterbur are called "kogomi" and are one of the wild vegetables. In the present invention, it is preferable to use kogomi from the viewpoint of the effect of enhancing telomerase expression.

[0058] In the present invention, the harvested Terminalia chebula Retz. can be processed and used. Examples of processed Terminalia chebula Retz. products include chip-like substances, crushed products, squeezed juices, extracts, and their dry powders. Considering the pharmaceutical properties, the Terminalia chebula Retz. used in the present invention is preferably a crushed product, squeezed juice, extract, or their dry powder because it is easy to apply. From the perspective of enhancing the telomerase expression effect, it is more preferably an extract or its dry powder. The processed Terminalia chebula Retz. product may be manufactured by methods commonly known to those skilled in the art or may be a product currently on the market. The extraction solvent, extraction method, etc. can be carried out in the same manner as for Terminalia chebula Retz.

[0059] The telomerase expression enhancer of the present invention can be used as an oral preparation or an external preparation applied to the skin or scalp. For example, it can be used as pharmaceuticals (including quasi-drugs), specific health foods, nutritional functional foods, functional display foods, and other so-called health foods whose efficacy has been approved by a predetermined agency, cosmetics, toiletries, etc.

[0060] The telomerase expression enhancer of the present invention is not particularly limited as long as it contains the material of the present invention and can be distinguished from other products as a product in terms of being used for enhancing telomerase expression. For example, any of the main body, packaging, instruction manual, and promotional materials (advertising media) of the product according to the present invention that display the telomerase expression enhancing function and various functions resulting therefrom are included in the scope of the present invention. Note that the telomerase expression enhancer of the present invention is not limited to those in which the material of the present invention is indicated as an active ingredient on the packaging of the product, etc. For example, it may not specify the active ingredient. Also, even if it is a general food, as long as it is manufactured and sold with a suggested use, it is included in the scope of the present invention.

[0061] Specifically, in so-called health foods, examples include those labeled with "enhancing the expression of telomerase," "preventing cell aging," "preventing telomere shortening," "elongating telomeres," "anti-aging," "anti-aging," "promoting hair growth," and the like. The subjects for ingesting the telomerase expression enhancer of the present invention are not particularly limited as long as they are people who need to enhance the expression of telomerase, but preferably include those who aim for anti-aging or anti-aging.

[0062] When the telomerase expression enhancer of the present invention is an oral preparation, examples of its form include tablet form, capsule form, powder form, granule form, liquid form, granule form, rod form, plate form, block form, solid form, round form, paste form, cream form, caplet form, gel form, chewable tablet form, stick form, and the like. Among these, tablet form, capsule form, powder form, granule form, round form, and chewable tablet form are preferred, and tablet form, capsule form, round form, and chewable tablet form are more preferred.

[0063] When the composition of the present invention is in the form of tablets, rounds, or chewable tablets, it is preferable to add one or more of excipients, lubricants, and fluidizing agents to enhance the moldability and improve the storage stability of the obtained tablets, rounds, or chewable tablets. In particular, the use of excipients and lubricants can further enhance the storage stability.

[0064] An excipient is added to improve the handling or molding of the composition or to make taking it convenient. There is no particular limitation on the excipients that can be used in the present invention, and examples include starches such as starch, pregelatinized starch, partially pregelatinized starch, starch degradation products, or their derivatives, crystalline cellulose, sugar alcohols, lactose, brewer's yeast, low-substituted hydroxypropyl cellulose, hydroxypropyl cellulose, refined white sugar, light anhydrous silicic acid, calcium silicate, titanium oxide, precipitated calcium carbonate, and the like. These may be used alone or in combination of two or more.

[0065] A lubricant is used to reduce the friction between the tableting machine pestle and the tablet when compressing the powder for tablets, and to prevent tableting obstacles such as sticking. The lubricant that can be used in the present invention is not particularly limited as long as it is a component capable of achieving the above object. For example, stearic acid or its salts such as stearic acid, calcium stearate, magnesium stearate, sodium stearyl fumarate, sucrose fatty acid ester, talc, polyethylene glycol, vegetable oil, hydrogenated oil, etc. may be mentioned. These may be used alone or in combination of two or more.

