WO2022168939A1 - Ceramide synthase gene expression promoting agent, skin improving agent, functional food for skin improvement, cosmetic, and usage method for dipeptide - Google Patents

Ceramide synthase gene expression promoting agent, skin improving agent, functional food for skin improvement, cosmetic, and usage method for dipeptide Download PDF

Info

Publication number
WO2022168939A1
WO2022168939A1 PCT/JP2022/004425 JP2022004425W WO2022168939A1 WO 2022168939 A1 WO2022168939 A1 WO 2022168939A1 JP 2022004425 W JP2022004425 W JP 2022004425W WO 2022168939 A1 WO2022168939 A1 WO 2022168939A1
Authority
WO
WIPO (PCT)
Prior art keywords
gly
ile
skin
ceramide synthase
dipeptide
Prior art date
Application number
PCT/JP2022/004425
Other languages
French (fr)
Japanese (ja)
Inventor
邦義 清水
充寛 寺本
Original Assignee
国立大学法人九州大学
佳秀工業株式会社
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 国立大学法人九州大学, 佳秀工業株式会社 filed Critical 国立大学法人九州大学
Publication of WO2022168939A1 publication Critical patent/WO2022168939A1/en

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/17—Amino acids, peptides or proteins
    • A23L33/18—Peptides; Protein hydrolysates
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • A61K38/05—Dipeptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/64—Proteins; Peptides; Derivatives or degradation products thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/06—Dipeptides
    • C07K5/06008—Dipeptides with the first amino acid being neutral
    • C07K5/06017—Dipeptides with the first amino acid being neutral and aliphatic

Definitions

  • the present invention relates to a method for using a ceramide synthase gene expression promoter, a skin quality improving agent, a functional food and cosmetic for improving skin quality, and a dipeptide.
  • a beautiful and healthy skin is one of the important factors that bring out the attractiveness of a person's appearance. Therefore, skin care has been conventionally performed by both men and women.
  • ingredients responsible for moisturizing the skin are important, and examples of such moisturizing ingredients include ceramide.
  • Ceramide is a type of sphingolipid that is biosynthesized in cells, and is the main component of the stratum corneum that forms the surface of the epidermal layer of human skin. In addition, ceramide suppresses excessive evaporation of moisture and also acts as a barrier against microorganisms. need to keep
  • ceramide synthesis promoters containing fungal cultures containing nicotinic acid and/or nicotinamide as active ingredients have been proposed.
  • ceramide synthesis accelerator improvement of rough skin and improvement of various skin diseases can be expected by activating ceramide synthesis of the epidermal cells themselves within the skin surface layer and improving the skin barrier function.
  • the present invention has been made in view of such circumstances, and a ceramide synthase gene capable of increasing the expression level of the ceramide synthase gene by employing a predetermined dipeptide as a main component for inducing function.
  • An expression promoter is provided.
  • the present invention also provides a skin quality improving agent, a functional food for improving skin quality, cosmetics, and a method of using the same dipeptide containing a predetermined dipeptide having a ceramide synthase gene expression promoting function.
  • the ceramide synthase gene expression promoter according to the present invention contains a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function. did.
  • the ceramide synthase gene expression promoter according to the present invention is also characterized in that the ceramide synthase gene is the ceramide synthase 3 (CERS3) gene.
  • CERS3 ceramide synthase 3
  • the skin quality-improving agent, functional food for skin quality improvement, and cosmetics according to the present invention contain a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function.
  • a dipeptide represented by Gly-Ile and/or Ile-Gly is used as a main ingredient for inducing the function of a ceramide synthase gene expression promoter or as a participating ingredient for the purpose of improving skin quality. It was decided to.
  • the dipeptide represented by Gly-Ile and/or Ile-Gly is contained as a main component for inducing function. It is possible to provide a ceramide synthase gene expression promoter that employs a predetermined dipeptide as a component and is capable of increasing the expression level of the ceramide synthase gene.
  • the ceramide synthase gene is the ceramide synthase 3 (CERS3) gene, it is possible to target ceramide synthase 3, which is mainly expressed in skin and keratinocytes, and to achieve more efficient ceramide synthase 3. It can be used as a ceramide synthase gene expression promoter.
  • CERS3 ceramide synthase 3
  • the skin quality improving agent, skin quality improving functional food, and cosmetics according to the present invention contain a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function. , a skin quality improving agent, a functional food for improving skin quality, and a cosmetic that can increase the expression level of the ceramide synthase gene.
  • a dipeptide represented by Gly-Ile and/or Ile-Gly is used as a main ingredient for inducing the function of the ceramide synthase gene expression promoter or as a participating ingredient for the purpose of improving skin quality. Since it was decided to use it, it is possible to enjoy the effect derived from the increase in the expression level of the ceramide synthase gene by the dipeptide represented by Gly-Ile and/or Ile-Gly.
  • FIG. 4 is an explanatory diagram showing the cell viability of HaCaT cells by addition of each dipeptide.
  • FIG. 4 is an explanatory diagram showing changes in the expression of ceramide synthase gene in HaCaT cells due to the addition of each dipeptide.
  • FIG. 4 is an explanatory diagram showing changes in the expression of the ceramide synthase gene in HaCaT cells due to the addition of a small amount of dipeptide.
  • FIG. 3 is an explanatory diagram showing the test results regarding the amount of water when the test food was ingested.
  • FIG. 3 is an explanatory diagram showing the test results regarding the amount of transepidermal water loss when the test food was ingested.
  • FIG. 3 is an explanatory diagram showing test results regarding total elasticity R2 of skin when test foods were ingested.
  • FIG. 3 is an explanatory diagram showing test results regarding skin viscoelasticity (R5 (net elasticity)) when test foods were ingested.
  • FIG. 3 is an explanatory diagram showing test results regarding skin recovery power R7 when a test food is ingested.
  • the present invention provides a ceramide synthase gene expression promoter that employs a predetermined dipeptide as a main component for inducing function and is capable of increasing the expression level of the ceramide synthase gene.
  • ceramide synthase gene expression promoter is not particularly limited. It can be used not only as an external agent but also, for example, as a reagent for testing and research.
