Skin barrier function improver and skin condition improver

Abstract

To provide a skin barrier function improver and a skin condition improver.SOLUTION: Lactic acid bacteria belonging to the genus Fructobacillus, a culture of the lactic acid bacteria, a culture supernatant of the lactic acid bacteria, and / or an extract of the lactic acid bacteria are suitable as active ingredients in skin barrier function improvers and skin condition improvers.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to a skin barrier function improving agent and a skin condition improving agent. [Background technology]

[0002] Wrinkles are a symptom of skin aging caused by factors such as aging, stress, and UV exposure. They are easily noticeable and have a significant impact on the appearance of the face. For this reason, there is a great deal of interest in wrinkles and how to improve them.

[0003] Previously, moisturizing skin care products containing water-retaining polymers such as mucopolysaccharides and collagen have been used to improve wrinkles, but this alone has not been sufficient to improve wrinkles. However, the mechanism of wrinkle formation is complex and difficult to reproduce experimentally, so that the mechanism has not yet been fully elucidated. Nevertheless, recent research has revealed that in addition to aging being an important factor in skin aging symptoms, factors such as dryness, oxidation, glycation, and ultraviolet rays also play a major role. Specifically, the factors, especially ultraviolet rays exposure, cause cell damage and accelerated cell apoptosis, a decrease in the proliferation activity of fibroblasts, the main cells in the dermis, and a decrease in the synthesis function of collagen, resulting in a decrease in the turnover rate of fibrous components such as collagen, the breakdown of fiber bundles due to the enhancement of inflammatory cytokines, the accumulation of waste products due to a decrease in the vascular system, and a decrease in nutrient supply, which are thought to result in a loss of skin elasticity and the development of wrinkles.

[0004] Patent Document 1 describes that aminocarboxylic acid derivatives having specific structures and acid addition salts thereof exhibit excellent wrinkle-reducing effects. Patent Document 2 describes that the adhesion and flexibility of the applied film to the skin are improved by combining an ethyl betaine methacrylate / acrylates copolymer with polyvinylpyrrolidone.

[0005] Patent Document 3 describes a skin condition improving agent or a food composition for improving skin condition, which contains inulin as an active ingredient. Patent Document 4 describes an orally ingested agent for suppressing a decrease in skin viscoelasticity that utilizes α-glycosyl hesperidin.

[0006] Patent Document 5 describes a transepidermal water loss inhibitor containing, as an active ingredient, lactic acid bacteria (Leuconostoc mesenteroides) derived from kudzu vines. [Prior art documents] [Patent documents]

[0007] [Patent Document 1] Japanese Patent Application Publication No. 2019-206483 [Patent Document 2] Japanese Patent Application Publication No. 2020-164431 [Patent Document 3] Japanese Patent Publication No. 2022-72515 [Patent Document 4] Japanese Patent Application Laid-Open No. 2010-100634 [Patent Document 5] Japanese Patent Publication No. 2022-72433 Summary of the Invention [Problem to be solved by the invention]

[0008] An object of the present invention is to provide a skin barrier function improving agent and a skin condition improving agent. [Means for solving the problem]

[0009] The present inventors have newly discovered that certain lactic acid bacteria and / or components derived from certain lactic acid bacteria have the effect of improving skin barrier function (skin barrier function improvement) and the effect of improving skin condition (skin condition improvement effect), and have completed the present invention.

[0010] That is, the present invention provides the following aspects. Item 1. A skin barrier function improving agent comprising lactic acid bacteria belonging to the genus Fructobacillus, a culture of said lactic acid bacteria, a culture supernatant of said lactic acid bacteria, and / or an extract of said lactic acid bacteria. Item 2. The skin barrier function improving agent according to Item 1, wherein the lactic acid bacterium is Fructobacillus fructosus. Item 3. The skin barrier function improving agent according to Item 1, wherein the lactic acid bacterium is Fructobacillus fructosus OS-1010 strain (accession number: NITE BP-03818). Item 4. The skin barrier function improving agent according to Item 1, which is used to improve dry skin, suppress transepidermal water loss, or suppress skin thickening. Item 5. A food, drink, cosmetic, or pharmaceutical product comprising the skin barrier function improving agent according to Item 1. Item 6. A skin condition improving agent comprising lactic acid bacteria belonging to the genus Fructobacillus, a culture of said lactic acid bacteria, a culture supernatant of said lactic acid bacteria, and / or an extract of said lactic acid bacteria. Item 7. The skin condition improving agent according to Item 6, wherein the lactic acid bacterium is Fructobacillus fructosus. Item 8. The skin condition improving agent according to claim 6, wherein the lactic acid bacterium is Fructobacillus fructosus strain OS-1010 (accession number: NITE BP-03818). Item 9. The skin condition improving agent according to Item 6, which is used for improving or inhibiting a decrease in skin viscoelasticity, improving or inhibiting skin wrinkles, improving or inhibiting skin sagging, or improving skin firmness and elasticity. Item 10. A food, drink, cosmetic, or pharmaceutical product comprising the skin condition improving agent according to Item 6. [Effects of the Invention]

[0011] The present invention provides novel materials useful as active ingredients for skin barrier function improving agents and skin condition improving agents. Specifically, the present invention provides lactic acid bacteria belonging to the genus Fructobacillus, cultures of the lactic acid bacteria, culture supernatants of the lactic acid bacteria, and / or extracts of the lactic acid bacteria as active ingredients for skin barrier function improving agents and skin condition improving agents. [Brief explanation of the drawings]

[0012] [Figure 1] FIG. 1 is a diagram showing the amount of change in score for each item of a visual analogue scale (hereinafter referred to as VAS) questionnaire. DETAILED DESCRIPTION OF THE INVENTION

[0013] The skin barrier function improving agent and skin condition improving agent of the present invention are characterized in that they contain, as active ingredients, lactic acid bacteria belonging to the genus Fructobacillus, a culture of the lactic acid bacteria, a culture supernatant of the lactic acid bacteria, and / or an extract of the lactic acid bacteria. The skin barrier function improving agent and the skin condition improving agent will be described in detail below.