[0066] A fluidizing agent is used to improve the fluidity of the mixed powder and granules. There is no particular limitation on the fluidizing agent that can be used in the present invention. For example, silicon dioxide, aluminum silicate, magnesium aluminosilicate, calcium phosphate, magnesium carbonate, magnesium oxide, etc. may be mentioned. These may be used alone or in combination of two or more. In the present invention, commercially available products can be used for the excipient, lubricant, and fluidizing agent.

[0067] When the telomerase expression enhancer of the present invention is an external preparation, its forms include, for example, lotion, emulsion, gel, cream, ointment, powder, granule, etc. Specifically, lotion, makeup cream, emulsion, cream, pack, hair tonic, hair cream, shampoo, hair rinse, treatment, body shampoo, facial cleanser, soap, foundation, powder, lipstick, lip gloss, blush, eye shadow, hair styling agent, hair growth agent, aqueous ointment, oily ointment, eye drops, eye wash, ship, gel, etc. may be mentioned. Also, when applied to the oral cavity, dentifrice, mouthwash, spray, etc. may be mentioned.

[0068] The content of the material of the present invention in the telomerase expression enhancer of the present invention may be appropriately contained within the range where the effect is exhibited.

[0069] Specifically, when the telomerase expression enhancer of the present invention is in the form of tablets, pills, capsules, or chewable tablets, it is preferable that the material of the present invention is contained in an amount of 0.000001 to 60% by mass in terms of dry mass, more preferably 0.00001 to 50% by mass, still more preferably 0.0001 to 40% by mass, and particularly preferably 0.01 to 30% by mass from the viewpoint of enhancing telomerase expression.

[0070] When the telomerase expression enhancer of the present invention is in the form of powder or granules, it is preferable that the material of the present invention is contained in an amount of 0.000001 to 100% by mass in terms of dry mass, more preferably 0.00001 to 90% by mass, still more preferably 0.0002 to 80% by mass, and particularly preferably 0.01 to 70% by mass from the viewpoint of enhancing telomerase expression.

[0071] When the telomerase expression enhancer of the present invention is in the form of a liquid beverage, it is preferable that the material of the present invention is contained in an amount of 0.0000001 to 50% by mass in terms of dry mass, more preferably 0.00001 to 25% by mass, still more preferably 0.0002 to 10% by mass, and particularly preferably 0.001 to 5% by mass from the viewpoint of enhancing telomerase expression.

[0072] When the telomerase expression enhancer of the present invention is an oral preparation, the intake amount is not particularly limited. However, for adults, it is preferable to ingest the material of the present invention in an amount of 10 mg / day or more, more preferably 20 mg / day or more, and still more preferably 30 mg / day or more from the viewpoint of enhancing telomerase expression. The upper limit is, for example, 100000 mg / day, preferably 80000 mg / day, and more preferably 60000 mg / day.

[0073] When the telomerase expression enhancer of the present invention is an oral preparation, it may be appropriately designed so that the daily intake amount is the above-mentioned intake amount, and it may be taken once or divided into multiple times. For example, in the case of tablets, capsules, pills or chewable tablets, the number of intakes per day is 1 to 4 times, and it is sufficient if the total amount can be taken as the above-mentioned intake amount. In the case of beverages, it is sufficient if the above-mentioned intake amount is blended in the daily intake amount. The telomerase expression enhancer of the present invention can be accommodated as a one-day portion in one container or, for example, divided into 2 to 3 multiple containers so that the daily intake amount is the above-mentioned intake amount.

