JP7442079B2 - Telomerase expression enhancer - Google Patents

Description

Article 30, Paragraph 2 of the Patent Act applies Publication name: Proceedings of the 65th Annual Conference of the Japanese Society of Food Science and Technology, page 118 Publisher: Secretariat of the 65th Annual Conference of the Japan Society of Food Science and Technology Publication date: 2018 August 22nd Meeting name: 65th Annual Meeting of the Japan Society for Food Science and Technology Event date: August 24th, 2018 Publication start date: September 10th, 2018 Publication address: http://www. toyoshinyaku. co. jp/%E3%80%8E%E3%83%95%E3%83%A9%E3%83%90%E3%83%B3%E3%82%B8%E3%82%A7%E3%83%8E %E3%83%BC%E3%83%AB%E3%80%8F%E3%81%8C%E3%83%86%E3%83%AD%E3%83%A1%E3%83%A9%E3 %83%BC%E3%82%BC%E3%82%92%E6%B4%BB%E6%80%A7/ Publication start date: September 19, 2018 Publication address: https://www. kenko-media. com/food devlp/archives/2944 Posting start date: September 27, 2018 Posting address: https://www. bci. co. jp/nichiryu/article/4420

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

In recent years, against the backdrop of increasing health consciousness and safety consciousness, products using natural materials are preferred in the food and cosmetics fields, and there is a desire to make better use of the useful functions of such natural materials. There is.

For example, barley grass is commonly used as a material for health foods and green juices, and its functionality, such as its bowel movement improving effect and antihypertensive effect, has been widely studied (Patent Documents 1 to 3).

Japanese Patent Application Publication No. 2006-151951 Japanese Patent Application Publication No. 2001-314170 Patent No. 4183886

An object of the present invention is to provide a telomerase expression enhancer containing a natural material and having an excellent telomerase expression enhancement effect.

As a result of investigating the functions of various natural materials, the present inventors discovered that a specific natural material has an excellent effect of enhancing telomerase expression, and completed the present invention.

That is, the present invention is as follows.
[1] A telomerase expression enhancer containing at least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, terminaria, oysters, bark, and cycad.
[2] The telomerase expression enhancer according to [1], which is an oral preparation.
[3] The telomerase expression enhancer according to [1], which is an external preparation.

The telomerase expression enhancer of the present invention exhibits an excellent telomerase expression enhancement effect.

FIG. 2 is a diagram showing the relative expression level of the human telomere reverse transcriptase (hTERT) gene when the telomerase expression enhancer of the present invention is applied.

The telomerase expression enhancer of the present invention is characterized in that it contains at least one material selected from pine bark, barley grass, kudzu flower, black ginger, banaba, terminaria, oyster, chinpi, and cycad cycad. . The telomerase expression enhancer of the present invention may contain one type, two types, or three or more types of materials. When containing two or more types, it is preferable to combine those that are highly effective individually.

The telomerase expression enhancer of the present invention enhances the expression of telomerase (telomere synthase). Therefore, cell telomeres can be elongated and cell aging can be prevented.

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 explained below.

[Pine bark]
Examples of the raw material pine for the pine bark used in the present invention include French maritime pine (Pinus Martima), larch, Japanese black pine, Japanese red pine, Japanese pine, Japanese pine, Korean pine, Japanese pine, Japanese pine, Japanese pine, Japanese pine, white pine, etc. Among these, it is preferable to use French maritime pine, which has a high effect of enhancing telomerase expression.

In the present invention, pine bark can be processed and used, and examples of processed pine bark products include chips, pulverized products, squeezed products, extracts, and dry powders thereof. The pine bark used in the present invention is preferably a crushed product, a squeezed product, an extract, or a dry powder thereof, since it is easy to apply in consideration of formulation properties, and from the viewpoint of the effect of enhancing telomerase expression, More preferably, it is an extract or a dry powder thereof. The processed pine bark product may be one manufactured by a method commonly known by those skilled in the art, or one that is available on the market. For example, pine bark extract manufactured by Toyo Shinyaku Co., Ltd. can be used.

Examples of the extraction solvent used when obtaining the pine bark extract include water, organic solvents, and water-containing organic solvents (hydrous alcohols such as hydrous ethanol). Examples of organic solvents include methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, butane, acetone, hexane, cyclohexane, propylene glycol, aqueous ethanol, aqueous propylene glycol, ethyl methyl ketone, and glycerin. , methyl acetate, ethyl acetate, diethyl ether, dichloromethane, edible fats and oils, 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.

There are no particular limitations on the method for obtaining the pine bark extract, but examples include heating extraction method, supercritical fluid extraction method, liquid carbon dioxide batch method, liquid carbon dioxide reflux method, supercritical carbon dioxide reflux method, etc. It will be done. Further, a plurality of extraction methods may be combined. By combining multiple extraction methods, it is possible to obtain pine bark extracts with various compositions.

The supercritical fluid extraction method is a method of extraction using a supercritical fluid, which is a fluid that exceeds the critical point (critical temperature, critical pressure) of gas and liquid of a substance. As the supercritical fluid, carbon dioxide, ethylene, propane, nitrous oxide (laughing gas), etc. are used, and carbon dioxide is preferably used.