  • the promoter uses a dipeptide represented by Gly-Ile (glycine-isoleucine) and/or Ile-Gly (isoleucine-glycine) to induce function. contains as a major component of Gly-Ile (glycine-isoleucine) and/or Ile-Gly (isoleucine-glycine) to induce function. contains as a major component of Gly-Ile (glycine-isoleucine) and/or Ile-Gly (isoleucine-glycine) to induce function. contains as a major component of Gly-Ile (glycine-isoleucine) and/or Ile-Gly (isoleucine-glycine) to induce function. contains as a major component of Gly-Ile (glycine-isoleucine) and/or Ile-Gly (isoleucine-glycine) to induce function. contains as a major component of Gly-Ile (glycine-isoleucine) and/or Ile-Gly
  • Gly-Ile and Ile-Gly are dipeptides in which glycine and isoleucine are peptide-bonded.
  • the origin of Gly-Ile and Ile-Gly is not particularly limited. It may be of any origin as long as it does not contradict the purpose. In this specification, unless otherwise specified, the left side of the amino acid sequence is described as the N-terminus.
  • Gly-Leu and Leu-Gly are known as dipeptides that are said to have a positive effect on the skin, but the dipeptide proposed in the present application is a substance that is structurally completely different from these substances, and It is clear from the test results described later that it has a higher ceramide synthase gene expression promoting effect than Gly-Leu or Leu-Gly.
  • the skin quality improving agent is characteristic in that it contains a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function.
  • the skin quality improving agent may be taken orally, or may be used as an external skin preparation.
  • External agents for skin include pharmaceuticals, quasi-drugs, and cosmetics (including medicated cosmetics), but are not bound by these frameworks and are included in this concept as long as they are externally applied to the skin. .
  • the present application also provides a cosmetic, which is one aspect of this external preparation for skin. That is, the cosmetic according to the present embodiment is characteristic in that it contains a dipeptide represented by Gly-Ile and/or Ile-Gly as an active ingredient.
  • cosmetics include not only so-called makeup cosmetics, but also basic cosmetics, hair tonics, perfumes, toothpastes, shampoos, rinses, soaps and cleansers used for washing the body, and toiletry products such as bath agents. It also includes medicinal cosmetics claiming preventive effects.
  • makeup cosmetics include foundations, eyebrows, mascara, eye shadows, eyeliners, lipsticks, glosses, cheeks, face powders, and nail polishes.
  • Basic cosmetics include lotions, milky lotions, Facial cleansers, cleansers, serums, creams and the like can be mentioned. It should be noted that the above descriptions of external preparations for skin, cosmetics, and the like are examples of corresponding articles, etc., in order to facilitate understanding of the present invention, and the interpretation of each term is based on the articles, etc., listed above. It is not limited. However, it does not preclude the applicant from limiting the present invention to these articles and the like in obtaining the right of the present application.
  • the present application also provides a method of using a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing the function of a ceramide synthase gene expression promoter.
  • a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing the function of a ceramide synthase gene expression promoter.
  • the present application also provides a method of using dipeptides represented by Gly-Ile and/or Ile-Gly as participating ingredients in functional foods for the purpose of improving skin quality.
  • dipeptides represented by Gly-Ile and/or Ile-Gly as participating ingredients in functional foods for the purpose of improving skin quality.
  • Several ingredients have been proposed to improve skin quality through oral ingestion. Not proposed.
  • the present application also provides a functional food for improving skin quality, and in particular, provides a functional food for improving skin quality that can exert an excellent moisturizing effect on the skin when taken orally.
  • Functional food is a concept that includes all foods that are said to contribute to the maintenance and promotion of health that do not contain pharmaceutical ingredients. It also includes food with health claims such as food for specified health use, food with nutrient function claims, and food with function claims.
  • the dosage form is not particularly limited, and any dosage form such as tablets, capsules, fine granules, pills, lozenges, liquids, and jelly-like can be selected. can be done.
  • Concentration of Gly-Ile and/or Ile-Gly as a component for inducing function to be contained in the ceramide synthase gene expression promoter, skin quality improving agent, functional food for improving skin quality, cosmetics, etc. according to the present embodiment is not particularly limited as long as the concentration is capable of inducing the above function.
  • the concentration of Gly-Ile or Ile-Gly contained in the functional food for improving skin quality is, for example, an amount that allows 20 to 600 ⁇ g of Gly-Ile or Ile-Gly to be ingested per day, The total amount of Gly-Ile and Ile-Gly can be taken in an amount of 20 to 600 ⁇ g per day.
  • the amount of Gly-Ile and/or Ile-Gly contained in one tablet is 20 to 600 ⁇ g. If two tablets are a guideline, the amount of Gly-Ile and/or Ile-Gly contained in the intake unit amount such as one tablet, one capsule, and one packet may be adjusted appropriately so that one tablet is 10 to 300 ⁇ g. It is possible.
  • the cosmetic may contain 0.03 to 200 ⁇ g/g of Gly-Ile or Ile-Gly, or the total concentration of Gly-Ile and Ile-Gly may be 0.03 to 200 ⁇ g/g.
  • concentrations of Gly-Ile and Ile-Gly in the ceramide synthase gene expression accelerator and the skin quality improving agent are adjusted to concentrations suitable for use in experiments, etc., and suitable as raw materials for the production of foods and cosmetics. concentration.
  • the ceramide synthase gene expression promoter, skin quality improving agent, skin quality improving functional food, cosmetics, etc. may contain Gly-Ile and/or Ile-Gly as a component for inducing function.
  • it may contain excipients, additives, auxiliary ingredients, and the like according to dosage forms, food forms, and the like.
  • excipients examples include lactose, crystalline cellulose, and starch when the dosage form is a solid formulation.
  • additives include stabilizers, surfactants, solubilizers, plasticizers, sweeteners, antioxidants, flavoring agents, coloring agents, preservatives, inorganic fillers, and the like.
  • the skin is composed of the epidermis and dermis, which are divided into the stratum basale, the stratum spinosum, the stratum granulosum and the stratum corneum.
  • a lamellar structure composed of lipids such as ceramides between stratum corneum cells located in the outermost layer of the skin plays a role as a physical barrier to retain water in the body and prevent invasion of bacteria and viruses.
  • Gly-Ile and Ile-Gly (hereinafter also referred to as the dipeptide of the present application) on epidermal cells was evaluated by examining changes in the expression level of the ceramide synthase gene.
