JP2018095580A - Anti-anxiety composition - Google Patents

Abstract

A novel anxiolytic composition is provided. According to the present invention, one or more selected from the group consisting of theaflavins, resveratrol, β-eudesmol, isoxanthohumol, sphingolipid and its constituents are used as active ingredients. An anti-anxiety composition comprising is provided. The composition of the present invention can be used for suppressing or improving anxiety caused by stress. [Selection figure] None

Description

  The present invention relates to an anxiolytic composition.

  Conventionally, anxiety has been considered as one of psychiatric disorders, and symptoms such as neuropathy, mood disorder, personality disorder, and behavioral disorder are known. On the other hand, even those who have not been diagnosed with mental illness may become uneasy or uncomfortable due to stress in everyday life, as well as stress from events and uncertain situations that they have never experienced. According to a survey by the Cabinet Office (2008 White Paper on National Life), there are various factors in stress, and about half of them feel that they are stressed. Therefore, it can be said that there is a potential demand for suppressing or alleviating anxiety caused by stress.

  So far, ingredients derived from food ingredients related to mental state such as anxiety and brain function are known. For example, Patent Document 1 discloses that theanine, which is a kind of amino acid contained in a large amount in tea leaves, is effective in treating depression of mood disorders. Patent Document 2 discloses that a pharmaceutical composition containing hops is effective in suppressing and improving depression and anxiety.

JP 2003-63958 A JP 2005-272342 A

  An object of the present invention is to provide a novel anxiolytic composition and an anxiolytic agent.

  The present inventors have recently confirmed that ingredients contained in food ingredients such as theaflavins are effective in improving anxiety caused by stress in the elevated plus maze test and the light / dark box test, which are evaluation systems for anxiety. I found it. The present invention is based on these findings.

According to the present invention, the following inventions are provided.
[1] Anti-anxiety comprising, as an active ingredient, one or more selected from the group consisting of theaflavins, resveratrol, β-eudesmol, isoxanthohumol and sphingolipid and its constituent components Compositions and anxiolytic agents.
[2] The composition and the agent according to the above [1], which are used for suppressing or improving anxiety caused by stress.
[3] The composition and agent according to the above [1] or [2], comprising theaflavins as active ingredients.
[4] The composition and agent according to any one of [1] to [3], which are food compositions.
[5] The composition and the agent according to the above [4], wherein the food composition is a food containing a black tea extract, black tea extract or theaflavin preparation.
[6] The composition and agent according to [4] or [5] above, wherein the food composition is a tea beverage.

  ADVANTAGE OF THE INVENTION According to this invention, the composition for anti-anxiety and an anxiolytic agent which comprise the component contained in foodstuffs and food raw materials, such as theaflavins, as an active ingredient are provided. Ingredients used as active ingredients in the present invention are ingredients derived from foods and food ingredients that have been ingested by humans for many years, and are advantageous in that they have no side effects even if they are used continuously and have high safety.

FIG. 1 is a diagram showing the results of an elevated plus maze test. A shows the open arm stay time (s) in the elevated plus maze. * Indicates p <0.05 (Bon feronni test) vs control group. B indicates the number of times of entry into the open arm of the elevated plus maze. 2 is a diagram showing the results of a light and dark box test. A shows the stay time (s) of the light room in the light and dark box. B indicates the number of times the light-dark box has entered the light room.

Detailed description of the invention

  In the present invention, it is selected from the group consisting of theaflavins, resveratrol, β-eudesmol, isoxanthohumol, and sphingolipid and its constituents (hereinafter sometimes referred to as “active ingredient of the present invention”). One type or two or more types can be used as active ingredients. Of these, theaflavin is a dimer of catechin and is a kind of polyphenol. Theaflavins can be a mixture containing one or more selected from the group consisting of theaflavins, theaflavin monogallate and theaflavin digallate.