The supercritical fluid extraction method includes an extraction step in which a target component is extracted using a supercritical fluid, and a separation step in which the target component and the supercritical fluid are separated. In the separation step, any of extraction separation using a pressure change, extraction separation using a temperature change, and extraction separation using an adsorbent or absorbent may be performed.

Alternatively, supercritical fluid extraction may be performed using an entrainer addition method. In this method, for example, ethanol, propanol, n-hexane, acetone, toluene, other aliphatic lower alcohols, aliphatic hydrocarbons, aromatic hydrocarbons, ketones are added to the extraction fluid at 2 to 20 W/V. By adding about 1.9% and performing supercritical fluid extraction using this fluid, the solubility of target extracts such as OPC (oligomeric proanthocyanidin) and catechins in the extraction solvent can be dramatically increased. This is a method for increasing the selectivity of separation or for obtaining an efficient pine bark extract.

Supercritical fluid extraction has the advantage that it can be operated at relatively low temperatures, so it can be applied to substances that change or decompose at high temperatures. It also has the advantage that no extraction fluid remains, and because it allows the circulation of the solvent, it can be used for desolvation. This has the advantage that steps can be omitted and the process becomes simple.

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

The pine bark extract used in the present invention contains proanthocyanidins as one of the main components. Proanthocyanidins are a group of compounds consisting of condensation polymers having a degree of polymerization of 2 or more and having flavan-3-ol and/or flavan-3,4-diol as constituent units.

The pine bark extract used in the present invention preferably contains a condensation polymer having a degree of polymerization of 2 or more as proanthocyanidin. In particular, proanthocyanidins containing a large amount of condensation polymers with a low degree of polymerization are preferred. Examples of condensation polymers with a low degree of polymerization include condensation polymers with a degree of polymerization of 2 to 30 (dimers to 30mers), and condensation polymers with a degree of polymerization of 2 to 10 (dimers to 10mers). Preferably, condensation polymers (dimers to tetramers) having a degree of polymerization of 2 to 4 are more preferable. In this specification, a polymer having 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.

[Barley grass]
Barley (Hordeum vulgare L.) is said to be native to Central Asia, is an annual or perennial herb belonging to the Poaceae family, and is broadly classified into two-rowed barley, six-rowed barley, etc., depending on the ear shape. The barley grass used in the present invention is not particularly limited as long as it is normally available, and any young barley leaves such as two-rowed barley or six-rowed barley may be used. Also, any variety may be used. The barley grass only needs to contain young barley leaves, and may contain other parts such as stems.

The form of young barley leaves is not particularly limited, and for example, young leaves may be harvested as they are, or young leaves may be further processed after being harvested. Processing treatments for barley grass include, for example, dry powder treatment, shred treatment and its dry powder treatment, juice extraction treatment and its dry powder treatment, extract extraction treatment and its dry powder treatment, etc., and these are known in the art. The adopted processing method can be applied. That is, examples of processed products include chips, pulverized products, juice, and extracts of young barley leaves, and examples of pulverized products include powders, granules, and the like. Examples of methods for producing a pulverized product include drying barley leaves and/or stems, coarsely pulverizing them, heating them at 110°C or higher, and further pulverizing them into fine particles (see JP-A No. 2003-033151); , a method in which barley leaves and/or stems are blanched, dried, and then crushed (see Japanese Patent Application Laid-Open No. 2002-065204).

The juice and extract of young barley leaves may be in liquid form, but may also be used in the form of paste or dry powder. The method for squeezing young barley leaves is not particularly limited, and any method commonly used by those skilled in the art can be applied. Extraction/separation methods and synthesis methods from barley grass are not particularly limited and can be appropriately selected depending on the purpose. For example, extraction solvents commonly used by those skilled in the art such as ethanol, water, and aqueous ethanol Examples include a method in which the mixture is added and extracted by heating if necessary.

The barley grass used in the present invention is preferably a ground product, a squeezed product, an extract, or a dry powder thereof, since it is easy to apply in consideration of formulation properties, and from the viewpoint of the effect of enhancing telomerase expression, It is more preferable to use a crushed product, a squeezed product, or a dry powder thereof. The processed barley grass product may be one produced by a method commonly known by those skilled in the art, or one that is available on the market.

[Kuzu no Hana]
Kudzu is a climbing perennial plant belonging to the Fabaceae family and the genus Kudzu. Kudzu flowers may be collected at any stage from bud to fully open flower, or a mixture of kudzu flowers collected at each stage may be used. The type of kudzu is not particularly limited, but examples include Pueraria thomsonii, Pueraria lobata, Pueraria thunbergiana, and the like.

In the present invention, harvested kudzu flowers can be processed and used, and examples of processed kudzu flowers include chips, crushed products, squeezed products, extracts, and dry powders thereof. I can do it. Kudzu flowers used in the present invention are preferably ground products, squeezed products, extracts, or dry powders thereof, since they are easy to apply in consideration of formulation properties, and from the viewpoint of the effect of enhancing telomerase expression. , an extract or a dry powder thereof. The kudzu flower processed product may be produced by a method commonly known by those skilled in the art, or may be commercially available.