  • Gly-Leu and Leu-Gly (hereinafter also referred to as comparative dipeptides) were used as controls for comparison, and differences from the dipeptides of the present application were also examined.
  • HaCaT cells Human epidermal keratinocytes (HaCaT cells) were cultured in ⁇ 10 cm dishes using Dulbecco's Modified Eagle Medium (DMEM) (high glucose) (containing 1% penicillin-streptomycin and 10% fetal bovine serum (FBS)) until confluent. was pre-cultured.
  • DMEM Dulbecco's Modified Eagle Medium
  • FBS fetal bovine serum
  • the cells were washed with phosphate buffered saline (PBS), resuspended in medium, seeded in a 24-well plate at a concentration of 1.0 ⁇ 10 5 cells/well, and placed in a CO 2 incubator (37°C, 5% CO 2 ). was cultured overnight.
  • PBS phosphate buffered saline
  • serum-free DMEM high glucose
  • penicillin-streptomycin containing either Gly-Ile, Ile-Gly, Gly-Leu, or Leu-Gly at a predetermined final concentration
  • cDNA was synthesized from the extracted total RNA using ReverTra Ace qPCR RT Master Mix with gDNA Remover (TOYOBO).
  • TOYOBO ReverTra Ace qPCR RT Master Mix with gDNA Remover
  • 5'-CCGATTACCTGCTGGAGTCAG-3' SEQ ID NO: 1
  • 5'-GGCGAAGACGATGAAGATGTTG-3' SEQ ID NO: 2
  • FIG. 1 shows the cell viability of HaCaT cells with the addition of each dipeptide.
  • FIG. 2 shows changes in the expression of the ceramide synthase gene in HaCaT cells with the addition of the dipeptides of the present application, Ile-Gly and Gly-Ile, and the comparative dipeptides, Leu-Gly and Gly-Leu.
  • FIG. 2(a) shows the expression level of the ceramide synthase CERS2 gene relative to the control, similarly FIG. 2(b) shows the expression level of the ceramide synthase CERS3 gene, and FIG. is shown as a relative value.
  • the ceramide synthase CERS2 gene is highly expressed in many cell tissues such as kidney, liver and intestine.
  • the ceramide synthase CERS3 gene is mainly expressed in skin and keratinocytes.
  • the ceramide synthase CERS4 gene is expressed not only in skin but also in leukocytes, heart and liver.
  • the addition of Gly-Ile showed a significant concentration-dependent increase in the expression level, and it is extremely interesting that at a concentration of 200 ⁇ g/mL, the expression level increased 3-fold compared to the control.
  • ceramide synthase CERS4 gene As shown in Fig. 2(c), a marked increase in the expression level was confirmed with the addition of Gly-Ile.
  • the dipeptide of the present application can be expected to have a cell activating effect, and that it has the effect of enhancing the expression of the ceramide synthase CERS3 and CERS4 genes. It was also suggested that this effect is particularly high with Gly-Ile among the dipeptides of the present application.
  • the dipeptide of the present application is found to have the effect of promoting the expression of the ceramide synthase gene, and is expected to have the effect of promoting ceramide synthesis, which plays an important role in the skin barrier function.
  • the dipeptide represented by Gly-Ile and/or Ile-Gly is a main component for inducing function in skin quality improving agents, a component involved in improving skin quality in functional foods, and a cosmetic component. It was shown that it can be used as an active ingredient for improving skin quality in cosmetics.
  • sample solution S1 containing at least 0.00038 ⁇ g/mL of Gly-Ile and at least 0.00097 ⁇ g/mL of Ile-Gly with a total concentration of both dipeptides of 0.00135 ⁇ g/mL and of Gly-Ile and at least 0.00195 ⁇ g/mL of Ile-Gly, with a total concentration of both dipeptides of 0.00271 ⁇ g/mL (double concentration solution of sample solution S1) and at least 0.00152 ⁇ g/mL of Ile-Gly.
  • Sample solution S3 containing 1 mL of Gly-Ile and at least 0.0039 ⁇ g/mL of Ile-Gly and having a total concentration of 0.00542 ⁇ g/mL of both dipeptides was used as the test solution.
  • 1 mL of final serum-free DMEM (high glucose) (containing 1% penicillin-streptomycin) constituting any of the above test solutions was added to each well, and placed in a CO 2 incubator (37°C). , 5% CO 2 ) for 48 hours.
  • Fig. 3(a) shows the cell viability of HaCaT cells with the addition of each sample solution S1 to S3. As shown in FIG. 3(a), no significant difference was observed between the sample solutions S1 to S3 and the control.
  • Fig. 3(b) shows changes in the expression of the ceramide synthase gene in HaCaT cells after the addition of each sample solution S1 to S3.
  • the CERS3 and CERS4 genes which have been reported to be expressed in the skin, were upregulated by about 2 to 4 times compared to controls.
  • the CERS3 gene showed a more pronounced increase in expression compared to the CERS4 gene.
  • Gly-Ile and Ile-Gly which are the dipeptides of the present application, have no cytotoxicity and have the effect of enhancing the expression of the ceramide synthase CERS3 and CERS4 genes. Therefore, Gly-Ile and Ile-Gly, which are the dipeptides of the present application, can be expected to promote ceramide synthesis, which plays an important role in skin barrier function, at a total concentration of 0.00135 ⁇ g/mL or higher.
  • the total concentration of both dipeptides is 0.03 ⁇ g/mL. can be effective.
  • the dosage form of the functional food for testing was a hard capsule. Specifically, 200 mg of powdered dextrin is used as a base material, Gly-Ile and/or Ile-Gly obtained by peptide synthesis are added and mixed, and the obtained mixed powder is used as a capsule with gelatin as a shell material. The body was filled and capped to obtain a functional food A in the form of a hard capsule.
  • Table 4 shows the dipeptide content of functional food A using a calibration curve. It was confirmed that one grain of functional food A contained 26.25 ⁇ 0.38 ⁇ g of Gly-Ile and 22.57 ⁇ 0.48 ⁇ g of Ile-Gly.
  • test was conducted on 20 healthy general women aged 40 to 50, divided into 2 groups: 10 test product groups and 10 placebo groups.
  • the intake was 2 tablets per day for both the test product group (functional food A) and the placebo group (placebo food).