  In the present invention, theaflavins can use theaflavins derived from foods and food materials in addition to enzyme-treated products and chemically synthesized products. Examples of foods that can be used include fermented tea (including semi-fermented tea) such as black tea and oolong tea. The theaflavins from the fermented tea extract can be prepared according to a conventional method, and in the case of preparation from a black tea extract, for example, it can be prepared according to the description in JP-A-2009-173652.

  In the present invention, theaflavins may be those contained in black tea extract or black tea extract. In the present invention, the theaflavins may be powders, tablets or other preparations containing theaflavins (for example, theaflavin preparations). The black tea extract or black tea extract containing theaflavins can be prepared according to a conventional method, for example, according to the description in JP-A-2009-173652.

  In the present invention, the black tea extract may contain 0.1% by mass or more, preferably 0.15% by mass or more of theaflavins, for example, 0.1 to 3.0% by mass, preferably 0.165%. It can contain -2.5 mass% theaflavins.

  In the present invention, the black tea extract can contain 0.05% by mass or more of theaflavins on a solid content basis, for example, 0.05 to 1.3% by mass of theaflavins.

  In the present invention, the theaflavin preparation can contain 20% by mass or more, preferably 30% by mass or more of theaflavins on the basis of the solid content, for example, 30-100% by mass, preferably 35-100% by mass of theaflavin. Can be included.

  In the present invention, the theaflavins content is measured as theaflavins, theaflavin monogallate and theaflavin digallate by high performance liquid chromatography (HPLC method), and the total amount is defined as theaflavins content. For example, the theaflavins content can be measured according to the method of Hara et al. (Journal of Japanese Society for Agricultural Chemistry, 61 (7), 803-808, 1987).

  The black tea extract or black tea extract contains polyphenols other than theaflavins, and it goes without saying that compositions and agents containing polyphenols other than theaflavins such as thearubidine are also within the scope of the present invention.

  Resveratrol (3,5,4-trihydroxystilbene) which is an active ingredient of the present invention is a phytoalexin (antibacterial substance) having an antioxidant action, and as a polyphenol contained in red wine or red grape juice Is also known. In the present invention, resveratrol can be used as a resveratrol derived from a plant body, a food, and a food material in addition to a synthetic product. Examples of the plant that can be used include grapes, peanuts, and weavers. Preparation of resveratrol from a plant can be performed according to a conventional method, for example, it can be prepared from grape according to the description of JP-A-2001-8695.

  Β Eudesmol, which is an active ingredient of the present invention, is known as an aromatic component with a refreshing scent like cypress contained in essential oils such as hops and eucalyptus. In the present invention, β-eudesmol can be used as a β-eudesmol derived from plants, foods, and food materials in addition to synthetic products. Examples of plants that can be used include hops and eucalyptus trees. Preparation of β-eudesmol from a plant can be performed according to a conventional method.

  Isoxanthohumol, which is an active ingredient of the present invention, is a hop prenylflavonoid (polyphenol) produced in the lupulin gland of hop cones, and is an isomer of xanthohumol. In the present invention, as isoxanthohumol, in addition to synthetic products, isoxanthohumol derived from plants, foods and food materials can be used. Examples of plants that can be used include hops. Preparation of isoxanthohumol from a plant body can be performed according to a conventional method, for example, it can be prepared from hops as described in International Publication No. 2004/0889359.

  The sphingolipid which is an active ingredient of the present invention refers to a complex lipid having sphingoids (long chain aliphatic amine having 2 or 3 hydroxyl groups at the terminal) as a long chain base component, and a glycosphingolipid such as ganglioside, Examples include sphingophospholipids such as sphingomyelin. Ganglioside has a structure in which sialic acid (N-acetylneuraminic acid) is bound on a sugar chain, and examples include ganglioside GD3 and ganglioside GM3. Sphingomyelin has a structure composed of phosphorylcholine and ceramide. Examples of the component of sphingolipid include ceramide, which is a common structure of sphingolipid. Ceramide has a structure in which sphingosine and a fatty acid are amide-bonded, and examples thereof include glucosylceramide and lactoceramide.