[0074] When the telomerase expression enhancer of the present invention is an external preparation, it is preferable that the material of the present invention is contained in an amount of 0.0000001 to 50% by mass in terms of dry mass, more preferably 0.000001 to 20% by mass, still more preferably 0.00001 to 10% by mass, and particularly preferably 0.0001 to 5% by mass from the viewpoint of enhancing telomerase expression.

[0075] When the telomerase expression enhancer of the present invention is an external preparation, the amount used is not particularly limited, and it can be appropriately selected in consideration of various factors such as the age, weight, and physical constitution of the individual to be used.

[0076] The telomerase expression enhancer of the present invention can be produced by a known method by adding other components other than the material of the present invention, if necessary. When the telomerase expression enhancer of the present invention is an oral preparation, examples of other components include vitamins such as water-soluble vitamins (vitamin B1, B2, B3, B5, B6, B7, B9, B12, B13, B15, B17, vitamin C, vitamin P, choline, inositol, PABA), fat-soluble vitamins (vitamin A, vitamin D, vitamin E, vitamin K); minerals such as magnesium, phosphorus, zinc, iron; sulfur-containing compounds contained in taurine, garlic, etc.; flavonoids or flavonoid-like substances such as hesperidin, quercetin; proteins such as collagen; peptides; amino acids; animal fats and oils; vegetable fats and oils; pulverized products or extracts of animals and plants, etc.

[0077] When the telomerase expression enhancer of the present invention is an external preparation, components that can be added include various medicinal components (active oxygen scavengers, antioxidants, anti-inflammatory agents, cell activators, vitamins, hormones, extracts derived from animals and plants having these actions, etc.), other oils (unsaturated fatty acids such as linoleic acid, linolenic acid, palmitic acid, DHA, EPA and their derivatives, oils extracted from animals and plants such as linseed oil, coconut oil, jojoba oil, olive oil, squalane, squalene, horse oil, rice bran oil, castor oil and their derivatives, etc.), humectants (collagen or its degradation products, collagen-like peptides contained in carrot extract, etc., soy peptides, amino acids, mucopolysaccharides such as hyaluronic acid, amino sugars such as chondroitin, saccharides such as trehalose, seaweeds, water-soluble dietary fibers such as alginic acid, glucomannan, pectin, etc.), surfactants (lecithin, fatty acid esters, amino acid derivatives, etc.), ultraviolet absorbers (zinc oxide, titanium oxide, etc.), ultraviolet absorption promoters, and the like.

Example

[0078] [Preparation of Sample (Test Substance)] As the pine bark, an extract (dry powder) of the bark of Pinus pinaster from the French coast was used. The extract of the pine bark was prepared according to the following method. First, water was added to the pine bark, and extraction was carried out at 95 °C or higher for 1 hour or more. Then, the extract obtained by filtration was purified and dried, and the obtained dry powder was used as the extract of the pine bark.

[0079] As the young barley leaves, a squeezed juice (dry powder) of young barley leaves was used. The squeezed juice of young barley leaves was prepared according to the following method. First, the young barley leaves were made into a paste. Then, after squeezing, the squeezed juice of young barley leaves was dried, and the obtained dry powder was used as the squeezed juice of young barley leaves.

[0080] As the flowers of kudzu, an extract (dry powder) of the flowers of kudzu was used. The extract of the flowers of kudzu was prepared according to the following method. First, water was added to the dried kudzu flowers, and hot water extraction was performed. Next, a solid-liquid separation operation was carried out to remove insoluble solids, and then a kudzu flower extract was obtained. After that, the extract was dried, and the resulting dry powder was used as the extract of kudzu flowers.

[0081] As the black ginger, an extract (dry powder) of the rhizome of black ginger was used. The extract of black ginger was prepared according to the following method. First, water-containing ethanol was added to the rhizome of black ginger for extraction. Next, a solid-liquid separation operation was carried out to remove insoluble solids, and then an extract was obtained. After that, dextrin was added to the extract, and then the extract was dried, and the resulting dry powder was used as the extract of black ginger.