[0014] [1. Active ingredient] The active ingredients of the skin barrier function improving agent and skin condition improving agent of the present invention are lactic acid bacteria belonging to the genus Fructobacillus, a culture of the lactic acid bacteria, a culture supernatant of the lactic acid bacteria, and / or an extract of the lactic acid bacteria.

[0015] [1-1. Lactic acid bacteria belonging to the genus Fructobacillus] Lactic acid bacteria belonging to the genus Fructobacillus (hereinafter also referred to as "predetermined lactic acid bacteria") are microorganisms that produce nicotinamide mononucleotide (NMN) and / or nicotinamide adenine dinucleotide (NAD). Furthermore, lactic acid bacteria belonging to the genus Fructobacillus are preferably microorganisms that produce at least NAD, and more preferably microorganisms that produce both NMN and NAD.

[0016] Examples of lactic acid bacteria belonging to the genus Fructobacillus include Fructobacillus durionis, Fructobacillus tropaeoil, and Fructobacillus fructosus.

[0017] More specific examples of lactic acid bacteria belonging to the genus Fructobacillus include the Fructobacillus durionis strain RD011727, the Fructobacillus durionis strain NBRC113239, the Fructobacillus tropaeoil strain RD012353, the Fructobacillus tropaeoil strain RD012354, the Fructobacillus fructosus strain NBRC3516, and the Fructobacillus fructosus strain OS-1010.

[0018] The Fructobacillus durionis RD011727 strain was internationally deposited at the National Institute of Technology and Evaluation, Patent Microorganisms Depositary (NPMD) (Room 122, 2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, Japan) on October 28, 2020, under the deposit number NITE BP-02764.

[0019] The Fructobacillus tropaeoil RD012353 strain was internationally deposited at the National Institute of Technology and Evaluation, Patent Microorganisms Depositary (NPMD) (Room 122, 2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, Japan) on October 28, 2020, under the deposit number NITE BP-02765.

[0020] The Fructobacillus tropaeoil RD012354 strain was internationally deposited at the National Institute of Technology and Evaluation, Patent Microorganisms Depositary (NPMD) (Room 122, 2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, Japan) on October 28, 2020, under the deposit number NITE BP-02766.

[0021] The Fructobacillus fructosus OS-1010 strain has been internationally deposited with the National Institute of Technology and Evaluation, Patent Microorganisms Depositary (NPMD) (Room 122, 2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, Japan) on January 5, 2024, under the accession number NITE BP-03818.

[0022] These lactic acid bacteria belonging to the genus Fructobacillus may be used alone or in combination of two or more species.

[0023] From the viewpoint of more effectively improving the effect of improving the skin barrier function (skin barrier function improvement), the lactic acid bacteria belonging to the genus Fructobacillus are preferably Fructobacillus tropaeoil and Fructobacillus fructosus, and more preferably Fructobacillus fructosus.

[0024] Among these lactic acid bacteria belonging to the genus Fructobacillus, from the viewpoint of more effectively improving the effect of improving the skin barrier function (skin barrier function improvement), the Fructobacillus durionis strain RD011727, the Fructobacillus durionis strain NBRC113239, the Fructobacillus tropaeoil strain RD012353, the Fructobacillus tropaeoil strain RD012354, the Fructobacillus fructosus strain NBRC3516, and the Fructobacillus fructosus strain OS-1010 are preferred, and the Fructobacillus tropaeoil strain RD012353, the Fructobacillus tropaeoil strain RD012354, the Fructobacillus fructosus strain NBRC3516, and the Fructobacillus fructosus strain OS-1010 are more preferred. Examples of suitable strains include the Fructobacillus fructosus OS-1010 strain, more preferably the Fructobacillus fructosus NBRC3516 strain, and the Fructobacillus fructosus OS-1010 strain, particularly preferably the Fructobacillus fructosus OS-1010 strain.

[0025] From the viewpoint of more effectively improving the effect of improving skin condition (skin condition improving effect), the lactic acid bacteria belonging to the genus Fructobacillus are preferably Fructobacillus tropaeoil and Fructobacillus fructosus, and more preferably Fructobacillus fructosus.

[0026] Among these lactic acid bacteria belonging to the genus Fructobacillus, from the viewpoint of further effectively improving the effect of improving skin conditions (skin condition improving effect), the Fructobacillus durionis strain RD011727, the Fructobacillus durionis strain NBRC113239, the Fructobacillus tropaeoil strain RD012353, the Fructobacillus tropaeoil strain RD012354, the Fructobacillus fructosus strain NBRC3516, and the Fructobacillus fructosus strain OS-1010 are preferred, and the Fructobacillus tropaeoil strain RD012353, the Fructobacillus tropaeoil strain RD012354, the Fructobacillus fructosus strain NBRC3516, and the Fructobacillus fructosus strain OS-1010 are more preferred. Examples of suitable strains include the Fructobacillus fructosus OS-1010 strain, more preferably the Fructobacillus fructosus NBRC3516 strain, and the Fructobacillus fructosus OS-1010 strain, particularly preferably the Fructobacillus fructosus OS-1010 strain.

[0027] [1-2. Form of active ingredient] The active ingredient is in the form of at least one of the above-mentioned specified lactic acid bacteria, a culture of the lactic acid bacteria, a culture supernatant of the lactic acid bacteria, and an extract of the lactic acid bacteria. The active ingredient may be used in any one of these forms alone or in combination of two or more forms.