  • the placebo food does not contain Gly-Ile and Ile-Gly.
  • the ingestion period was 4 weeks.
  • the skin quality was measured after the ingestion period. Specifically, the water content, water loss, and viscoelasticity of the skin were measured. The measurement site was the left arm.
  • the standard error was calculated, and a paired t-test and an unpaired t-test were performed for comparison between pre-measurement and post-measurement (within-group comparison) and between-group comparisons.
  • the significance level was set to 5%, and less than 5% was judged to have a significant difference (p ⁇ 0.05:**), and less than 10% was judged to have a significant tendency (p ⁇ 0.1:*).
  • FIG. 4 shows test results regarding water content.
  • the water content is the amount of water contained in the stratum corneum, and is a value referred to as an index of skin moisture.
  • FIG. 4(a) shows changes in water content measurements before and after ingestion of the test food or placebo food
  • FIG. 4(b) shows changes in water content in the placebo group and the test product group, respectively. .
  • Fig. 5 shows the test results regarding transepidermal water loss.
  • the transepidermal water loss is the amount of water that evaporates from the surface of the skin, and is an indicator of the barrier function.
  • Figure 5 (a) shows the change in transepidermal water loss before and after ingestion of the test food or placebo food
  • Figure 5 (b) shows the change in the amount of transepidermal water loss between the placebo group and the test product group. each shown.
  • FIG. 6 shows the test results regarding the total skin elasticity (R2).
  • Total elasticity is an index representing the overall elasticity of the skin. Skin elasticity is maintained by the fiber and water-retaining components of the dermis layer.
  • FIG. 6(a) shows changes in total elasticity before and after ingestion of the test food or placebo food
  • FIG. 6(b) shows changes in total elasticity of the placebo group and the test article group, respectively.
  • FIG. 7 shows the test results regarding skin viscoelasticity (R5 (net elasticity)). Viscoelasticity is a value that serves as an index of skin firmness and elasticity.
  • FIG. 7(a) shows changes in viscoelasticity before and after ingestion of the test food or placebo food
  • FIG. 7(b) shows changes in viscoelasticity between the placebo group and the test product group, respectively.
  • Fig. 8 shows test results regarding skin recovery power R7 (return rate).
  • the resilience is a value representing the instantaneous contraction rate with respect to the stretched length of the skin at the time of suction, and shows a high correlation with firmness and sagging of the skin.
  • FIG. 8(a) shows changes in resilience before and after ingestion of the test food or placebo food
  • FIG. 8(b) shows changes in resilience between the placebo group and the test article group, respectively.
  • the ceramide synthase gene expression promoter contains a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function. It is possible to provide a ceramide synthase gene expression-promoting agent that employs a predetermined dipeptide as a main component for inducing function and that is capable of increasing the expression level of the ceramide synthase gene.
  • the skin quality improving agent, skin quality improving functional food, and cosmetics according to the present embodiment contain a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function. Therefore, it is possible to increase the expression level of the ceramide synthase gene, and use it as a skin quality improving agent, a functional food for improving skin quality, or a cosmetic that can improve skin quality.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Medicinal Chemistry (AREA)
  • Organic Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Epidemiology (AREA)
  • Dermatology (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Molecular Biology (AREA)
  • General Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Biochemistry (AREA)
  • Nutrition Science (AREA)
  • Immunology (AREA)
  • Biophysics (AREA)
  • Genetics & Genomics (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Mycology (AREA)
  • Birds (AREA)
  • Food Science & Technology (AREA)
  • Polymers & Plastics (AREA)
  • Coloring Foods And Improving Nutritive Qualities (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Cosmetics (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Peptides Or Proteins (AREA)

Abstract

Provided is a ceramide synthase gene expression promoting agent in which a prescribed dipeptide is employed as a main ingredient for bringing about a function, and which is capable of increasing the expression level of a ceramide synthase gene.　A dipeptide represented by Gly-Ile and/or Ile-Gly is contained as the main ingredient for bringing about a function. The present invention is also characterized in that the ceramide synthase gene is the ceramide synthase 3 (CERS3) gene.

Description

セラミド合成酵素遺伝子発現促進剤、肌質改善剤、肌質改善用機能食品及び化粧料並びにジペプチドの使用方法Ceramide synthase gene expression promoter, skin quality improving agent, functional food and cosmetic for improving skin quality, and usage of dipeptide
　本発明は、セラミド合成酵素遺伝子発現促進剤、肌質改善剤、肌質改善用機能食品及び化粧料並びにジペプチドの使用方法に関する。 The present invention relates to a method for using a ceramide synthase gene expression promoter, a skin quality improving agent, a functional food and cosmetic for improving skin quality, and a dipeptide.
　美しく健康的な肌は、その人の外観的な魅力を引き出す重要な要素の一つである。それ故、従来より肌の手入れは男女問わず行われている。 A beautiful and healthy skin is one of the important factors that bring out the attractiveness of a person's appearance. Therefore, skin care has been conventionally performed by both men and women.
　特に、若々しい肌を保つためには、張りや十分な潤いが大切である。従って、肌の保湿を担う成分は重要であり、このような保湿成分としては例えばセラミドが挙げられる。 In particular, to maintain youthful skin, firmness and sufficient moisture are important. Therefore, ingredients responsible for moisturizing the skin are important, and examples of such moisturizing ingredients include ceramide.
　セラミドは、細胞内にて生合成されるスフィンゴ脂質の一種であり、ヒトの皮膚の表皮層の表面を形成する角質層の主成分である。またセラミドは、過度な水分の蒸散を抑制すると共に、微生物に対するバリアとしての役割も有しており、肌を瑞々しく若々しい状態とするためには、肌のセラミド量を適切な量に保つ必要がある。 Ceramide is a type of sphingolipid that is biosynthesized in cells, and is the main component of the stratum corneum that forms the surface of the epidermal layer of human skin. In addition, ceramide suppresses excessive evaporation of moisture and also acts as a barrier against microorganisms. need to keep
　しかし、セラミドは加齢と共に失われがちである。 However, ceramide tends to be lost with aging.
　そこで、皮膚の表皮細胞におけるセラミドの生合成を促進すべく、ニコチン酸及び／又はニコチンアミドを含む菌培養物を有効成分としたセラミド合成促進剤が提案されている。 Therefore, in order to promote ceramide biosynthesis in epidermal cells of the skin, ceramide synthesis promoters containing fungal cultures containing nicotinic acid and/or nicotinamide as active ingredients have been proposed.