[0082] As the banaba, an extract (dry powder) of banaba leaves was used. The extract of banaba leaves was prepared according to the following method. First, the dried banaba leaves were ground to obtain a ground product. Next, water-containing ethanol was added to the ground product for extraction to obtain an extract. After that, the extract was dried and then ground, and the resulting dry powder was used as the extract of banaba leaves.

[0083] As the Terminalia, an extract (dry powder) of the fruits of Terminalia bellirica ricca was used. The water extract of the fruits of Terminalia bellirica was prepared according to the following method. First, the remaining part obtained by removing seeds from the fruits of Terminalia bellirica was mixed with water to prepare a mixed solution, and hot water extraction was performed. After extraction, the filtrate obtained by filtering the mixed solution was freeze-dried, and the resulting dry powder was used as the extract of the fruits of Terminalia bellirica.

[0084] As the oyster, an extract (dry powder) of the oyster meat was used. The extract of the oyster meat was prepared according to the following method. First, the oyster meat was heated, then cooled, after which an enzyme was added and pressure was applied. Subsequently, it was heat-sterilized and dried. The obtained dry powder was used as an extract of oyster meat.

[0085] As the dried tangerine peel extract (dry powder) with a sweet and neutral taste was used as the tangerine peel. The dried tangerine peel extract was prepared according to the following method. First, water was added to the sweet and neutral tangerine peel, heated, and hot water extraction was carried out. Then, it was filtered to collect the filtrate, and the obtained filtrate was further filtered to collect the filtrate, obtaining an extract of the sweet and neutral tangerine peel. The filtrate was dried, and the obtained dry powder was used as the extract of the sweet and neutral tangerine peel.

[0086] As the young shoots of Kusasotetsu, the dry powder of kogomi (young shoots of Kusasotetsu) was used. The dry powder of kogomi was prepared according to the following method. The young shoots of Kusasotetsu were washed, sterilized, steamed, and then dried. Subsequently, the dried material was pulverized, and the obtained dry powder was used as the dry powder of kogomi.

[0087] Each material was dissolved in DMSO at a concentration of 10 mg / mL and diluted to 10 μg / mL in DMEM medium containing 10% FBS serum to prepare a sample (test substance).

[0088] [Quantitative Reverse transcriptase-PCR (RT-PCR) reaction] 1. Preparation of total RNA For the preparation of total RNA, High Pure RNA Isolation Kit (Roche) was used and carried out according to its product protocol. Also, reagents and instruments used from total RNA preparation to the end of the reverse transcription reaction were RNase Free.

[0089] First, 5.0×10 in a 5 mL cell culture dish 4Cells were seeded at cells / well and cultured at 37°C in DMEM medium containing 10% FBS serum. The next day, various samples (test substances) were added and cultured at 37°C for 48 hours in DMEM medium containing 10% FBS serum. For the comparison control (control), a DMEM medium containing 0.1% DMSO and 10% FBS serum was added. After 48 hours, the medium was completely removed, 200 μL of 1×PBS was added for washing, and then 400 μL of the cell lysate contained in the High Pure RNA Isolation Kit was added. After the cell lysate was evenly spread over the entire dish and dissolved, the entire amount of the cell lysate was collected into a 1.5 mL tube.

[0090] The collected samples were thoroughly suspended for 60 seconds using a vortex mixer and gently spun down. The High Pure filter tube and the collection tube in the Kit were assembled, and the cell lysate solution was added to the filter tube. Centrifugation was performed at room temperature at 10,000×g for 15 seconds, the liquid discharged into the collection tube was discarded, and the filter tube and the collection tube were assembled again. 90 μL of DNase incubation buffer and 10 μL of DNase I were added to a 1.5 mL tube and mixed. This mixed solution was added to the previous filter tube and incubated at room temperature for 15 minutes. After 15 minutes, 500 μL of Wash Buffer I in the Kit was added to the filter tube, and centrifugation was performed at room temperature at 10,000×g for 15 seconds. After centrifugation, the liquid discharged into the collection tube was discarded, and the filter tube and the collection tube were assembled again.