[0028] [1-2-1.Lactic acid bacteria] When the active ingredient is in the form of lactic acid bacteria, the active ingredient includes bacterial components of the lactic acid bacteria and products that are or were present in the lactic acid bacteria. The lactic acid bacteria may be either live or killed. In the case of killed bacteria, the bacterial cells may be crushed. In the case of crushed bacteria, some of the bacterial components may be removed. Furthermore, the lactic acid bacteria may be in the form of bacterial cell powder dried by means of freeze-drying, shelf drying, spray drying, or the like. Among these, bacterial cell powder dried by, for example, freeze-drying is preferred from the viewpoint of further enhancing the effect of improving skin barrier function (skin barrier function improvement) and the effect of improving skin condition (skin condition improvement effect).

[0029] Lactic acid bacteria can be obtained by a production method including a step of culturing one or more of the above-mentioned specific lactic acid bacteria and a step of separating and recovering the bacterial cells. The separation and recovery step involves solid-liquid separation and washing of the bacterial cells. The production method can include other steps in addition to the above steps. These other steps include a resting cell reaction step (which can be carried out after the culturing step, preferably after the separation and recovery step), a drying step (which can be carried out after the separation and recovery step or the resting cell reaction), and a bacterial cell disruption step (which can be carried out after the separation and recovery step, the resting cell reaction step, or the drying step).

[0030] Media that can be used in the culturing step include those used for expansion culture (pre-culture medium) and those used for production culture (main culture medium). The main culture medium can be prepared based on the medium used as the pre-culture medium, with further additives added. The medium is preferably a liquid medium, but may also be an agar medium. In addition to a carbon source, the medium generally contains a nitrogen source, minerals, etc.

[0031] Carbon sources include carbohydrates and carbohydrate materials. Carbohydrates include sugars (monosaccharides, disaccharides, oligosaccharides), polysaccharides, and sugar alcohols. Examples of carbohydrates include lactose, sucrose, glucose, starch, xylitol, dextrose, and the like. The carbohydrate material may be any organic composition containing carbohydrates, such as milk and its processed products (skim milk powder, whey, milk powder, condensed milk, etc.), soy milk and its processed products (soy milk hydrolysate, etc.), grains, fruits, and vegetables. Milk may be derived from any mammal, such as cow, goat, sheep, buffalo, camel, llama, donkey, yak, horse, or reindeer. Carbohydrates may be isolated or contained in a carbohydrate material. For example, fructose (a carbohydrate) may be used in the form contained in fruit (a carbohydrate material). These carbon sources may be used alone or in combination. Among these carbon sources, glucose is preferred.

[0032] The concentration of the carbon source in the medium is not particularly limited and may be set appropriately depending on the type of medium, the culture method, etc., but may be, for example, 0.5 to 4 w / w%, preferably 1 to 3 w / w%, and more preferably 1.5 to 2.5 w / w%.

[0033] Any inorganic or organic nitrogen source can be used as the nitrogen source. Examples include proteins such as yeast extract (e.g., brewer's yeast), meat extract, and casein; protein hydrolysates such as peptone (e.g., protease peptone); peptides such as peptides; and nitrogen-containing salts such as ammonium salts (e.g., ammonium citrate) and nitrates. These nitrogen sources may be used alone or in combination.

[0034] The concentration of the nitrogen source in the medium is not particularly limited and may be set appropriately depending on the type of medium, culture method, etc., but in the case of proteins, the concentration may be, for example, 0.3 to 4 w / w%, preferably 0.5 to 3 w / w%, and more preferably 1 to 2 w / w%; in the case of peptides, the concentration may be, for example, 0.1 to 2 w / w%, preferably 0.3 to 1.8 w / w%, and more preferably 0.5 to 1.5 w / w%; and in the case of nitrogen-containing salts, the concentration may be, for example, 0.03 to 1.5 w / w%, preferably 0.05 to 1 w / w%, and more preferably 0.1 to 0.5 w / w%.

[0035] Examples of minerals include manganese (e.g., manganese salts such as manganese sulfate), zinc, iron, sodium (e.g., sodium salts such as sodium acetate), potassium (e.g., potassium salts such as dipotassium hydrogen sulfate and dipotassium hydrogen phosphate), magnesium (e.g., magnesium salts such as magnesium sulfate), calcium, phosphorus (e.g., phosphates such as dipotassium hydrogen phosphate), sulfur (e.g., sulfates such as manganese sulfate, potassium hydrogen sulfate, and magnesium sulfate), and trace elements. These minerals may be used alone or in combination. Of these minerals, manganese, sodium, magnesium, and potassium are preferred.

[0036] The concentration of minerals in the medium is not particularly limited and may be set appropriately depending on the type of medium, culture method, etc. In the case of manganese salts, for example, the concentration is 0.001 to 0.01 w / w%, preferably 0.003 to 0.008 w / w%; in the case of sodium salts, for example, the concentration is 0.05 to 1.5 w / w%, preferably 0.1 to 1 w / w%; in the case of magnesium salts, for example, the concentration is 0.001 to 0.02 w / w%, preferably 0.005 to 0.015 w / w%; in the case of potassium salts, for example, the concentration is 0.05 to 1 w / w%, preferably 0.1 to 0.5 w / w%; in the case of phosphates, for example, the concentration is 0.05 to 1 w / w%, preferably 0.1 to 0.5 w / w%; and in the case of sulfates, the concentration is 0.001 to 0.04 w / w%, preferably 0.005 to 0.02 w / w%.

[0037] In addition to the above components, the medium may contain other components such as vitamins (such as B vitamins), surfactants (nonionic surfactants (such as Tween), anionic surfactants (such as SDS)), antibacterial agents (such as triclosan), and antibiotics (such as monesin). These other components may be used alone or in combination. Among these other components, surfactants are preferred, and nonionic surfactants are more preferred.

[0038] The concentration of other components in the medium is not particularly limited and may be set appropriately depending on the types of other components, the type of medium, the culture method, etc. When a surfactant is contained, the concentration of the surfactant may be, for example, 0.01 to 0.5 w / w%, preferably 0.05 to 0.3 w / w%.