Kudzu flower extract can be obtained by extraction using a suitable solvent, such as water; lower alcohols such as ethanol, methanol, isopropanol, butanol; ethyl acetate, methyl acetate, etc. Examples include lower esters, acetone, and mixed solvents of these and water. The temperature of the extraction solvent can be appropriately set from room temperature to below the boiling point depending on 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 a mixed solvent of ethanol, methanol, isopropanol, butanol, ethyl acetate, methyl acetate, acetone, and water. I can do it. The temperature of the extraction solvent can be appropriately set from room temperature to below the boiling point depending on the solvent used. In the present invention, hot water extracts are preferred from the viewpoint of enhancing telomerase expression.

[Black ginger]
The black ginger (Kaempferia parviflora) used in the present invention is not particularly limited as long as it is a plant of the Zingiberaceae family, Kaempferia parviflora, which is known to grow naturally in Southeast Asia. Parts such as roots, stems, leaves, flowers, and branches can be used, but roots and stems are preferred.

In addition to the unprocessed black ginger that is collected, the black ginger may be a processed product that is obtained by subjecting it to a predetermined treatment after being collected. Examples of processed products include chips, pulverized products, squeezed products, extracts, and dry powders thereof. The black ginger used in the present invention is preferably a ground product, a squeezed product, an extract, or a dry powder thereof, since it is easy to apply in consideration of formulation properties, and from the viewpoint of the effect of enhancing telomerase expression, More preferably, it is an extract or a dry powder thereof. In addition, from the viewpoint of ease of processing, storage, transportation, etc., and versatility in usage, a dry powder is preferable. The processed black ginger product may be one manufactured by a method commonly known by those skilled in the art, or one that is available on the market.

The extract of black ginger is not particularly limited as long as the components of black ginger are extracted, but for example, the extract obtained by extracting the components contained in black ginger with a solvent according to a conventional method, or a diluted solution thereof. and concentrated liquids, their dried products, and their powders. For example, components contained in black ginger include, but are not limited to, polymethoxyflavonoids such as 5,7-dimethoxyflavone.

Solvents used to obtain the black ginger extract include, for example, water such as room temperature water, hot water, and hot water; lower alcohols such as ethanol, methanol, isopropanol, and butanol; lower esters such as ethyl acetate and methyl acetate; Examples include acetone; a mixed solvent of these and water; and the like. Examples of the mixed solvent include acetone/water (2/8 to 8/2, volume ratio) mixture, ethanol/water (2/8 to 8/2, volume ratio) mixture, and the like.

There are no particular limitations on the method for obtaining the black ginger extract, but for example, add a solvent 2 to 20 times the mass of black ginger, and add it to the reflux temperature of the solvent for several minutes to several tens of hours at a temperature of 0°C to the reflux temperature of the solvent. Examples include methods of performing extraction under arbitrary conditions such as standing still, shaking, stirring, and refluxing. After the extraction operation, it is preferable to perform a solid-liquid separation operation such as filtration or centrifugation to remove insoluble solids. An extract can be obtained by performing operations such as dilution and concentration as necessary. Furthermore, the same operation may be repeated to extract insoluble matter, and the resulting extract may be used in combination with the previous extract. These extracts may be further purified and used by purification methods commonly used by those skilled in the art.

The method for obtaining a dried product from an extract is not particularly limited, and examples include methods of subjecting the extract or its concentrate to drying treatments commonly used by those skilled in the art, such as spray drying, freeze drying, reduced pressure drying, and fluidized drying. It will be done. Furthermore, the dried product thus obtained can be used after being powdered using a method known to those skilled in the art.

[Banaba]
Banaba (Lagerstroemia speciosa, Linn. or Pers.) is a plant that belongs to the family Myrtaceae and the family Lythraceae. It is also commonly known as "Lagerstroemia speciosa," and it grows widely in the Philippines, India, Malaysia, southern China, and other Southeast Asian countries, as well as Australia. It is a plant that has

In the present invention, Banaba flowers, leaves, bark, roots, or seeds can be used, but from the viewpoint of the effect of enhancing telomerase expression, it is preferable to use leaves. Banaba leaves may include stems. Since banaba leaves are prone to rot, it is preferable to dry them immediately after harvesting to make a dried product. The fresh leaves may be dried naturally or in the air, but it is preferable to forcefully dry them by air-drying. Drying is performed so that the moisture content is 20% by mass or less, preferably 10% by mass or less, thereby preventing corrosion of microorganisms and stably retaining the components in banaba leaves.

In the present invention, banaba can be processed and used, and processed banaba products include, for example, chips, pulverized products, squeezed products, extracts, and dry powders thereof. Banaba used in the present invention is preferably a crushed product, a squeezed product, an extract, or a dry powder thereof, since it is easy to apply in consideration of formulation properties. It is more preferable to use a substance or a dry powder thereof. Banaba processed products may be produced by methods commonly known by those skilled in the art, or may be commercially available.

There are no particular limitations on the method for obtaining the crushed banaba, for example, after washing and drying in the sun or in a dryer, the processed product may be obtained as is or by cutting into appropriate shapes and sizes. It can be obtained by grinding using a grinding device. As the crushing device, a wide variety of commonly used devices can be used, and for example, a crusher consisting of a raw material hopper, a crusher, a classifier, a product holder, etc. can be used.