  In the present invention, theaflavins, resveratrol, β-eudesmol, isoxanthohumol, and one or more of sphingolipids and components thereof are used as an anti-anxiety composition and an active ingredient of an anxiolytic agent. And can be used as an active ingredient in methods for suppressing or alleviating anxiety.

  That is, according to the present invention, there is provided a method for suppressing or alleviating anxiety, comprising ingesting or administering an effective amount of the active ingredient of the present invention to a human or non-human animal. When the method of the present invention is carried out, an effective amount of the active ingredient of the present invention is administered to a mammal (human, mouse, rat, rabbit, dog, cat, cow, horse, pig, monkey, etc.) in need thereof. Can be implemented.

  The present invention also provides the use of the active ingredients of the present invention for the manufacture of anti-anxiety compositions and foods. The present invention further provides the use of the active ingredient of the present invention for the manufacture of an anxiolytic agent. The present invention further provides an active ingredient of the present invention for use in suppressing or improving anxiety.

  In the present invention, the term “anti-anxiety” means to suppress or improve anxiety, to stabilize mentally, or to prevent or improve psychological / physical disorders caused by anxiety. It shall be used in the meaning including suppressing or improving the anxiety state caused.

  In the present invention, the degree of the “anti-anxiety” action can be evaluated using an anxiety level evaluation system (see Examples 2 and 3) such as an elevated plus maze test and a light / dark box test. Specifically, in at least one of the elevated plus maze test and the light / dark box test, the anxiolytic effect is exhibited when the anxiety level after intake or administration of the test substance is lower than the anxiety level before intake or administration. Can be determined.

  According to the examples described later, the active ingredient of the present invention also suppresses inflammation caused by microglia in the brain. Therefore, according to another aspect of the present invention, one or more selected from the group consisting of theaflavins, resveratrol, β-eudesmol, isoxanthohumol and sphingolipid and its constituents are effective. A composition for use in suppressing brain inflammation and a brain inflammation inhibitor, which are included as components, are provided.

  According to the present invention, there is also provided a method for suppressing inflammation in the brain, comprising ingesting or administering an effective amount of the active ingredient of the present invention to a human or non-human animal. When the method of the present invention is carried out, an effective amount of the active ingredient of the present invention is administered to a mammal (human, mouse, rat, rabbit, dog, cat, cow, horse, pig, monkey, etc.) in need thereof. Can be implemented.

  The present invention also provides the use of the active ingredient of the present invention for the manufacture of a composition and food for use in the suppression of brain inflammation. The present invention further provides use of the active ingredient of the present invention for the production of a brain inflammation inhibitor. The present invention further provides the active ingredient of the present invention for use in suppressing brain inflammation.

  In the present invention, “intracerebral inflammation” mainly refers to excessive activation of microglia caused by various environmental factors such as waste accumulation in the brain, psychological and physical stress, fatigue, invasion of foreign matter, and aging. It refers to inflammation in the brain caused by it, and is intended to include inflammation in the brain in which increased production of inflammatory cytokines, inflammatory chemokines, and reactive oxygen species occurs.

  Active oxygen produced by overactivated microglia and inflammatory cytokines such as TNF-α are closely related to mood disorders including depression, chronic fatigue syndrome, pain, dementia, and multiple sclerosis (Berta, T., et al., 2014, J. Clin. Invest., Vol. 124 (3), pp. 1173-1186; Riazi, K., et al., 2008, Proc. Natl Acad. Sci. USA, Vol. 105 (44), pp. 17151-17156; Grinberg, YY, et al., 2013, J. Neurochem., Vol. 126 (5), pp. 662-672; Couch, Y., et al., 2013, Brain Behav. Immun., Vol.29, pp.139-146; Yasui, M., et al., 2014, Glia, doi: 10.1002 / glia.22687; Nakatomi, Y. , et al., 2014, J. Nucl. Med., Vol. 55, pp. 945-95; Felger and Lotrich, 2013, Neuroscience, Vol. 246, pp. 199-229). Therefore, according to another aspect of the present invention, one or more selected from the group consisting of theaflavins, resveratrol, β-eudesmol, isoxanthohumol and sphingolipid and its constituents are effective. A pharmaceutical composition for use in the treatment and / or prevention of a disease comprising a component for which the suppression of inflammation in the brain is effective for treatment and / or prevention, and the suppression of inflammation in the brain is effective for treatment and / or prevention Provided are therapeutic and prophylactic agents for certain diseases. Diseases for which suppression of inflammation in the brain is effective for treatment and / or prevention include mood disorders including depression, chronic fatigue syndrome, pain, dementia, and multiple sclerosis.