[0039] The culture conditions are not particularly limited as long as they allow the growth of lactic acid bacteria belonging to the genus Fructobacillus.

[0040] The culture temperature may be any temperature that is optimal for the lactic acid bacteria belonging to the genus Fructobacillus to be cultured, and may be, for example, 26 to 40° C., preferably 27 to 38° C., more preferably 28 to 36° C., and even more preferably 29 to 34° C. The culture time may be appropriately set depending on the type of lactic acid bacteria belonging to the genus Fructobacillus to be actually cultured, and may be, for example, 4 to 48 hours, preferably 8 to 36 hours, and more preferably 12 to 24 hours.

[0041] Regarding the operation during the culture, stirring of the culture solution during the culture is not an issue. Furthermore, as a specific example of the culture procedure, the above-mentioned culture is carried out as a production culture (main culture) for a certain period of time, and before that, an expansion culture (preculture) can be carried out in a small amount of medium (for example, 1 / 6 to 1 / 4 of the volume of the main culture medium). The culture conditions for the preculture can be appropriately set according to the type of lactic acid bacteria belonging to the genus Fructobacillus, and the above-mentioned conditions can be used. Furthermore, in the main culture, the culture obtained in the preculture can be inoculated into the main culture medium so that the OD660 becomes, for example, 0.01 to 0.04, preferably 0.01 to 0.03.

[0042] In the separation and recovery step, solid-liquid separation of the culture solution can be carried out by filtration using filter paper, centrifugation, decantation, screw press, roller press, rotary drum screen, belt screen, vibrating screen, multi-plate vibrating filter, vacuum dehydration, pressurized dehydration, belt press, centrifugal concentration dehydration, multi-disk dehydration, or the like, and the obtained bacterial cells can be washed.

[0043] The resting cell process can increase the nicotinamide mononucleotide content in lactic acid bacteria.

[0044] The liquid used in the resting cell reaction step is not particularly limited as long as it allows the resting cell reaction of lactic acid bacteria belonging to the genus Fructobacillus, and examples thereof include water, buffer solutions, and organic solvents.

[0045] The pH of the liquid used in the resting cell reaction is, for example, 4.0 to 10.0, preferably 5.0 to 9.0, and more preferably 5.5 to 7.5.

[0046] When the liquid used in the resting cell reaction is a buffer solution, examples of the buffer solution include acetate buffer, phosphate buffer, borate buffer, carbonate buffer, citrate buffer, Tris buffer, and HEPES buffer.

[0047] More specific examples of buffer solutions include KHC8H4O4-NaOH (pH 4.0), CH3COOH-CH3COONa (pH 4.0), MES-NaOH (pH 5.0), CH3COOH-CH3COONa (pH 5.0), KH2PO4-K2HPO4 (pH 6.0), MES-NaOH (pH 6.0), KH2PO4-K2HPO4 (pH 7.0), and PIPES-NaOH (pH 7.0), HEPES-NaOH (pH 8.0), H3BO4-NaOH (pH 8.0), CHES-NaOH (pH 9.0), H3BO4-NaOH (pH 9.0), H2CO3-NaHCO3 (pH 10.0), CHES-NaOH (pH 10.0), etc. are included, and preferred are CH3COOH-CH3COONa, KH2PO4-K2HPO4, H3BO4-NaOH, and more preferred are KH2PO4-K2HPO4.

[0048] When the liquid used in the resting cell reaction is an organic solvent, examples of the organic solvent include aromatic compounds such as benzene and benzonitrile, ketones such as acetone, acetylacetone and methyl ethyl ketone, fatty acid esters such as ethyl acetate, butyl acetate, ethyl butyrate and ethyl formate, ethers such as diethyl ether, dipropyl ether, dibutyl ether, tetrahydrofuran and 1,4-dioxane, halogenated hydrocarbons such as dichloromethane, chloroform and dichloroethane, 1,2-propanediol ... Examples of the diols include butanediol, 1,3-butanediol, 1,4-butanediol, 2,3-butanediol, 1,2-hexanediol, 1,6-hexanediol, 1,2-pentanediol, 1,5-pentanediol, 2-methyl-2,4-pentanediol, and 3-methyl-1,5-pentanediol; alcohols having a linear or branched alkyl group having 1 to 7 carbon atoms; and alcohols such as cyclohexanol, 3-methoxy-3-methyl-1-butanol, and 3-methoxy-1-butanol.

[0049] Of the above liquids, preferred examples of the liquid used in the resting cell reaction include water and buffer solutions.

[0050] The reaction temperature in the resting cell reaction is, for example, 21 to 37° C., preferably 24 to 28° C. The reaction time is, for example, 0.1 to 24 hours, preferably 6 to 12 hours.

[0051] The resting cell reaction can be carried out by suspending lactic acid bacteria belonging to the genus Fructobacillus in the above liquid and leaving the suspension to stand, stirring or shaking.

[0052] Alternatively, the resting cell reaction may be carried out without using the above liquid by adjusting the pH of a culture solution containing lactic acid bacteria belonging to the genus Fructobacillus to a range of 4.0 to 10.0, preferably 5.0 to 9.0, and more preferably 5.5 to 7.5.

[0053] In the drying step, the cells can be dried using drying methods such as freeze drying, tray drying, and spray drying.

[0054] In the cell disruption step, the dried cells can be disrupted by any method. In the cell disruption step, some of the cell components may be removed from the disrupted cells.

[0055] [1-2-2. Lactic acid bacteria culture] When the active ingredient is in the form of a culture of lactic acid bacteria, the active ingredient contains, together with the lactic acid bacteria, medium components containing the product of the lactic acid bacteria. The lactic acid bacteria culture may be a culture produced using one of the above-mentioned specified lactic acid bacteria species alone, a culture produced using a combination of multiple species, or a mixture of a culture produced using one species alone and a culture produced using another species alone.