There are no particular limitations on the method for obtaining the juice of banaba, and examples thereof include a method of squeezing banaba or its fragments, a method of centrifuging or filtering the fragments of banaba, and the like. As a specific example of a method for producing squeezed juice, juice is squeezed using mechanical crushing means such as a mixer or juicer, and if necessary, crude solids are removed by means such as sieving or filtration to obtain squeezed juice. There are several ways to obtain this. The squeezed product may be concentrated as necessary, or it may be processed into a dry powder by freeze drying, hot air drying, spray drying, or the like.

There are no particular limitations on the method for obtaining the banaba extract, and for example, the extract obtained by extracting the components contained in banaba leaves with a solvent in accordance with a conventional method, its diluted or concentrated solution, or its drying. Examples include substances and their powders.

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 a mixed solvent of these and water. In the present invention, a mixed solvent of an organic solvent and water can be used, such as a mixed solvent of ethanol, methanol, isopropanol, butanol, ethyl acetate, methyl acetate, acetone, and water. Preferred examples include acetone/water (2/8 to 8/2, volume ratio) mixture and ethanol/water (2/8 to 8/2, volume ratio) mixture. The temperature of the extraction solvent can be appropriately set from room temperature to below the boiling point depending on the solvent used.

The extraction method includes, for example, adding a solvent 2 to 20 times, preferably 5 to 15 times, more preferably 8 to 10 times the mass of banaba, preferably at 50°C or higher, more preferably about 50 to 85°C. Examples include a method in which extraction is carried out under arbitrary conditions such as standing, shaking, stirring, and refluxing at a temperature ranging from several minutes to several tens of hours, preferably about 30 minutes to 2 hours. When extracting at a high temperature, the organic solvent (for example, ethanol) may evaporate and the extraction efficiency may decrease, so 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. An extract can be obtained by performing operations such as dilution and concentration as necessary. Furthermore, the same operation may be repeated to extract insoluble matter, and the resulting extract may be used in combination with the previous extract. These extracts may be further purified and used by purification methods commonly used by those skilled in the art.

There are no particular limitations on the method of obtaining a dried product from the extract, and examples include methods of 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, and fluidized drying. can be mentioned. Furthermore, the dried product thus obtained can be used after being powdered using a method known to those skilled in the art. For example, banaba extract can be obtained according to the method described in JP-A-2005-263650. According to this method, Banaba leaves have a high content of corosolic acid 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 collecting the precipitate. Banaba extract can be obtained.

[Terminaria]
Terminalia is a broad-leaved tree belonging to the genus Momotamana in the family Shicunaceae. In the present invention, for example, Terminalia bellirica (belerica), Terminalia catappa, Terminalia tomentosa, Terminalia citrina, Terminalia phellocarpa, Terminalia Inalia coplandii, Terminalia brassi, Terminalia ivorensis, Terminalia superba, Terminalia arjuna, Terminalia chebula, etc. Among these, Terminaria bellirica (belerica) and Terminalia chebula are preferred, and Terminaria bellirica (belerica), which has a high effect of enhancing telomerase expression, is particularly preferred. Furthermore, in the present invention, it is preferable to use the fruit of Terminalia from the viewpoint of the effect of enhancing telomerase expression.

In the present invention, the harvested Terminaria can be processed and used, and examples of processed Terminaria include chips, pulverized products, squeezed products, extracts, and dry powders thereof. The terminalia used in the present invention is preferably a pulverized product, a squeezed product, an extract, or a dry powder thereof, since it is easy to apply in consideration of formulation properties. It is more preferable to use a substance or a dry powder thereof. The terminalia processed product may be manufactured by a method commonly known by those skilled in the art, or may be commercially available.

Extracts of Terminalia include, for example, extracts obtained by extracting the components contained in Terminaria with a solvent in accordance with a conventional method, diluted or concentrated liquids thereof, dried products thereof, powders thereof, etc. .

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 a mixed solvent of these and water. In the present invention, a mixed solvent of an organic solvent and water can be used, such as a mixed solvent of ethanol, methanol, isopropanol, butanol, ethyl acetate, methyl acetate, acetone, and water. Preferred examples include acetone/water (2/8 to 8/2, volume ratio) mixture and ethanol/water (2/8 to 8/2, volume ratio) mixture. In the present invention, water, ethanol, methanol, or a mixed solvent thereof is preferably used from the viewpoint of the effect of enhancing telomerase expression. The temperature of the extraction solvent can be appropriately set from room temperature to below the boiling point depending on the solvent used.

[Oysters]
The oysters used in the present invention are not particularly limited as long as they belong to the oyster family, and examples thereof include oysters belonging to the genus Perilla and oysters belonging to the genus oyster. In the present invention, it is preferable to use oyster meat from the viewpoint of the effect of enhancing telomerase expression.

In the present invention, oyster meat can be processed and used, and examples of processed oyster products include crushed products, crushed products, squeezed products, extracts (including enzyme-treated products), and dry powders thereof. be able to. The oysters used in the present invention are preferably ground products, squeezed products, extracts, or dry powders thereof because they are easy to apply in consideration of formulation properties. It is more preferable to use a substance or a dry powder thereof. Note that the extraction method using a solvent is the same as for Terminalia. Further, as an example of the enzyme treatment, one in which oyster meat is hydrolyzed with an enzyme can be exemplified. The hydrolytic enzyme may be of any type as long as it hydrolyzes food materials (proteins).