  According to the present invention, there is also provided a method for treating and / or treating a disease in which suppression of inflammation in the brain is effective for treatment and / or prevention, comprising administering an effective amount of the active ingredient of the present invention to a human or non-human animal. A method of preventing is provided. When the method of the present invention is carried out, an effective amount of the active ingredient of the present invention is administered to a mammal (human, mouse, rat, rabbit, dog, cat, cow, horse, pig, monkey, etc.) in need thereof. Can be implemented.

  According to the present invention, there is also provided use of the active ingredient of the present invention for the manufacture of a pharmaceutical composition for use in the treatment and / or prevention of a disease for which inhibition of brain inflammation is effective for the treatment and / or prevention. Provided. The present invention further provides use of the active ingredient of the present invention for the manufacture of a therapeutic and prophylactic agent for a disease for which suppression of inflammation in the brain is effective for treatment and / or prevention. The present invention further provides the active ingredient of the present invention for use in the treatment and / or prevention of diseases for which the suppression of inflammation in the brain is effective for the treatment and / or prevention.

  The use of the active ingredients of the invention in the present invention may be in humans and non-human animals, and is intended for both therapeutic and non-therapeutic uses. As used herein, “non-therapeutic” means not including the act of operating, treating, or diagnosing a human (that is, a medical act on a human), and specifically, receiving a doctor or doctor's instructions It means that the person does not include a method of performing surgery, treatment or diagnosis on humans.

  The anti-anxiety composition and agent of the present invention, the composition and agent for suppressing intracerebral inflammation of the present invention, the pharmaceutical composition of the present invention, and the therapeutic agent and preventive agent of the present invention (hereinafter referred to as “the composition of the present invention”). Can be provided in the form of pharmaceuticals, quasi drugs, foods, feeds, etc., and can be carried out according to the description below. In addition, the method for suppressing or improving anxiety according to the present invention, the method for suppressing intracerebral inflammation according to the present invention, and the method for treating and preventing the present invention can be carried out according to the following description.

  The active ingredients of the present invention such as theaflavins can be orally administered to humans and non-human animals. Oral preparations include granules, powders, tablets (including sugar-coated tablets), pills, capsules, syrups, emulsions, and suspensions. These preparations can be formulated using a pharmaceutically acceptable carrier by a technique usually performed in this field. Pharmaceutically acceptable carriers include excipients, binders, diluents, additives, fragrances, buffers, thickeners, colorants, stabilizers, emulsifiers, dispersants, suspending agents, preservatives, etc. For example, magnesium carbonate, magnesium stearate, talc, sugar, lactose, pectin, dextrin, starch, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose, low melting point wax, cocoa butter can be used as a carrier.

  The production method of the oral preparation is not particularly limited, and any method can be used. Active ingredients include excipients (eg lactose, sucrose, starch, mannitol), disintegrants (eg calcium carbonate, carboxymethylcellulose calcium), binders (eg pregelatinized starch, gum arabic, carboxymethylcellulose, polyvinylpyrrolidone, Hydroxypropyl cellulose), stabilizers and / or lubricants (eg, talc, magnesium stearate, polyethylene glycol 6000) and then compression molded, then, if necessary, taste masking, intestinal It can be produced by coating by a known method for the purpose of solubility or durability. As the coating agent, for example, ethyl cellulose, hydroxymethyl cellulose, polyoxyethylene glycol, cellulose acetate phthalate, hydroxypropyl methylcellulose phthalate, Eudragit (Rohm) and the like can be used. Capsules may be produced by using the active ingredient as it is or by blending the active ingredient with pharmaceutical additives such as stabilizers, dispersants, colorants and / or preservatives. The capsule may be a soft capsule or a hard capsule, and the material of the capsule is not particularly limited. For example, gelatin, vegetable fiber, starch, or the like may be used, and an adhesion preventing measure may be taken.