[0056] The medium components contained in the culture may be those from which some of the medium components used in culturing the lactic acid bacteria have been removed. The lactic acid bacteria contained in the culture may be either live or dead. In the case of dead bacteria, the cells may be disrupted. In the case of disrupted bacteria, some of the cell components may be removed.

[0057] A culture of lactic acid bacteria can be obtained by a production method including a step of culturing one or more of the above-mentioned specific lactic acid bacteria. The production method may include other steps in addition to the steps described above. These other steps may include a step of removing a portion of the medium components, a step of subjecting the lactic acid bacteria to a resting cell reaction, and / or a drying step. Details of each step are as described above in "1-2-1. Lactic Acid Bacteria."

[0058] [1-2-3. Lactic acid bacteria culture supernatant] When the active ingredient is in the form of a culture supernatant of lactic acid bacteria, the active ingredient comprises a medium component containing a product of the lactic acid bacteria. The culture supernatant of lactic acid bacteria may be a culture supernatant produced using a single species of the above-mentioned specified lactic acid bacteria, a culture supernatant produced using a combination of multiple species, or a mixture of a culture supernatant produced using a single species and a culture supernatant produced using another single species.

[0059] The medium components contained in the culture supernatant may be those from which some of the medium components used in culturing the lactic acid bacteria have been removed.

[0060] The culture supernatant of lactic acid bacteria can be obtained by a production method including the steps of culturing one or more of the above-mentioned specific lactic acid bacteria and separating and recovering the culture supernatant. The production method may include other steps in addition to the steps described above. These other steps may include removing a portion of the medium components, subjecting the lactic acid bacteria to a resting cell reaction, and / or drying. Details of each step are as described above in "1-2-1. Lactic Acid Bacteria."

[0061] [1-2-4. Lactic acid bacteria extract] When the active ingredient is in the form of a lactic acid bacterium extract, the active ingredient is a multi-component composition obtained by subjecting the above-mentioned lactic acid bacterium or a culture of lactic acid bacteria to an extraction treatment. Examples of extraction solvents used in preparing the extract include polar solvents such as water; monohydric lower alcohols having 1 to 4 carbon atoms, such as ethanol and isopropanol; polyhydric alcohols, such as 1,3-butylene glycol, propylene glycol, and glycerin; and mixtures thereof. The temperature conditions during extraction are preferably room temperature (e.g., 5 to 35°C, preferably 20 to 30°C). Specific methods for preparing the extract include preparing a suspension containing the above-mentioned lactic acid bacterium or a culture of lactic acid bacteria or a pulverized product thereof in an extraction solvent, and subjecting the suspension to solid-liquid separation to obtain an extract (liquid extract), or optionally drying the extract to obtain a powder (powdered extract).

[0062] [1-3. Ingredients contained in the active ingredients] The active ingredients used in the present invention, lactic acid bacteria belonging to the genus Fructobacillus, cultures of the lactic acid bacteria, culture supernatants of the lactic acid bacteria, and extracts of the lactic acid bacteria, all contain not only NMN and / or NAD produced by the lactic acid bacteria, but also other metabolites (other than NMN and NAD) produced by the lactic acid bacteria and / or bacterial components, etc. It is believed that the effect of improving skin barrier function (skin barrier function improvement) and the effect of improving skin condition (skin condition improvement effect) according to the present invention are achieved by the combination of these components.

[0063] When the active ingredient used in the present invention contains NAD and NMN, the content of NMN is not particularly limited, but the content (content ratio) of NMN per 1 part by weight of NAD is, for example, 0.001 to 10 parts by weight. From the viewpoint of more effectively improving the effect of improving skin barrier function (skin barrier function improvement) and the effect of improving skin condition (skin condition improvement effect), the content of NMN per 1 part by weight of NAD is preferably 0.005 to 5.0 parts by weight, more preferably 0.005 to 3.0 parts by weight, even more preferably 0.008 to 2.0 parts by weight, particularly preferably 0.01 to 1.5 parts by weight, and even more preferably 0.03 to 1.0 parts by weight.

[0064] The content (content ratio) of NMN per mole of NAD is not particularly limited, but may be, for example, 0.001 to 10 moles, preferably 0.005 to 5 moles. In a more preferred embodiment, the content of NMN per mole of NAD is preferably 0.01 to 3 moles, more preferably 0.02 to 2 moles, and even more preferably 0.03 to 1.5 moles.

[0065] [1-4. Content of active ingredients] The amount of active ingredient contained in each of the skin barrier function improving agent and skin condition improving agent of the present invention is not particularly limited, and can be determined appropriately depending on the effect of improving the skin barrier function (skin barrier function improvement) or the effect of improving the skin condition (skin condition improving effect) to be imparted.

[0066] The content of the active ingredient contained in each of the skin barrier function improving agent and the skin condition improving agent of the present invention is, for example, 0.00001 to 10% by weight, preferably 0.0001 to 1% by weight, calculated as the dry weight of lactic acid bacteria belonging to the genus Fructobacillus. The dry weight equivalent refers to the dry weight of the active ingredient when the active ingredient is a lactic acid bacterium or a culture of lactic acid bacteria, the dry weight of the lactic acid bacteria used to obtain the culture supernatant when the active ingredient is a culture supernatant of lactic acid bacteria, and the dry weight of the lactic acid bacteria used to obtain the extract when the active ingredient is an extract.

[0067] [2. Other ingredients] The skin barrier function improving agent and skin condition improving agent of the present invention may or may not contain other pharmacological ingredients in addition to the above-mentioned active ingredients, as necessary. Examples of such pharmacological ingredients include anti-inflammatory agents, antioxidants, disinfectants, cooling agents, vitamins, mucopolysaccharides, etc.