　そして、このようなセラミド合成促進剤によれば、皮膚表層内部で表皮細胞自身のセラミド合成を活発化させ皮膚バリア機能を改善することにより荒れ肌の改善および各種皮膚疾患の改善が期待できるとしている。 According to such a ceramide synthesis accelerator, improvement of rough skin and improvement of various skin diseases can be expected by activating ceramide synthesis of the epidermal cells themselves within the skin surface layer and improving the skin barrier function.
特開平１０－２５９１３５号公報JP-A-10-259135
　しかしながら、上記従来のニコチン酸やニコチンアミドを含む菌培養物を有効成分としたセラミド合成促進剤は、外用剤として提案されているものであり、経口摂取した際の効果については明らかにされていない。 However, the conventional ceramide synthesis accelerators containing nicotinic acid or nicotinamide-containing fungal cultures as active ingredients have been proposed as external preparations, and their effects when taken orally have not been clarified. .
　また、セラミドの生合成を促進するにあたり、機能惹起のための主要成分として更なるバリエーションを提供することは、使用者の選択肢を広げる点で望ましいと言える。 In addition, in promoting the biosynthesis of ceramide, it can be said that it is desirable to provide further variations as the main ingredient for inducing function in order to expand the options for users.
　本発明は、斯かる事情に鑑みてなされたものであって、機能惹起のための主要成分として所定のジペプチドが採用され、セラミド合成酵素遺伝子の発現量を増大させることが可能なセラミド合成酵素遺伝子発現促進剤を提供する。 The present invention has been made in view of such circumstances, and a ceramide synthase gene capable of increasing the expression level of the ceramide synthase gene by employing a predetermined dipeptide as a main component for inducing function. An expression promoter is provided.
　また本発明では、同じくセラミド合成酵素遺伝子発現促進機能を有する所定のジペプチドを含有させた肌質改善剤や肌質改善用機能食品、化粧料、更には同ジペプチドの使用方法についても提供する。 The present invention also provides a skin quality improving agent, a functional food for improving skin quality, cosmetics, and a method of using the same dipeptide containing a predetermined dipeptide having a ceramide synthase gene expression promoting function.
　上記従来の課題を解決するために、本発明に係るセラミド合成酵素遺伝子発現促進剤では、Gly-Ile及び／又はIle-Glyで表されるジペプチドを機能惹起のための主要成分として含有することとした。 In order to solve the conventional problems described above, the ceramide synthase gene expression promoter according to the present invention contains a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function. did.
　また本発明に係るセラミド合成酵素遺伝子発現促進剤では、前記セラミド合成酵素遺伝子は、セラミド合成酵素３(CERS3)遺伝子であることにも特徴を有する。 The ceramide synthase gene expression promoter according to the present invention is also characterized in that the ceramide synthase gene is the ceramide synthase 3 (CERS3) gene.
　また本発明に係る肌質改善剤や肌質改善用機能食品、化粧料では、Gly-Ile及び／又はIle-Glyで表されるジペプチドを機能惹起のための主要成分として含有することとした。 In addition, the skin quality-improving agent, functional food for skin quality improvement, and cosmetics according to the present invention contain a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function.
　また本発明では、セラミド合成酵素遺伝子発現促進剤における機能惹起のための主要成分としてや、肌質改善を目的とした関与成分として、Gly-Ile及び／又はIle-Glyで表されるジペプチドを使用することとした。 Further, in the present invention, a dipeptide represented by Gly-Ile and/or Ile-Gly is used as a main ingredient for inducing the function of a ceramide synthase gene expression promoter or as a participating ingredient for the purpose of improving skin quality. It was decided to.
　本発明に係るセラミド合成酵素遺伝子発現促進剤によれば、Gly-Ile及び／又はIle-Glyで表されるジペプチドを機能惹起のための主要成分として含有することとしたため、機能惹起のための主要成分として所定のジペプチドが採用され、セラミド合成酵素遺伝子の発現量を増大させることが可能なセラミド合成酵素遺伝子発現促進剤を提供することができる。 According to the ceramide synthase gene expression promoter of the present invention, the dipeptide represented by Gly-Ile and/or Ile-Gly is contained as a main component for inducing function. It is possible to provide a ceramide synthase gene expression promoter that employs a predetermined dipeptide as a component and is capable of increasing the expression level of the ceramide synthase gene.
　また、前記セラミド合成酵素遺伝子は、セラミド合成酵素３(CERS3)遺伝子であることとすれば、主に皮膚やケラチノサイトにて多く発現するセラミド合成酵素３を標的とすることができ、より効率的なセラミド合成酵素遺伝子発現促進剤とすることができる。 In addition, if the ceramide synthase gene is the ceramide synthase 3 (CERS3) gene, it is possible to target ceramide synthase 3, which is mainly expressed in skin and keratinocytes, and to achieve more efficient ceramide synthase 3. It can be used as a ceramide synthase gene expression promoter.
　また、本発明に係る肌質改善剤や肌質改善用機能食品、化粧料では、Gly-Ile及び／又はIle-Glyで表されるジペプチドを機能惹起のための主要成分として含有することとしたため、セラミド合成酵素遺伝子の発現量を増大させることが可能な肌質改善剤や肌質改善用機能食品、化粧料とすることができる。 In addition, the skin quality improving agent, skin quality improving functional food, and cosmetics according to the present invention contain a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function. , a skin quality improving agent, a functional food for improving skin quality, and a cosmetic that can increase the expression level of the ceramide synthase gene.
　また、本発明では、セラミド合成酵素遺伝子発現促進剤における機能惹起のための主要成分としてや、肌質改善を目的とした関与成分として、Gly-Ile及び／又はIle-Glyで表されるジペプチドを使用することとしたため、Gly-Ile及び／又はIle-Glyで表されるジペプチドによるセラミド合成酵素遺伝子の発現量の増大に由来した効果を享受することができる。 Further, in the present invention, a dipeptide represented by Gly-Ile and/or Ile-Gly is used as a main ingredient for inducing the function of the ceramide synthase gene expression promoter or as a participating ingredient for the purpose of improving skin quality. Since it was decided to use it, it is possible to enjoy the effect derived from the increase in the expression level of the ceramide synthase gene by the dipeptide represented by Gly-Ile and/or Ile-Gly.