  When the active ingredients of the present invention such as theaflavins are provided as food, they can be provided as food as they are, or can be provided by containing them in food. The food thus provided is a food containing an effective amount of the active ingredient of the present invention. In the present specification, “containing an effective amount” of the active ingredient of the present invention means that the active ingredient of the present invention is ingested within the range described later when an amount normally consumed in each food is ingested. Content. “Food” means health food, functional food, nutritional supplement, supplement, health functional food (eg, food for specified health use, functional nutrition food, functional indication food), food for special use (eg, food for infants) , Food for pregnant women, food for sick people).

  The form of “food” is not particularly limited, and may be, for example, a drink form, a semi-liquid form or a gel form. Examples of supplements include tablets produced by tableting after adding an excipient, a binder and the like to the active ingredient of the present invention, and capsules encapsulated in capsules.

  The food provided by the present invention is not particularly limited as long as it contains the active ingredient of the present invention. For example, foods and drinks containing carbohydrates such as rice, noodles, breads and pasta; cookies, cakes, etc. Western confectionery, Japanese confectionery such as Shantou and Yokan, candy, gum, frozen confectionery such as yogurt and pudding, and various confectionery such as ice confectionery; whiskey, bourbon, spirits, liqueur, wine, fruit liquor, Japanese sake, Chinese liquor, Alcoholic beverages such as shochu, beer, non-alcohol beer with alcohol content of 1% or less, happoshu, other miscellaneous liquor, strawberry high; beverages containing fruit juice, beverages containing vegetable juice, beverages containing fruit juice and vegetable juice, soft drinks, carbonated beverages , Milk, soy milk, milk drinks, drink-type yogurt, drink-type jelly, coffee, cocoa, tea drinks, energy drinks, sports Link, mineral water, non-alcoholic beverages such as near-water; processed products using egg, (including delicacies) processed products of seafood and meat (including offal lever or the like) and the like can be exemplified. The mineral water includes both foaming and non-foaming mineral water.

  Tea beverages include fermented tea, semi-fermented tea and non-fermented tea, for example, black tea, green tea, barley tea, brown rice tea, sencha tea, gyokuro tea, roasted tea, oolong tea, turmeric tea, puer tea, rooibos tea. , Rose tea, chrysanthemum tea, herbal tea (for example, mint tea, jasmine tea). When using theaflavins as an active ingredient of the composition and agent of the present invention, it contains a tea extract, a tea extract or a theaflavin preparation, a tea extract, a tea extract or a tea extract, Naturally included foods (eg, tea beverages) are within the scope of the food of the present invention.

  Examples of the fruit used in fruit juice-containing beverages and fruit and vegetable juice-containing beverages include apples, mandarin oranges, grapes, bananas, pears, peaches, mangoes, acais, blueberries, and umes. Examples of vegetables used in vegetable juice-containing beverages, fruit juices, and vegetable juice-containing beverages include tomato, carrot, celery, pumpkin, cucumber, and watermelon.

  The intake or dose of the active ingredient of the present invention can be determined depending on the recipient's sex, age and weight, symptoms, administration time, dosage form, administration route, combined drugs and the like. Among the active ingredients of the present invention, when taking or administering theaflavins for the purpose of anxiolytic action, the daily intake and dose for adults is 0.002 to 10 mg / kg body weight (preferably 0.002 to 0.002). 0.03 mg / kg body weight) and may be taken or administered in several doses depending on the subject. In addition, among the active ingredients of the present invention, when theaflavins are ingested or administered for the purpose of inhibiting inflammation in the brain (or treating and preventing diseases that can be treated and prevented by inhibiting brain inflammation), The intake and dose are 0.002 to 10 mg / kg body weight (preferably 0.002 to 0.03 mg / kg body weight). Depending on the subject, the intake and administration may be divided into several times. .