[0068] Furthermore, the skin barrier function improving agent and skin condition improving agent of the present invention may or may not contain bases and / or additives as necessary to form the desired formulation. Such bases and / or additives are not particularly limited as long as they are pharmaceutically acceptable, and examples thereof include aqueous bases such as water and monohydric lower alcohols having 1 to 4 carbon atoms (ethanol, isopropanol, etc.); oily bases such as naturally derived oils (vegetable oils, animal oils, and processed oils thereof), mineral oils, ester oils, fatty acid alkyl esters, fatty acids, fatty acid esters, and higher alcohols; surfactants; polyhydric alcohols (glycerin, propylene glycol, dipropylene glycol, 1,3-butylene glycol, etc.); and additives such as cooling agents, preservatives, flavoring agents, colorants, thickeners, pH adjusters, humectants, stabilizers, antioxidants, UV absorbers, chelating agents, adhesives, buffers, solubilizers, solubilizers, and preservatives.

[0069] [3. Formulation and product classification] The formulation form of the skin barrier function improving agent and skin condition improving agent of the present invention is not particularly limited, and may be any of liquid, semi-solid (cream, gel, ointment, paste), solid (granules, fine granules, powder, tablets, capsules), etc. Furthermore, the skin barrier function improving agent and skin condition improving agent of the present invention may be a non-emulsified formulation such as an aqueous formulation or an oily formulation, or may be an emulsified formulation such as an oil-in-water emulsion formulation or a water-in-oil emulsion formulation.

[0070] Furthermore, product categories of the skin barrier function improving agent and skin condition improving agent of the present invention include foods and beverages, cosmetics, and pharmaceuticals.

[0071] Food and beverages include, but are not limited to, fermented milk (drinkable yogurt, etc.), lactic acid bacteria drinks, dairy drinks (coffee milk, fruit milk, etc.), tea drinks (green tea, black tea, oolong tea, etc.), fruit and vegetable drinks (drinks containing fruit juices such as orange, apple, grape, etc., and vegetable juices such as tomato, carrot, etc.), alcoholic drinks (beer, happoshu, wine, etc.), carbonated drinks, soft drinks, water-based drinks, and other beverages; and processed foods such as fermented milk (set yogurt, soft yogurt, etc.), sweets, instant foods, and seasonings.

[0072] Foods and beverages also include functional foods. Functional foods refer to foods that have a certain functionality for the living body, and examples include functional health foods such as foods for specified health uses (including conditionally designated foods for specified health uses [FOSHU]) and foods with nutrient function claims, foods with functional claims, foods for special dietary uses, dietary supplements, health supplements, supplements (e.g., tablets, coated tablets, sugar-coated tablets, capsules, liquids, and other dosage forms), and beauty foods (e.g., diet foods). Furthermore, functional foods may be foods for special dietary uses, such as foods for the sick, powdered milk for pregnant and nursing women, infant formula, foods for the elderly, and foods for caregivers.

[0073] Cosmetics include topical preparations including topical preparations for the skin and topical preparations for the mucosa, and more specifically include basic cosmetics such as lotions, milky lotions, creams, essences, gels, packs, sheet masks, and lip balms; skin cleansers such as facial cleansers, makeup removers (including cleansing agents), exfoliants, and body shampoos; body care cosmetics such as sunscreens, body gels, body massage agents, antiperspirants, deodorants, hair removers, and bath additives; makeup cosmetics such as foundations, powders, lipsticks, blushers, eye shadows, eyeliners, mascaras, and eyebrow pencils; nail cosmetics such as nail polish and nail removers; hair cosmetics such as hair styling agents, shampoos, conditioners, rinses, and hair growth agents; and oral cosmetics such as liquid toothpastes, toothpastes, mouthwashes, and mouth sprays.

[0074] Pharmaceuticals include oral preparations and topical preparations (including quasi-drugs) including topical preparations for skin and mucous membranes. Oral preparations include tablets, coated tablets, sugar-coated tablets, capsules, and liquid preparations, while topical preparations include liquid preparations (including lotions, sprays, aerosols, and emulsions), foams, ointments, creams, gels, and patches.

[0075] Among these product categories, oral preparations and foods and beverages are preferred from the viewpoint of further enhancing the effect of improving the skin barrier function (skin barrier function improvement) or the effect of improving the skin condition (skin condition improvement effect).

[0076] [4.Application] The skin barrier function improving agent and skin condition improving agent of the present invention are used for purposes aimed at improving the skin barrier function (skin barrier function improvement) or improving the skin condition (skin condition improving effect), respectively.

[0077] Uses of skin barrier function improvers include improving dry skin, suppressing transepidermal water loss, and suppressing skin thickening. Of these, suppressing transepidermal water loss is preferred. Therefore, by ingesting a skin barrier function improver, it is expected to improve the dry condition of the skin (improving dry skin), prevent dry skin (preventing dry skin), and enhance or improve the stratum corneum barrier function of the skin. Along with these actions, it is also expected to prevent or improve rough skin caused by dryness. Improving dry skin means, for example, preventing the skin from becoming dry for a long time by preventing excess sebum secreted to replenish moisture from clogging the pores and preventing the formation of a sebum film on the skin surface, thereby preventing further progression of dryness. Suppression of transepidermal water loss means, for example, the effect of suppressing the amount of water loss from the skin surface, and includes the effect of moisturizing the skin or retaining moisture. Inhibition of skin thickening means, for example, inhibiting an increase in the thickness of the stratum corneum in the epidermis and / or dermis and / or subcutaneous layer of the skin (e.g., subcutaneous fat or muscle) and promoting skin cell turnover.