各ジペプチドの添加によるHaCaT細胞の細胞生存率を示す説明図である。FIG. 4 is an explanatory diagram showing the cell viability of HaCaT cells by addition of each dipeptide. 各ジペプチドの添加によるHaCaT細胞のセラミド合成酵素遺伝子の発現変化を示す説明図である。FIG. 4 is an explanatory diagram showing changes in the expression of ceramide synthase gene in HaCaT cells due to the addition of each dipeptide. ジペプチドの少量添加によるHaCaT細胞のセラミド合成酵素遺伝子の発現変化を示す説明図である。FIG. 4 is an explanatory diagram showing changes in the expression of the ceramide synthase gene in HaCaT cells due to the addition of a small amount of dipeptide. 試験食品を摂取させた際の水分量に関する試験結果を示す説明図である。FIG. 3 is an explanatory diagram showing the test results regarding the amount of water when the test food was ingested. 試験食品を摂取させた際の経皮水分蒸散量に関する試験結果を示す説明図である。FIG. 3 is an explanatory diagram showing the test results regarding the amount of transepidermal water loss when the test food was ingested. 試験食品を摂取させた際の肌の総弾力Ｒ２に関する試験結果を示す説明図である。FIG. 3 is an explanatory diagram showing test results regarding total elasticity R2 of skin when test foods were ingested. 試験食品を摂取させた際の肌の粘弾性（Ｒ５（正味弾性））に関する試験結果を示す説明図である。FIG. 3 is an explanatory diagram showing test results regarding skin viscoelasticity (R5 (net elasticity)) when test foods were ingested. 試験食品を摂取させた際の肌の回復力Ｒ７に関する試験結果を示す説明図である。FIG. 3 is an explanatory diagram showing test results regarding skin recovery power R7 when a test food is ingested.
　本発明は、機能惹起のための主要成分として所定のジペプチドが採用され、セラミド合成酵素遺伝子の発現量を増大させることが可能なセラミド合成酵素遺伝子発現促進剤を提供するものである。 The present invention provides a ceramide synthase gene expression promoter that employs a predetermined dipeptide as a main component for inducing function and is capable of increasing the expression level of the ceramide synthase gene.
　本実施形態に係るセラミド合成酵素遺伝子発現促進剤の用途は特に限定されるものではなく、食品や化粧料に肌質改善の機能を付与すべく添加原料として使用したり、肌質改善のための外用剤として使用するのは勿論のこと、例えば試験研究用試薬としても使用することができる。 The use of the ceramide synthase gene expression promoter according to the present embodiment is not particularly limited. It can be used not only as an external agent but also, for example, as a reagent for testing and research.
　本実施形態に係るセラミド合成酵素遺伝子発現促進剤の特徴として、同促進剤は、Gly-Ile（グリシン－イソロイシン）及び／又はIle-Gly（イソロイシン－グリシン）で表されるジペプチドを機能惹起のための主要成分として含有している。 As a feature of the ceramide synthase gene expression promoter according to the present embodiment, the promoter uses a dipeptide represented by Gly-Ile (glycine-isoleucine) and/or Ile-Gly (isoleucine-glycine) to induce function. contains as a major component of
　Gly-IleやIle-Glyは、グリシンとイソロイシンとがペプチド結合したジペプチドである。Gly-IleやIle-Glyの由来は特に限定されるものではなく、所定の素材や食材の乾燥物や抽出物であったり、所定のタンパク質の消化物由来であったり、ペプチド合成によるものなど、その由来が目的に反しないものであればいずれであっても良い。なお、本明細書では特に断りのない場合、アミノ酸配列は左側をＮ末端として記載している。 Gly-Ile and Ile-Gly are dipeptides in which glycine and isoleucine are peptide-bonded. The origin of Gly-Ile and Ile-Gly is not particularly limited. It may be of any origin as long as it does not contradict the purpose. In this specification, unless otherwise specified, the left side of the amino acid sequence is described as the N-terminus.
　実験結果を参照しつつ追って説明するが、本発明者らの鋭意研究により、Gly-IleやIle-Glyはセラミド合成酵素遺伝子の発現を促進することが明らかになっており、皮膚中のセラミド量を維持したり増加させる効果が期待できる。 As will be described later with reference to the experimental results, the present inventors' intensive research has revealed that Gly-Ile and Ile-Gly promote the expression of the ceramide synthase gene. can be expected to maintain or increase
　なお、肌に好影響を及ぼすとされているジペプチドとしてGly-LeuやLeu-Glyが知られているが、本願にて提案するジペプチドはこれら物質とは構造的に全く異なる物質であり、また、Gly-LeuやLeu-Glyと比較してより高度なセラミド合成酵素遺伝子発現促進効果を有することは、後述する試験結果から明らかである。 Gly-Leu and Leu-Gly are known as dipeptides that are said to have a positive effect on the skin, but the dipeptide proposed in the present application is a substance that is structurally completely different from these substances, and It is clear from the test results described later that it has a higher ceramide synthase gene expression promoting effect than Gly-Leu or Leu-Gly.
　また本願では、肌質改善剤についても提供する。本実施形態に係る肌質改善剤は、Gly-Ile及び／又はIle-Glyで表されるジペプチドを機能惹起のための主要成分として含有する点で特徴的である。本願において肌質改善剤は、経口的に摂取されても良いし、また、皮膚外用剤として使用されても良い。皮膚外用剤は、医薬品や医薬部外品、化粧品（薬用化粧品も含む）を意味するのは勿論のこと、これらの枠にとらわれず、皮膚に対して外用する剤であればこの概念に含まれる。 This application also provides a skin quality improving agent. The skin quality improving agent according to the present embodiment is characteristic in that it contains a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function. In the present application, the skin quality improving agent may be taken orally, or may be used as an external skin preparation. External agents for skin include pharmaceuticals, quasi-drugs, and cosmetics (including medicated cosmetics), but are not bound by these frameworks and are included in this concept as long as they are externally applied to the skin. .