These solvent extraction treatments and enzyme treatments can be carried out by heating and/or pressurizing, if necessary. Further, in order to obtain more active ingredients, it is preferable to crush the oyster meat in advance before extraction treatment and enzyme treatment. For example, pulverized oyster meat is extracted with water at a temperature of 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 performed to obtain liquid or powdered oyster meat. It is possible to obtain an extract concentrate (oyster extract) such as oyster extract. In addition, the crushed oyster meat is hydrolyzed with enzymes in water at about 40 to 80 degrees Celsius, and after the enzymes are inactivated, unnecessary substances such as oyster meat residue are removed, concentrated, and processed as needed. Spray drying, freeze drying, etc. can be performed to obtain an extraction concentrate (oyster extract) in liquid or powder form.

Although the extract of the present invention may be an extract itself, it is preferably a concentrated liquid, semi-solid, or solid concentrate. For example, a commercially available product containing oyster extract can be used.

[Chen Pi]
In the present invention, peel refers to the peel of a citrus fruit. In China, dried peels of ripe mandarin oranges are generally used as peels, and in Japan, dried peels of ripe Satsuma mandarin oranges are used. The material is not limited to , but any citrus peel may be used. Citrus fruits may be, for example, domestically produced or foreign. In the present invention, from the viewpoint of the effect of enhancing telomerase expression, for example, Kanpei, which is a type of citrus fruit developed in Ehime Prefecture, is preferable.

In the present invention, processed skin peel can be used, and processed skin peel products include, for example, chips, pulverized products, squeezed products, extracts, and dry powders thereof. Considering the formulation properties, it is preferable that the chinenpi used in the present invention is a crushed product, a squeezed product, an extract, or a dry powder thereof, since it is easy to apply. It is more preferable to use a substance or a dry powder thereof. The peel processed product may be produced by a method commonly known by those skilled in the art, or may be commercially available.

The extract of Chinpi can be obtained by extraction using a suitable solvent. The extraction solvent, extraction method, and purification method can be carried out in the same manner as in the case of pine bark.

[Cycad cycad]
A cycad is a perennial fern of the family Iwadendae or the family Mesidae. The young shoots of the cycad are called ``kogomi'' and are a type of wild vegetable. In the present invention, it is preferable to use Kogomi from the viewpoint of the effect of enhancing telomerase expression.

In the present invention, harvested cycads can be processed and used, and examples of processed cycads include chips, pulverized products, squeezed products, extracts, and dry powders thereof. Considering the formulation properties, it is preferable that the cycad used in the present invention is a crushed product, a squeezed product, an extract, or a dry powder thereof, since it is easy to apply. It is more preferable to use a substance or a dry powder thereof. The processed cycad can be manufactured by a method commonly known by those skilled in the art, or can be commercially available. Note that the extraction solvent, extraction method, etc. can be performed in the same manner as for Terminaria.

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, pharmaceuticals (including quasi-drugs), so-called health foods such as foods for specified health uses, foods with nutritional function claims, functional foods whose efficacy has been approved by designated organizations such as foods with functional claims, cosmetics, etc. , can be used as toiletries, etc.

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 in terms of being used for enhancing telomerase expression. For example, the scope of the present invention includes products that display on the main body, packaging, instruction manual, or advertising material (advertisement media) that the product has a telomerase expression enhancement function and various functions resulting from this. include. 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 product packaging or the like. For example, the active ingredient may not be specified. Moreover, even if it is a general food product, it is included in the scope of the present invention that is manufactured and sold with indication of its use.

Specifically, so-called health foods include ``enhancing the expression of telomerase'', ``preventing cell aging'', ``preventing telomere shortening'', ``lengthening telomeres'', ``anti-aging'' and ``anti-aging''. , "hair growth", etc. can be exemplified. Subjects who take the telomerase expression enhancer of the present invention are not particularly limited as long as they need to enhance telomerase expression, but preferred examples include people who are aiming for anti-aging or anti-aging. .

When the telomerase expression enhancer of the present invention is an oral preparation, its form includes, for example, tablet, capsule, powder, granule, liquid, granule, rod, plate, block, solid, round, Examples include paste, cream, caplet, gel, chewable tablets, and sticks. Among these, tablet, capsule, powder, granule, round, and chewable tablet forms are preferred, and tablet, capsule, round, and chewable tablet forms are more preferred.

When the composition of the present invention is made into a tablet, a round shape, or a chewable tablet, one or more of excipients, lubricants, and fluidizing agents can be added to improve moldability and improve the shape of the composition. It is preferred because it improves the storage stability of tablets, pills, or chewable tablets. In particular, storage stability can be further enhanced by using excipients and lubricants.

Excipients are added to improve the handling or shaping of the composition or to make it more convenient to administer. There are no particular limitations on excipients that can be used in the present invention, and examples include starch, pregelatinized starch, partially pregelatinized starch, starch derivatives such as starch decomposition products, crystalline cellulose, sugar alcohols, lactose, brewer's yeast, Examples include 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.

A lubricant is used to reduce friction between the tablet punch and die of a tablet when compressing powder for tablets, and to prevent tableting problems such as sticking. The lubricant that can be used in the present invention is not particularly limited as long as it is a component that can achieve the above purpose, and examples include stearic acid or its salts such as stearic acid, calcium stearate, and magnesium stearate, and fumaric acid. Examples include stearyl sodium, sucrose fatty acid ester, talc, polyethylene glycol, vegetable oil, and hydrogenated oil. These may be used alone or in combination of two or more.