  Since the composition and the preparation of the present invention use ingredients derived from foods and food ingredients that humans have taken for many years as active ingredients, there is no concern about side effects even if they are used continuously, and they are highly safe. For this reason, when the composition and agent of the present invention are used in combination with an existing drug, the dose of the existing drug can be reduced, and as a result, the side effects of the existing drug can be reduced or eliminated. In combination with other drugs, even if the composition and the agent of the present invention are separately prepared, the other drug and the composition and the agent of the present invention are combined in the same composition. Also good.

  The composition and the agent of the present invention can be provided as a composition comprising the active ingredient of the present invention in an amount effective for anxiolytic action. In this case, the composition and the preparation of the present invention may be packaged so that an effective intake can be ingested, and as long as the effective intake can be ingested, the packaging form may be one package or multiple packages. Also good.

  When the composition and agent of the present invention are provided in a package form, a description relating to the intake is made on the package so that an effective intake can be taken, or a document with the description is provided together Is desirable. Moreover, when providing the effective intake for one day with a plurality of packages, a plurality of packages with the effective intake for one day can be provided as a set for convenience of intake.

  When providing the composition and the preparation of the present invention in a package form, it can be provided in a unit package form from the viewpoint of convenience of intake, and is preferably provided in a unit package form per serving. Here, “unit packaging form per meal” is a form in which the intake amount per meal is determined in advance, for example, as a food or drink that can be taken orally in a specific amount, not only general foods , Beverages (including drinks), health supplements, health functional foods, supplements, and other forms.

  Even if the intake per meal determined in advance in the unit packaging form is the effective intake per day, the effective intake per day is divided into two or more (preferably two or three times). It may be the intake amount. Therefore, the active ingredient of the present invention can be blended in the unit packaging form with the daily intake of the adult, or half or one third of the daily intake of the adult. The active ingredient of the present invention can be blended in an amount.

  The packaging form for providing the composition and the preparation of the present invention is not particularly limited as long as it defines a certain amount. For example, wrapping paper, bag, soft bag, paper container, can, bottle, capsule, etc. Examples include containers that can be accommodated.

  The composition and the preparation of the present invention can exert their effects by single intake or single administration. The composition and agent of the present invention are also preferably administered or ingested continuously for 3 days or more in order to exert their effects over a longer period, and the administration and ingestion period is more preferably about 1 week, especially Preferably it is about 2 weeks. Here, “continuously” means to continue administration or ingestion every day. When the composition and the agent of the present invention are provided in a package form, an effective intake for a certain period (for example, one week) may be provided as a set for continuous intake.

The anti-anxiety composition and agent of the present invention may be labeled with an anti-anxiety action. In this case, the following part or all of the following indications may be attached to the anti-anxiety composition and the preparation of the present invention in order to make the indication easy for consumers to understand. In the present invention, it goes without saying that anxiolytic action and suppression or improvement of anxiety are used in the meaning including the following indications.
・ Lighten up feelings ・ Strengthen stress ・ Relieve stress ・ Relieve anxious feelings ・ Relieve calm feelings ・ Relieve thin feelings ・ Lighten up ・ Slightly less worried ・ Psychological burden・ Relieve mental load ・ Be positive

  The present invention will be described more specifically based on the following examples, but the present invention is not limited to these examples.

Example 1 Screening of Inflammatory Inhibitor in Brain (1) Isolation of Microglia Cells Brain tissue dispersion was obtained by removing brain from mice and treating with papain enzyme. After stopping the papain enzyme reaction, microglia cells are reacted with a CD11b antibody (manufactured by Miltenyi Biotec), which is a pan-microglia marker labeled with microbeads, and CD11b-positive cells are adsorbed on the column by the MACS method in which the cells on which the microbeads have acted are adsorbed. As microglia cells were isolated.