　また本願では、この皮膚外用剤の一態様でもある化粧料についても提供する。すなわち、本実施形態に係る化粧料は、Gly-Ile及び／又はIle-Glyで表されるジペプチドを有効成分として含有する点で特徴的である。 In addition, the present application also provides a cosmetic, which is one aspect of this external preparation for skin. That is, the cosmetic according to the present embodiment is characteristic in that it contains a dipeptide represented by Gly-Ile and/or Ile-Gly as an active ingredient.
　本願において化粧料は、いわゆるメーキャップ化粧品の他、基礎化粧品やヘアトニック、香水、歯磨き、シャンプー、リンス、身体の洗浄等に用いられる石鹸や洗浄料、入浴剤などのトイレタリー製品も含むものであり、また、予防効果等を謳う、薬用化粧品も含まれる。 In the present application, cosmetics include not only so-called makeup cosmetics, but also basic cosmetics, hair tonics, perfumes, toothpastes, shampoos, rinses, soaps and cleansers used for washing the body, and toiletry products such as bath agents. It also includes medicinal cosmetics claiming preventive effects.
　また、メーキャップ化粧品としては、ファンデーションや眉墨（アイブロー）、マスカラ、アイシャドー、アイライン、口紅、グロス、頬紅（チーク）、白粉、マニキュアなどが挙げられ、基礎化粧品としては、例えば化粧水や乳液、洗顔料、クレンジング、美容液、クリームなどが挙げられる。なお、上述した皮膚外用剤や化粧料等についての説明は、本発明の理解に供すべくこれらに相当する物品等の一例を列挙したものであり、各語句の解釈はこれら列挙された物品等に限定されるものではない。ただし、本出願人が本願を権利化するにあたり、本発明をこれら物品等に限定することを妨げない。 Examples of makeup cosmetics include foundations, eyebrows, mascara, eye shadows, eyeliners, lipsticks, glosses, cheeks, face powders, and nail polishes. Basic cosmetics include lotions, milky lotions, Facial cleansers, cleansers, serums, creams and the like can be mentioned. It should be noted that the above descriptions of external preparations for skin, cosmetics, and the like are examples of corresponding articles, etc., in order to facilitate understanding of the present invention, and the interpretation of each term is based on the articles, etc., listed above. It is not limited. However, it does not preclude the applicant from limiting the present invention to these articles and the like in obtaining the right of the present application.
　また本願では、Gly-Ile及び／又はIle-Glyで表されるジペプチドのセラミド合成酵素遺伝子発現促進剤における機能惹起のための主要成分としての使用方法についても提供する。セラミド合成酵素遺伝子発現促進効果を有する成分はこれまでに幾つか提案されているが、これまでGly-Ile及び／又はIle-Glyで表されるジペプチドを機能惹起のための主要成分として含有するセラミド合成酵素遺伝子発現促進剤は提案されていない。 The present application also provides a method of using a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing the function of a ceramide synthase gene expression promoter. Several components having a ceramide synthase gene expression-enhancing effect have been proposed so far. Synthase gene expression promoters have not been proposed.
　また、使用者に対し、セラミド合成酵素遺伝子発現促進作用乃至効果を惹起しうる新たな成分を提案することは、選択肢を増やす上で有意義である。 In addition, proposing new ingredients that can induce ceramide synthase gene expression promoting action or effect to users is significant in increasing options.
　また本願では、Gly-Ile及び／又はIle-Glyで表されるジペプチドの機能食品における肌質改善を目的とした関与成分としての使用方法についても提供する。経口摂取により肌質改善を図る成分は幾つか提案されているが、これまでGly-Ile及び／又はIle-Glyで表されるジペプチドを肌質改善を目的とした関与成分として含有する機能食品は提案されていない。 The present application also provides a method of using dipeptides represented by Gly-Ile and/or Ile-Gly as participating ingredients in functional foods for the purpose of improving skin quality. Several ingredients have been proposed to improve skin quality through oral ingestion. Not proposed.
　また本願では、肌質改善用機能食品についても提供するものであり、特に、経口摂取により優れた皮膚の保湿効果を発揮できる肌質改善用機能食品を提供するものである。 The present application also provides a functional food for improving skin quality, and in particular, provides a functional food for improving skin quality that can exert an excellent moisturizing effect on the skin when taken orally.
　機能食品は、医薬品成分を含まない健康の保持増進に寄与するとされる食品全般を包含する概念であり、例えば、栄養補助食品や健康補助食品、栄養調性食品のほか、所謂サプリメントなどの一般食品であったり、特定保健用食品や栄養機能食品、機能性表示食品の如き保健機能食品も含まれる。 Functional food is a concept that includes all foods that are said to contribute to the maintenance and promotion of health that do not contain pharmaceutical ingredients. It also includes food with health claims such as food for specified health use, food with nutrient function claims, and food with function claims.
　また、サプリメント様とした場合には、その剤形は特に限定されるものではなく、錠剤、カプセル剤、細粒剤、丸剤、トローチ剤、液剤、ゼリー様など、あらゆる剤形を選択することができる。 In addition, if it is a supplement-like, the dosage form is not particularly limited, and any dosage form such as tablets, capsules, fine granules, pills, lozenges, liquids, and jelly-like can be selected. can be done.
　なお、上述した外用剤や化粧料、機能食品についての説明は、本発明の理解に供すべくこれらに相当する物品等の一例を列挙したものであり、各語句の解釈はこれら列挙された物品等に限定されるものではない。ただし、本出願人が本願を権利化するにあたり、本発明をこれら物品等に限定することを妨げない。 It should be noted that the above descriptions of topical preparations, cosmetics, and functional foods are examples of corresponding articles, etc., for better understanding of the present invention. is not limited to However, it does not preclude the applicant from limiting the present invention to these articles and the like in obtaining the right of the present application.