[0091] Add 500 μL of Wash Buffer II to the filter tube and centrifuge at room temperature at 10,000 × g for 15 seconds. After centrifugation, discard the liquid discharged into the collection tube, and reassemble the filter tube and the collection tube. Further, add 200 μL of Wash Buffer II and centrifuge at room temperature at 15,000 × g for 2 minutes to wash the filter. After centrifugation, insert the filter tube into a new 1.5 mL tube, add 100 μL of elution buffer to the center of the filter tube, and let it stand at room temperature for 3 minutes. Then, centrifuge at room temperature at 10,000 × g for 1 minute. The eluate obtained by this operation was used as the RNA solution. The RNA concentration in the solution was calculated based on the absorbance value at 260 nm using a NanoDrop 2000 / 2000c spectrophotometer (Thermo Fisher Scientific) and used in subsequent experiments.

[0092] 2. cDNA Synthesis To 1.0 μg of total RNA extracted from cells, add 5 pmol of Oligo(dT) 20 primer (TOYOBO), and add RNase Free water so that the total volume becomes 13 μL. Perform a heat treatment reaction at 65 °C for 5 minutes using a Thermal Cycler PTC 200 (MJ Research), and immediately transfer it to ice for rapid cooling. During this time, advance the reverse transcriptase reaction program to the 42 °C stage and pause it. To the sample cooled on ice for 5 minutes, add a solution obtained by mixing 4 μL of reverse transcriptase reaction buffer, 2 μL of 10 mM dNTPs (GE Healthcare), and 0.5 μL of reverse transcriptase ReverTra Ace (TOYOBO) per sample, and mix gently. Then, synthesize cDNA by reacting at 42 °C for 20 minutes, 99 °C for 5 minutes, and 4 °C for 5 minutes. This cDNA was used as a template for quantitative PCR.

[0093] 3. Primer Design The target gene whose expression level was measured by RT-PCR was searched on NCBI (http: / / www.ncbi.nlm.nih.gov / gene / ), and based on its sequence, the primer sequences were determined and synthesized. The synthesis of the primers was commissioned to Sigma. The primers for detecting β-actin and hTERT, which are internal controls, are shown in Table 1.

[0094]

Table 1

[0095] 4. Quantitative Reverse transcriptase-PCR (RT-PCR) reaction The prepared cDNA was used as a template. 49 μL of sterilized water, 3.5 μL each of the forward and reverse primers diluted to 10 pmol / mL, 7.0 μL of the template cDNA, and 24.5 μL of the THUNDERBIRD SYBR qPCR Mix (TOYOBO), a master mix for high-efficiency real-time PCR, were placed in a 0.2 mL PCR tube and suspended well. Then, 25 μL each was added to 3 wells in a 96-well plate, and quantitative real-time PCR was performed using a Thermal Cycler Dice Real Time System (TAKARA BIO). The PCR reaction was carried out with a denaturation reaction at 95°C for 5 seconds, an annealing reaction at 60°C for 10 seconds, and an extension reaction at 72°C for 20 seconds, and this was repeated for 45 cycles (3 steps) and detected by FAM. β-actin was used as the primer for the calibration curve. Also, the relative gene expression level was obtained by dividing the measured value by the expression level value of β-actin.

[0096] Table 2 and Figure 1 show the relative expression levels of the human telomerase reverse transcriptase (hTERT) gene. Note that human telomerase reverse transcriptase (hTERT) is an enzyme contained in telomerase. As shown in Figure 1, the expression of the human telomerase reverse transcriptase (hTERT) gene was enhanced by adding various materials of the present invention.

[0097]

Table 2

[0098] Examples of various aspects of the present invention are given below, but the technical scope of the present invention is not limited thereto.