(2) Preparation of theaflavins Theaflavins are theaflavin TF40 (theaflavin, theaflavin monogallate, theaflavin digallate total concentration of about 40% w / w, manufactured by Yaizu Suisan Kagaku Kogyo Co., Ltd.) containing 0.1% phosphoric acid and 22% acetonitrile. It was dissolved in water, subjected to reverse phase preparative HPLC, washed with 22% acetonitrile water containing 0.1% phosphoric acid, and eluted with 70% acetonitrile water containing 0.1% phosphoric acid. The eluate was fractionated while monitoring the absorbance at a wavelength of 280 nm at which theaflavin has an absorption maximum. Among the fractions, fractions having a total purity of theaflavins of 90% or more were collected and added up.

  Acetonitrile in the combined fractions was removed under reduced pressure with an evaporator, ethyl acetate was added, and theaflavins were extracted as an ethyl acetate layer. The ethyl acetate layer was collected, washed with saturated brine, dehydrated with sodium sulfate, and then the ethyl acetate was removed under reduced pressure to obtain theaflavins solids. The obtained theaflavins solid was dissolved in a small amount of acetonitrile, and water was added to suspend it uniformly, followed by lyophilization. The purity of theaflavins in the brown powder after lyophilization was> 95%. The purity of theaflavins was confirmed according to the method described in Japanese Journal of Agricultural Chemistry 1987, 61, 803-808.

(3) Screening method The microglia cells isolated in the above (1) were cultured for 12 hours in a medium to which various test substances shown below were added at various concentrations. The test substances and their concentrations were as follows. That is, the theaflavins prepared in the above (2), resveratrol (manufactured by SIGMA-ALDRICH), β-eudesmol (manufactured by SIGMA-ALDRICH) and isoxanthohumol (manufactured by SIGMA-ALDRICH) are each 0. Concentrations of 12.5, 50, and 100 μM, 3-sialic acid (manufactured by Nagara Science), 6-sialic acid (manufactured by Nagara Science), sphingomyelin (manufactured by Nagara Science), ganglioside GD3 (manufactured by Nagara Science) Ganglioside GM3 (manufactured by Nagara Science), glycosylceramide (manufactured by Nagara Science) and lactceramide (manufactured by Nagara Science) were adjusted to concentrations of 0, 10, 20, 40 and 80 μg / mL, respectively.

  After culturing microglial cells for 12 hours in the presence of the test substance, in order to induce the production of inflammatory cytokines accompanying the induction of inflammation in the microglia cells, a medium containing lipopolysaccharide (manufactured by SIGMA-ALDRICH) at a concentration of 5 ng / mL In addition, interferon gamma (manufactured by R & D Systems) was added to the medium to a concentration of 0.5 ng / mL and further cultured for 12 hours. After culturing, TNF-α contained in the culture supernatant was quantified using an ELISA kit (manufactured by eBioscience) to evaluate the inflammatory state of microglia cells cultured in the presence of each test substance.

(4) Screening results Table 1 shows the results.

  From the results shown in Table 1, in the presence of resveratrol, theaflavins, β-eudesmol, isoxanthohumol, 3-sialic acid, 6-sialic acid, sphingomyelin, ganglioside GD3, ganglioside GM3, glycosylceramide, and lactoceramide It was confirmed that the microglial cells that received inflammation stimulation in (2) had a reduced TNF-α concentration in the culture supernatant depending on the concentration of the test substance, and the inflammation was suppressed.

Example 2: Evaluation of anti-anxiety action (1)
(1) Administration of theaflavins Five-week-old ddy male mice (obtained from Japan SLC Co., Ltd., the same applies hereinafter) were divided into groups (a total of 3 groups, control group and 2 experimental groups, each group n = 10). The test substance should be distilled water so that the theaflavin dosage is 0 mg / kg body weight in the control group, and distilled water so that the theaflavin dosage is 1 mg / kg body weight and 5 mg / kg body weight in the experimental group. The prepared theaflavins solution was forcibly administered into the stomach once each day for 5 consecutive days.