[0078] The skin condition improving agent can be used for improving or inhibiting a decrease in skin viscoelasticity, improving or inhibiting skin wrinkles, improving or inhibiting skin sagging, and improving skin firmness and elasticity. Of these, improving or inhibiting skin viscoelasticity or improving or inhibiting skin wrinkles is preferred. Improving or inhibiting a decrease in skin viscoelasticity means, for example, improving skin viscoelasticity or improving skin flexibility or inhibiting a decrease in skin viscoelasticity. The target skin is not particularly limited, and examples thereof include the face (e.g., forehead, cheeks, mouth, corners of the eyes, etc.), arms, elbows, backs of hands, fingertips, feet, knees, shins, heels, neck, armpits, back, etc. Improving or inhibiting skin wrinkles means, for example, inhibiting an increase in skin wrinkles or promoting a decrease in wrinkles, thereby preventing apparent skin aging. The improvement or suppression of skin sagging means, for example, restoring skin firmness that has been lost due to aging or the like, and improving or suppressing skin sagging that has occurred due to aging or the like. Improving skin firmness and elasticity means, for example, improving the firmness of skin that has lost its firmness due to thickening of subcutaneous fat or cellulite, suppressing skin aging that occurs with age, and improving skin elasticity and spots caused by ultraviolet rays and active oxygen.

[0079] [5.Dose] When the skin barrier function improving agent and skin condition improving agent of the present invention are administered internally, the amount of the active ingredient can be, for example, 0.001 to 1000 mg / kg / day, preferably 0.01 to 500 mg / kg / day, and can be administered internally 1 to 5 times a day. [Example]

[0080] The present invention will be explained in more detail below by showing examples, but the present invention is not limited to these examples.

[0081] [Preparation of dried bacterial powder (killed bacteria) of OS-1010 strain] Fructobacillus fructosus OS-1010 strain was inoculated into 30 ml of Difco MRS medium (preculture medium) and cultured statically at 30°C for 24 hours. The resulting culture was inoculated into 1 L of MRS medium (main culture medium) to an OD660 of 0.02 and cultured with stirring at 30°C for 12 hours at pH 6.0-7.0. The resulting culture was centrifuged to collect the bacterial cells. The collected cells were washed with 1 L of 0.85 w / w% KCl aqueous solution. The washed cells were centrifuged again to collect the bacterial cells. The collected bacterial cells were suspended in 50 ml of distilled water, sterilized at 65°C for 30 minutes, and lyophilized to obtain a dried powder of OS-1010 strain cells (killed cells). Tablets were prepared using the dried powder of OS-1010 strain cells (killed cells) and used for evaluation testing.

[0082] Using the HPLC analysis conditions described below, the amounts of nicotinamide mononucleotide (NMN) and nicotinamide adenine dinucleotide (NAD) produced in the recovered lactic acid bacteria powder were measured. The ratio of NMN per part by weight of NAD was 0.03 parts by weight (0.06 moles of NMN per mole of NAD).

[0083] (MRS medium composition) 2w / w% glucose 1w / w% protease peptone 1w / w% beef extract 0.5w / w% yeast extract 0.2w / w% ammonium citrate 0.1w / w% Tween 80 0.5w / w% sodium acetate 0.01w / w% magnesium sulfate 0.005w / w% manganese sulfate 0.2w / w% dipotassium hydrogen phosphate

[0084] (HPLC analysis conditions) How to measure NMN Column: DaisoPak SP-100-5-ODS-P (4.6 x 150 mm) x 2 Column temperature: 25℃ Eluent: 75mM ammonium phosphate aqueous solution (pH 6.0) Flow rate: 0.6ml / min Detector: UV detector (260 nm) Detection time: 14.5 minutes How to measure NAD Column: DAISOPAK SP-100-5-ODS-P (4.6 x 250 mm) x 2 Column temperature: 25℃ Eluent: 75 mM ammonium phosphate aqueous solution (pH 6.0):methanol = 95:5 (w:w) Flow rate: 0.7mL / min Detector: UV detector (260 nm) Detection time: 33 minutes

[0085] (Method for quantifying NMN and NAD) A calibration curve was created using purchased NMN and NAD as standard samples, and the amounts of NMN and NAD produced in the OS-1010 strain dried cell powder (killed cells) were quantified from the peak area ratio of their HLPC.

[0086] A human study was conducted to examine the effect of taking tablets containing dried powder of Fructobacillus fructosus strain OS-1010 (killed bacteria) on improving skin conditions.

[0087] (1) Test Contents This study was conducted on 60 healthy men and women aged 30 to 59. The subjects were randomly divided into the following two groups. (Tablet a) Placebo group: A group taking placebo tablets (i.e., tablets that do not contain dried bacterial powder (killed bacteria) of Fructobacillus fructosus OS-1010 strain). (Tablet b) OS-1010 strain intake group (lactic acid bacteria inoculation group): A group taking tablets containing dried bacterial powder (killed bacteria) of Fructobacillus fructosus OS-1010 strain (lactic acid bacteria content: 12 billion per tablet)

[0088] The compositions of the tablets used in the test are as follows: The placebo group received tablet A, and the lactic acid bacteria group received tablet B. Tablet B contained 12 billion killed cells of Fructobacillus fructosus OS-1010 strain per tablet. In both the placebo and lactic acid bacteria inoculation groups, subjects took one tablet per day for eight weeks. Transepidermal water loss, skin viscoelasticity, wrinkle area ratio, total mean wrinkle depth, and maximum mean wrinkle depth were measured before and eight weeks after ingestion of the tablets.