[0099] <Production Examples 1 to 8> Using the formulation in Table 3 below, at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, Terminalia, oysters, tangerine peel, and Psoralea corylifolia was blended, mixed uniformly, and then tablets were produced by molding using a tableting device (250 mg per tablet). The obtained food composition can be expected to have a high telomerase expression enhancing effect.

[0100]

Table 3

[0101] <Production Examples 9 to 16> Using the formulation in Table 4 below, at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, Terminalia, oysters, tangerine peel, and Psoralea corylifolia was blended, mixed uniformly, and filled into a gelatin-containing coating to produce hard capsules (350 mg per capsule). The obtained food composition can be expected to have a high telomerase expression enhancing effect.

[0102]

Table 4

[0103] <Production Examples 17 to 24> With the formulation shown in Table 5 below, at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, Terminalia, oysters, tangerine peel, and Psoralea corylifolia was formulated, mixed uniformly, and encapsulated with a film containing gelatin and glycerin to produce soft capsules (300 mg per capsule). The obtained food composition is expected to have a high telomerase expression enhancing effect.

[0104]

Table 5

[0105] <Production Examples 25 to 32> With the formulation shown in Table 6 below, at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, Terminalia, oysters, tangerine peel, and Psoralea corylifolia was formulated and mixed uniformly to produce a powdered beverage. For example, by mixing 3 g of the obtained food composition with 150 mL of water and drinking it, a high telomerase expression enhancing effect can be expected.

[0106]

Table 6

[0107] <Production Examples 33 to 40> With the formulation shown in Table 7 below, at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, Terminalia, oysters, tangerine peel, and Psoralea corylifolia was respectively formulated, put into a fluidized bed granulator, mixed with an air stream for several minutes, and granulated by spraying 60 L of water at 2000 mL per minute. Subsequently, the obtained granulated product was sieved through a 30-mesh sieve to produce granules. The obtained food composition is expected to have a high telomerase expression enhancing effect.

[0108]

Table 7

[0109] <Production Examples 41 to 56> At the formulations shown in Tables 8 to 11 below, at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, Terminalia, oysters, tangerine peel, and Kusasotetsu was respectively formulated to produce a lotion. The obtained external preparation can be expected to have a high telomerase expression enhancing effect.

[0110]

Table 8

[0111]

Table 9

[0112]

Table 10

[0113]

Table 11

[0114] <Production Examples 57 to 64> At the formulations shown in Tables 12 and 13 below, at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, Terminalia, oysters, tangerine peel, and Kusasotetsu was respectively formulated to produce a cream. The obtained external preparation can be expected to have a high telomerase expression enhancing effect.

[0115]

Table 12

[0116]

Table 13

[0117] <Production Examples 65 to 72> Using the formulations shown in Table 14 and Table 15 below, at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, Terminalia, oysters, tangerine peel, and Kusasotetsu was blended respectively to produce an emulsion. The obtained external preparation is expected to have a high telomerase expression enhancing effect.

[0118]

Table 14

[0119]

Table 15

[0120] <Production Examples 73 to 80> Using the formulations shown in Table 16 and Table 17 below, at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, Terminalia, oysters, tangerine peel, and Kusasotetsu was blended respectively to produce a gel agent. The obtained external preparation is expected to have a high telomerase expression enhancing effect.

[0121]

Table 16

[0122]

Table 17

[0123] <Production Examples 81 to 88> Using the formulations shown in Table 18 and Table 19 below, at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, Terminalia, oysters, tangerine peel, and Kusasotetsu was blended respectively to produce an aerosol agent. The obtained external preparation is expected to have a high telomerase expression enhancing effect.

[0124]

Table 18

[0125]

Table 19

Industrial Applicability

[0126] Since the telomerase expression enhancer of the present invention can be used as a health food or the like, it is industrially useful.

Claims

1. A telomerase expression enhancer characterized by containing young barley leaves.

2. The telomerase expression enhancer according to Claim 1, which is an oral preparation.

3. The telomerase expression enhancer according to Claim 1, which is a topical preparation.

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