A fluidizing agent is used to improve the fluidity of mixed powder or granules. The fluidizing agent that can be used in the present invention is not particularly limited, and examples include silicon dioxide, aluminum silicate, magnesium aluminate silicate, calcium phosphate, magnesium carbonate, and magnesium oxide. These may be used alone or in combination of two or more. In the present invention, commercially available excipients, lubricants, and fluidizing agents can all be used.

When the telomerase expression enhancer of the present invention is an external preparation, its form includes, for example, lotion, emulsion, gel, cream, ointment, powder, and granule. Specifically, lotions, cosmetic creams, emulsions, creams, packs, hair tonics, hair creams, shampoos, hair rinses, treatments, body shampoos, facial cleansers, soaps, foundations, whitening powders, lipsticks, lip glosses, blushers, and eye shadows. , hair conditioners, hair growth agents, water-based ointments, oil-based ointments, eye drops, eye washes, eye drops, gels, etc. When applied to the oral cavity, examples include toothpaste, mouthwash, and spray.

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 can be achieved.

Specifically, when the telomerase expression enhancer of the present invention is in the form of a tablet, round, capsule, or chewable tablet, the material of the present invention is contained in an amount of 0.000001 to 60% by mass of the total in terms of dry mass. The content is preferably 0.00001 to 50% by mass, more preferably 0.0001 to 40% by mass, and from the viewpoint of enhancing telomerase expression, 0.01 to 30% by mass. It is particularly preferable that the content is % by mass.

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, and preferably 0.00001 to 90% by mass in terms of dry mass. It is more preferable that the content is 0.0002 to 80% by mass, and even more preferably 0.01 to 70% by mass from the viewpoint of enhancing telomerase expression.

When the telomerase expression enhancer of the present invention is 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, and preferably 0.00001 to 25% by mass in terms of dry mass. It is more preferable that the amount is 0.0002 to 10% by mass, and even more preferably 0.001 to 5% by mass from the viewpoint of enhancing telomerase expression.

When the telomerase expression enhancer of the present invention is an oral preparation, there is no particular restriction on the intake amount, but it is preferable that the intake amount of the material of the present invention is 10 mg/day or more per adult. It is more preferable to take in an amount of 20 mg/day or more, and even more preferably to take in an amount of 30 mg/day or more in terms of enhancing telomerase expression. The upper limit is, for example, 100,000 mg/day, preferably 80,000 mg/day, and more preferably 60,000 mg/day.

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 at once or divided into multiple doses. Good too. For example, in the case of tablets, capsules, pills, or chewable tablets, it is sufficient that the intake is taken 1 to 4 times per day, and the above-mentioned intake can be taken as a total amount, and in the case of drinks, the intake should be added to the daily intake. It is sufficient if the amount is mixed. The telomerase expression enhancer of the present invention can be stored in one container or divided into a plurality of containers, for example, 2 to 3, so that the daily intake amount is the above-mentioned intake amount. .

When the telomerase expression enhancer of the present invention is an external preparation, the material of the present invention is preferably contained in an amount of 0.0000001 to 50% by mass, and preferably 0.000001 to 20% by mass, based on dry mass. It is more preferable that the content is 0.00001 to 10% by mass, and even more preferably 0.0001 to 5% by mass is contained from the viewpoint of enhancing telomerase expression.

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

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, other ingredients include, for example, water-soluble vitamins (vitamins 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; Contained in taurine, garlic, etc. flavanoids or flavonoids such as hesperidin and quercetin; proteins such as collagen; peptides; amino acids; animal fats and oils; vegetable fats and oils; ground products or extracts of animals and plants.

When the telomerase expression enhancer of the present invention is an external preparation, the ingredients that can be added include various medicinal ingredients (active oxygen scavengers, antioxidants, anti-inflammatory agents, cell activators, vitamins, hormones, etc.). (e.g., extracts derived from plants and animals having Oils extracted from animals and plants such as horse oil, rice bran oil, and castor oil and their derivatives), moisturizers (such as collagen or its decomposition products, collagen-like peptides contained in carrot extract, soybean peptides, amino acids, hyaluronic acid, etc.) Mucopolysaccharides, amino sugars such as chondroitin, sugars such as trehalose, seaweed, alginic acid, glucomannan, water-soluble dietary fibers such as pectin, etc.), surfactants (lecithin, fatty acid esters, amino acid derivatives, etc.), ultraviolet absorbers (zinc oxide, titanium oxide, etc.), ultraviolet absorption promoters, etc.

[Preparation of sample (test substance)]
As the pine bark, an extract (dry powder) of French maritime pine bark was used. The pine bark extract was prepared according to the method below.
First, water was added to pine bark, and extraction was performed at 95°C or higher for 1 hour or more. Next, the extract obtained by filtration was purified and dried, and the obtained dry powder was used as a pine bark extract.

As the young barley leaves, a squeezed product (dry powder) of young barley leaves was used. In addition, the juice of young barley leaves was prepared according to the following method.
First, young barley leaves were made into a paste. Next, after pressing, the young barley leaf juice was dried, and the obtained dry powder was used as a young barley leaf juice.