(2) Elevated plus maze test The elevated plus maze test is an evaluation system for anxiety levels. The elevated cross maze consists of a cross-shaped platform installed 50cm above the floor, the two open arms facing each other (30cm x 6cm) have no walls, and the other two closed arms have walls These arms are connected at the center. Although the mouse has a desire for a new search, it feels anxiety and fear because it is placed in an open state with no walls on the open arm, and the stay time in the open arm is usually short. The increase in the open arm residence time is considered to reflect the anxiolytic effect of the administered test substance. The mouse of (1) was subjected to an elevated plus maze 1 hour after the final administration of theaflavins and allowed to freely explore the maze for 2 minutes and 30 seconds. The open arm stay time and the number of intrusions into the open arm during the search time were measured. The measured value was expressed as an average value ± standard error.

(3) Results The results are shown in FIG. As shown in FIG. 1, the open arm residence time increased in the theaflavin administration group in a concentration-dependent manner, and the open arm residence time in the theaflavin 5 mg / kg body weight administration group was significant compared to the control group (risk rate of less than 5%) ). That is, it was confirmed by the elevated plus maze test that theaflavins have an anxiolytic action. In addition, a single administration of theaflavins also showed a tendency to exert anxiolytic effects in the elevated plus maze test (data not shown). In the results of FIG. 1, the number of intrusions into the open arm of the control group tended to be larger than that in the theaflavins administration group. This is thought to be due to the large number of times of exiting immediately after entering.

Example 3: Evaluation of anti-anxiety action (2)
(1) Administration of theaflavins 5-week-old ddy male mice were divided into 3 groups (a total of 3 groups: a control group and 2 experimental groups). For the test substance, distilled water was used so that the theaflavin dosage was 0 mg / kg body weight in the control group (n = 10), and the theaflavins dosage was 1 mg / kg body weight (n = 9) and 5 mg in the experimental group. A theaflavin solution prepared with distilled water so as to be / kg body weight (n = 10) was forcibly administered into the stomach once a day for 6 consecutive days.

(2) Light / dark box test The light / dark box test is an evaluation system for anxiety levels. The light / dark box is composed of a light room (23 cm × 24 cm) where light enters and a light-shielded dark room of the same size as the light room. Since the mouse has a habit of preferring a dark place and dislikes a bright place, the staying time in a bright room is usually reduced. The increase in staying time in the light room is considered to reflect the anxiolytic effect of the test substance. The mouse of (1) was subjected to a light and dark box 1 hour after the final administration of the theaflavins and allowed to freely search the light and dark box for 5 minutes. The staying time in the light room and the number of intrusions into the light room during the search time were measured. The measured value was expressed as an average value ± standard error.

(3) Results The results are shown in FIG. As shown in FIG. 2, in the theaflavins administration group, the staying time in the bright room increased in a concentration-dependent manner, and the number of times of entering the bright room increased. That is, it was confirmed by the light and dark box test that theaflavins have an anxiolytic action. In addition, a single administration of theaflavins also showed a tendency to exert anxiolytic effects in the light / dark box test (data not shown).

Claims (6)

  An anti-anxiety composition comprising, as an active ingredient, one or more selected from the group consisting of theaflavins, resveratrol, β-eudesmol, isoxanthohumol and sphingolipid and its constituent components.   The composition according to claim 1, which is used for suppressing or improving anxiety caused by stress.   The composition of Claim 1 or 2 which comprises theaflavins as an active ingredient.   The composition according to any one of claims 1 to 3, which is a food composition.   The composition according to claim 4, wherein the food composition is a food containing a black tea extract, a black tea extract or a theaflavin preparation.   The composition according to claim 4 or 5, wherein the food composition is a tea beverage.

Images