[0089] Tablet a (placebo group) composition Dextrin: 132mg Powdered cellulose: 16 mg Fine silicon dioxide: 4 mg Sucrose fatty acid esters: 4mg Maltose; 24mg Lactose; 20mg Total (1 tablet): 200mg

[0090] Tablet B (for lactic acid bacteria inoculation group) composition Dextrin: 129mg Powdered cellulose: 16 mg Fine silicon dioxide: 4 mg Sucrose fatty acid esters: 4mg Maltose; 24mg Lactose; 20mg Dried powder of Fructobacillus fructosus strain OS-1010 (killed cells); 3mg Total (1 tablet): 200mg

[0091] (2) Example 1: Test on transepidermal water loss Using a Tewameter (Integral Corporation), measurements were taken five times on the arm (inside the forearm). Transepidermal water loss was measured for 30 subjects before and eight weeks after intake, and the average value for each group was calculated. Table 1 shows the values, with the average value before intake set at 100. The lower the value, the greater the effect.

[0092] [Table 1]

[0093] Compared to the placebo group, the increase in transepidermal water loss was suppressed in the lactic acid bacteria intake group, and it was confirmed that the lactic acid bacteria intake has the effect of improving the skin's barrier function, such as improving dry skin (improving dry skin) and preventing dry skin (preventing dry skin).

[0094] (3) Example 2: Test on skin viscoelasticity Measurements were taken five times on the cheek using a Cutometer (Integral Corporation). The viscoelasticity (R0: flexibility, R2: total elasticity, R5: net elasticity, R7: recovery elasticity) of 30 subjects was measured before and 8 weeks after intake, and the average value for each group was calculated. Table 2 shows the values, with the average value before intake set at 100. The higher the value, the greater the effect.

[0095] [Table 2]

[0096] Compared to the placebo group, the lactic acid bacteria intake group had improved R0 (flexibility) and R7 (recovery elasticity), confirming that intake of lactic acid bacteria strains has the effect of improving skin elasticity and skin firmness.

[0097] (4) Example 3 Test on Skin Viscoelasticity (Subgroup Analysis) A subgroup analysis was further conducted on the results of Example 2, focusing on subjects whose skin sebum levels before initiating intake were equal to or greater than the median. Based on the average values ​​of viscoelasticity (R0: flexibility, R2: total elasticity, R5: net elasticity, R7: recovery elasticity) for the subjects (placebo group: 17 subjects, lactic acid bacteria intake group: 15 subjects) before intake and 8 weeks after intake, the values ​​are shown in Table 3, with the average value before intake set to 100. The higher the value, the greater the effect.

[0098] [Table 3]

[0099] Compared to the placebo group, the lactic acid bacteria intake group had improved R2 (total elasticity), R5 (net elasticity), and R7 (recovery elasticity), confirming that intake of lactic acid bacteria strains has the effect of improving skin elasticity and skin firmness.

[0100] (5) Example 4: Test on crow's feet Measurements were taken at the outer corners of the eyes using PRIMOS (Integral Corporation). Before and eight weeks after ingestion, the wrinkle area ratio, total average wrinkle depth, and maximum average wrinkle depth were measured for the wrinkles around the eyes of 30 subjects. Table 4 shows the values, with the average value before ingestion set to 100. The smaller the value, the greater the effect.

[0101] [Table 4]

[0102] Compared to the placebo group, the lactic acid bacteria intake group showed improvements in all values, including wrinkle area ratio, total average wrinkle depth, and maximum average wrinkle depth, confirming that intake of the OS-1010 strain has the effect of improving skin wrinkles.

[0103] (5) Example 5: Visual Analogue Scale Questionnaire The following test was conducted on healthy men and women aged 20 to 64 years old. Each subject took two tablets of Tablet C shown below per day for 12 weeks. Before taking the tablets, and 4, 8, and 12 weeks after taking them, subjects were asked to fill out a visual analogue scale (hereinafter referred to as VAS) questionnaire about their skin condition.

[0104] Tablet c (OS-1010 strain combination) composition Dextrin: 12mg Powdered cellulose: 24 mg Fine silicon dioxide: 4 mg Sucrose fatty acid esters: 4mg Maltose; 40mg Lactose; 16mg Dried powder of Fructobacillus fructosus OS-1010 (killed cells); 100mg Total (1 tablet): 200mg

[0105] The VAS questionnaire consisted of seven items: "skin condition," "skin moistness (moisture)," "skin texture," "fine lines around the eyes," "skin radiance," "skin clarity," and "skin firmness and elasticity." The VAS questionnaire measures the distance (mm) from the left end of a 100mm line where the subject marks their condition, with the left end representing the best condition (no symptoms) and the right end representing the worst condition (worst symptoms ever).The smaller the number, the better the evaluation. Figure 1 shows the change in VAS questionnaire scores. In a VAS questionnaire on physical sensation, the score was lower than before taking the lactic acid bacteria, confirming that taking the lactic acid bacteria improved the physical sensation of skin condition.

Claims

1. A skin dryness alleviating agent containing lactic acid bacteria Fructobacillus fructosus OS-1010 strain (accession number: NITE BP-03818).

2. A transepidermal water loss inhibitor containing Fructobacillus fructosus OS-1010 strain (accession number: NITE BP-03818) lactic acid bacteria.

3. A skin thickening inhibitor containing Fructobacillus fructosus OS-1010 strain (accession number: NITE BP-03818) lactic acid bacteria.

4. An agent for improving or inhibiting a decrease in skin viscoelasticity, comprising Fructobacillus fructosus OS-1010 strain (accession number: NITE BP-03818) lactic acid bacteria.

5. A skin wrinkle improving or inhibiting agent containing lactic acid bacteria Fructobacillus fructosus OS-1010 strain (accession number: NITE BP-03818).

6. An agent for improving or inhibiting sagging skin, comprising lactic acid bacteria Fructobacillus fructosus OS-1010 strain (accession number: NITE BP-03818).

7. A skin firmness and elasticity improving agent containing Fructobacillus fructosus OS-1010 strain (accession number: NITE BP-03818) lactic acid bacteria.

8. A food or drink comprising any one of the agents according to claims 1 to 7.

9. A cosmetic comprising any one of the agents according to claims 1 to 7.

10. A pharmaceutical comprising any one of the agents according to claims 1 to 7.

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