Kudzu flower extract (dry powder) was used as the kudzu flower. In addition, the extract of Kudzu flower was prepared according to the following method.
First, hot water extraction was performed by adding water to dried kudzu flowers. Next, a solid-liquid separation operation was performed to remove insoluble solids, and then a kudzu flower extract was obtained. Thereafter, the extract was dried, and the resulting dry powder was used as a kudzu flower extract.

As black ginger, black ginger rhizome extract (dry powder) was used. In addition, the extract of black ginger was prepared according to the following method.
First, extraction was performed by adding aqueous ethanol to black ginger rhizomes. Next, a solid-liquid separation operation was performed to remove insoluble solids, and then an extract was obtained. After that, dextrin was added to the extract, and the extract was dried, and the obtained dry powder was used as a black ginger extract.

Banaba leaf extract (dry powder) was used as banaba. In addition, the banaba leaf extract was prepared according to the following method.
First, a dried banaba leaf was crushed to obtain a crushed product. Next, aqueous ethanol was added to the pulverized material for extraction to obtain an extract. Thereafter, the extract was dried and pulverized, and the resulting dry powder was used as a banaba leaf extract.

As Terminaria, Terminaria belli
ricca) fruit extract (dry powder) was used. In addition, the water extract of the fruit of Terminalia Berylica was prepared according to the following method.
First, seeds were removed from the fruit of Terminalia Berylica, the remaining part was mixed with water to prepare a mixed solution, and hot water extraction was performed. After extraction, the mixed solution was filtered and the filtrate was freeze-dried, and the resulting dry powder was used as an extract of Terminalia bellirica fruit.

Oyster flesh extract (dry powder) was used as the oyster. The oyster meat extract was prepared according to the method below.
First, the oyster meat was heated, then cooled, enzymes were added, and the mixture was pressurized. Thereafter, it was heat sterilized and dried. The obtained dry powder was used as an oyster meat extract.

As the peel, an extract (dry powder) of the peel of Ganping was used. In addition, the extract of the pericarp of Kanpei was prepared according to the following method.
First, water was added to the peel of Kanpei and heated to perform hot water extraction. Next, the filtrate was collected by filtration, and the obtained filtrate was further filtered and the filtrate was collected to obtain an extract of Kanpei pericarp. The filtrate was dried, and the resulting dry powder was used as an extract of Kanpei pericarp.

Dried powder of Kogomi (young buds of Cycad cycad) was used as the cycad. In addition, the dry powder of Kogomi was prepared according to the following method.
Young cycad shoots were washed, sterilized, steamed, and then dried. Thereafter, the dried product was pulverized and the obtained dry powder was used as a dry powder of Kogomi.

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

[Quantitative Reverse transcriptase-PCR (RT-PCR) reaction]
1. Preparation of total RNA Total RNA was prepared using High Pure RNA Isolation Kit (Roche) according to the product protocol. Furthermore, the reagents and equipment used from the preparation of total RNA to the completion of the reverse transcription reaction were RNase-free.

First, cells were seeded at 5.0×10 ⁴ cells/well in a 5 mL cell culture dish, 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 a DMEM medium containing 10% FBS serum. For comparison (control), DMEM medium containing 0.1% DMSO and 10% FBS serum was added. After 48 hours, completely remove the medium, add 200 μL of 1x PBS for washing, add 400 μL of cell lysate included in the High Pure RNA Isolation Kit, and thoroughly distribute the cell lysate over the entire dish. After dispersion and lysis, the entire cell lysate was collected into a 1.5 mL tube.

The collected sample was thoroughly suspended in a vortex mixer for 60 seconds and lightly spun down. The High Pure filter tube and recovery tube in the kit were assembled, and the cell lysate solution was added to the filter tube. Centrifugation was performed at room temperature and 10,000×g for 15 seconds, the liquid discharged into the collection tube was discarded, and the filter tube and collection tube were reassembled. To a 1.5 mL tube, 90 μL of DNase incubation buffer and 10 μL of DNase I were added and mixed per tube. This mixture was added to the 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 centrifuged at 10,000×g for 15 seconds at room temperature. After centrifugation, the liquid discharged into the collection tube was discarded, and the filter tube and collection tube were reassembled.

500 μL of Wash Buffer II was added to the filter tube and centrifuged at 10,000×g for 15 seconds at room temperature. After centrifugation, the liquid discharged into the collection tube was discarded, and the filter tube and collection tube were reassembled. Furthermore, 200 μL of Wash Buffer II was added, and centrifugation was performed at room temperature and 15,000×g for 2 minutes to wash the filter. After centrifugation, the filter tube was inserted into a new 1.5 mL tube, 100 μL of elution buffer was added to the center of the filter tube, and the tube was left standing at room temperature for 3 minutes. Thereafter, centrifugation was performed at room temperature and 10,000×g for 1 minute. The eluate from this operation was used as an 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 was used in subsequent experiments.

2. Synthesis of cDNA 5 pmol of Oligo (dT) ₂₀ primer (TOYOBO) was added to 1.0 μg of total RNA extracted from cells, and RNase Free water was added so that the total volume was 13 μL. A heat treatment reaction was performed at 65° C. for 5 minutes using a Thermal Cycler PTC 200 (MJ Research), and immediately transferred to ice for quenching. In the meantime, the reverse transcriptase reaction program was advanced to the 42°C stage and then temporarily stopped. A solution containing 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 was added to the samples cooled on ice for 5 minutes, and mixed gently. did. Thereafter, cDNA was synthesized by reacting at 42°C for 20 minutes, at 99°C for 5 minutes, and at 4°C for 5 minutes. This cDNA was used as a template for quantitative PCR.

3. Design of primers Search NCBI (http://www.ncbi.nlm.nih.gov/gene/) for the target gene whose expression level is to be measured by RT PCR, and determine the primer sequence based on the sequence. Synthesized. Primer synthesis was outsourced to Sigma. Primers for detecting β-actin and hTERT, which are internal controls, are shown in Table 1.

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

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

Examples of various aspects of the present invention are listed below, but the technical scope of the present invention is not limited thereto.

<Production Example 1 to Production Example 8>
At least one material selected from pine bark, barley grass, kudzu flower, black ginger, banaba, terminaria, oyster, chinpi, and cycad cycad is blended in the formulation shown in Table 3 below, so that the mixture is uniform. Tablets were manufactured by mixing the mixture and molding using a tableting machine (250 mg per tablet). The resulting edible composition can be expected to have a high effect of enhancing telomerase expression.

<Production Example 9 to Production Example 16>
At least one material selected from pine bark, barley grass, kudzu flower, black ginger, banaba, terminaria, oyster, chinpi, and cycad cycad is blended in the formulation shown in Table 4 below to make it uniform. Hard capsules were manufactured by mixing the mixture with the following ingredients and filling them into a gelatin-containing film (350 mg per capsule). The resulting edible composition can be expected to have a high effect of enhancing telomerase expression.

<Production Example 17 to Production Example 24>
At least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, terminaria, oysters, chinese bark, and cycad cycad is blended in the formulation shown in Table 5 below, so that the mixture is uniform. Soft capsules (300 mg per capsule) were prepared by mixing the ingredients and encapsulating them with a film containing gelatin and glycerin. The resulting edible composition can be expected to have a high effect of enhancing telomerase expression.

<Production Example 25 to Production Example 32>
At least one material selected from pine bark, barley grass, kudzu flower, black ginger, banaba, terminaria, oyster, chinpi, and cycad cycad is blended in the formulation shown in Table 6 below to ensure a uniform consistency. A powdered beverage was produced by mixing with. For example, by mixing 3 g of the resulting edible composition with 150 mL of water and drinking it, a high telomerase expression enhancement effect can be expected.

<Production Example 33 to Production Example 40>
At least one material selected from pine bark, young barley leaves, kudzu flowers, black ginger, banaba, terminaria, oysters, chinpi, and cycads is mixed in the formulation shown in Table 7 below, and the fluidized bed granulation is carried out. The mixture was placed in a machine, mixed with an air stream for several minutes, and granulated by spraying 60 L of water at 2000 mL per minute. Subsequently, the obtained granules were sieved through a 30 mesh sieve to produce granules. The resulting edible composition can be expected to have a high effect of enhancing telomerase expression.

<Production Example 41 to Production Example 56>
A lotion is prepared by blending at least one material selected from pine bark, barley grass, kudzu flower, black ginger, banaba, terminaria, oyster, chinpi, and cycad in the formulations shown in Tables 8 to 11 below. was manufactured. The obtained external preparation can be expected to have a high telomerase expression enhancement effect.

<Production Example 57 to Production Example 64>
At least one material selected from pine bark, barley grass, kudzu flower, black ginger, banaba, terminaria, oyster, chinpi, and cycad cycad is blended in the formulations shown in Tables 12 and 13 below, and the cream is was manufactured. The obtained external preparation can be expected to have a high telomerase expression enhancement effect.

<Production Example 65 to Production Example 72>
At least one material selected from pine bark, barley grass, kudzu flower, black ginger, banaba, terminaria, oyster, chinpi, and cycad cycad is blended in the formulations shown in Tables 14 and 15 below, and a milky lotion is prepared. was manufactured. The obtained external preparation can be expected to have a high telomerase expression enhancement effect.

<Production Example 73 to Production Example 80>
At least one material selected from pine bark, barley grass, kudzu flower, black ginger, banaba, terminaria, oyster, chinpi, and cycad cycad is blended in the formulations shown in Tables 16 and 17 below to form a gel. The drug was manufactured. The obtained external preparation can be expected to have a high telomerase expression enhancement effect.

<Production Example 81 to Production Example 88>
At least one material selected from pine bark, barley grass, kudzu flower, black ginger, banaba, terminaria, oyster, chinpi, and cycad cycad is blended in the formulations shown in Tables 18 and 19 below to produce an aerosol. The drug was manufactured. The obtained external preparation can be expected to have a high telomerase expression enhancement effect.

The telomerase expression enhancer of the present invention is industrially useful because it can be used as a health food and the like.

Claims (3)

A telomerase expression enhancer characterized by containing a pine bark extract (however, a placenta extract extracted from the placenta of a mammal or a placenta of a plant species, oils and fats supplied from animals or plants, and a pine bark extract) (excluding compositions containing bark extract). The telomerase expression enhancer according to claim 1, which is an oral preparation. The telomerase expression enhancer according to claim 1, which is an external preparation.