　本実施形態に係るセラミド合成酵素遺伝子発現促進剤や肌質改善剤、肌質改善用機能食品、化粧料等に含有させる機能惹起のための成分としてのGly-Ile及び／又はIle-Glyの濃度は前記機能を惹起可能な濃度であれば特に限定されるものではない。敢えて一例を挙げるならば、肌質改善用機能食品に含まれるGly-IleやIle-Glyの濃度としては、例えば一日あたり20～600μｇのGly-Ile又はIle-Glyを摂取できる量としたり、Gly-IleとIle-Glyの合計の量として一日あたり20～600μｇ摂取できる量とすることができる。付言すれば、一日あたり１粒を目安に摂取する肌質改善用機能食品であれば、１粒に含まれるGly-Ile及び／又はIle-Glyの量を20～600μｇとしたり、一日あたり２粒が目安であれば１粒あたり10～300μｇとするように、１粒や１カプセル、一包など摂取単位量に含まれるGly-Ile及び／又はIle-Glyの量を適宜調整することも可能である。 Concentration of Gly-Ile and/or Ile-Gly as a component for inducing function to be contained in the ceramide synthase gene expression promoter, skin quality improving agent, functional food for improving skin quality, cosmetics, etc. according to the present embodiment is not particularly limited as long as the concentration is capable of inducing the above function. To give an example, the concentration of Gly-Ile or Ile-Gly contained in the functional food for improving skin quality is, for example, an amount that allows 20 to 600 μg of Gly-Ile or Ile-Gly to be ingested per day, The total amount of Gly-Ile and Ile-Gly can be taken in an amount of 20 to 600 µg per day. In addition, if it is a functional food for improving skin quality that is ingested as a guideline for one tablet per day, the amount of Gly-Ile and/or Ile-Gly contained in one tablet is 20 to 600 μg. If two tablets are a guideline, the amount of Gly-Ile and/or Ile-Gly contained in the intake unit amount such as one tablet, one capsule, and one packet may be adjusted appropriately so that one tablet is 10 to 300 μg. It is possible.
また、化粧料としては、0.03～200μg／gのGly-Ile又はIle-Glyを含ませたり、Gly-IleとIle-Glyの合計の濃度を0.03～200μg／gとすることができる。セラミド合成酵素遺伝子発現促進剤や肌質改善剤におけるGly-IleやIle-Glyの濃度は、それぞれ実験等での使用に適した濃度としたり、食品や化粧料の製造のための原料として適した濃度とすることができる。 In addition, the cosmetic may contain 0.03 to 200 µg/g of Gly-Ile or Ile-Gly, or the total concentration of Gly-Ile and Ile-Gly may be 0.03 to 200 µg/g. The concentrations of Gly-Ile and Ile-Gly in the ceramide synthase gene expression accelerator and the skin quality improving agent are adjusted to concentrations suitable for use in experiments, etc., and suitable as raw materials for the production of foods and cosmetics. concentration.
　また、本実施形態に係るセラミド合成酵素遺伝子発現促進剤や肌質改善剤、肌質改善用機能食品、化粧料等には、機能惹起のための成分としてのGly-Ile及び／又はIle-Glyの他に、剤形や食品形態等に応じた賦形剤や添加剤、補助成分などを含むこともできる。 Further, the ceramide synthase gene expression promoter, skin quality improving agent, skin quality improving functional food, cosmetics, etc. according to the present embodiment may contain Gly-Ile and/or Ile-Gly as a component for inducing function. In addition, it may contain excipients, additives, auxiliary ingredients, and the like according to dosage forms, food forms, and the like.
　賦形剤としては、例えば、剤形が固形剤の場合には、乳糖や結晶セルロース、デンプンなどとすることができる。 Examples of excipients include lactose, crystalline cellulose, and starch when the dosage form is a solid formulation.
　また添加剤としては、例えば、安定剤、界面活性剤、可溶化剤、可塑剤、甘味剤、抗酸化剤、着香剤、着色剤、保存剤、無機充填剤等を挙げることができる。 Examples of additives include stabilizers, surfactants, solubilizers, plasticizers, sweeteners, antioxidants, flavoring agents, coloring agents, preservatives, inorganic fillers, and the like.
　以下、本実施形態に係るセラミド合成酵素遺伝子発現促進剤、肌質改善剤、肌質改善用機能食品及び化粧料並びにジペプチドの使用方法に関し、実験過程や結果を参照しながら更に説明する。 The method of using the ceramide synthase gene expression promoter, skin quality improving agent, skin quality improving functional food and cosmetic, and dipeptide according to the present embodiment will be further described below with reference to the experimental process and results.
〔１．セラミド合成酵素遺伝子の発現試験〕
　皮膚は表皮および真皮により構成され、表皮は基底層、有棘層、顆粒層および角層に分類される。皮膚の最外層に位置する角層細胞間において、セラミドなどの脂質により構成されるラメラ構造は、体内の水分保持や細菌およびウイルスの侵入を防ぐ物理的なバリアとしての役割を担う。 [1. Expression test of ceramide synthase gene]
The skin is composed of the epidermis and dermis, which are divided into the stratum basale, the stratum spinosum, the stratum granulosum and the stratum corneum. A lamellar structure composed of lipids such as ceramides between stratum corneum cells located in the outermost layer of the skin plays a role as a physical barrier to retain water in the body and prevent invasion of bacteria and viruses.
　本検討では、セラミド合成酵素遺伝子の発現量の変化を検討することで、Gly-IleやIle-Gly（以下、本願ジペプチドともいう。）の表皮細胞の改善効果を評価した。また、比較対照として、Gly-LeuやLeu-Gly（以下、比較ジペプチドともいう。）を用い、本願ジペプチドとの違いについても検討した。 In this study, the improvement effect of Gly-Ile and Ile-Gly (hereinafter also referred to as the dipeptide of the present application) on epidermal cells was evaluated by examining changes in the expression level of the ceramide synthase gene. In addition, Gly-Leu and Leu-Gly (hereinafter also referred to as comparative dipeptides) were used as controls for comparison, and differences from the dipeptides of the present application were also examined.
　ヒト表皮角化細胞（HaCaT細胞）は、Dulbecco's Modified Eagle Medium (DMEM)(高グルコース)(含1%ペニシリン-ストレプトマイシンおよび10%ウシ胎児血清(FBS))を用いて、コンフルエントになるまでφ10cmディッシュにて前培養した。 Human epidermal keratinocytes (HaCaT cells) were cultured in φ10 cm dishes using Dulbecco's Modified Eagle Medium (DMEM) (high glucose) (containing 1% penicillin-streptomycin and 10% fetal bovine serum (FBS)) until confluent. was pre-cultured.
　その後、リン酸緩衝生理食塩水(PBS)で洗浄し、培地に再懸濁後、24穴プレートに1.0×105cells/wellの濃度で播種しCO2インキュベーター(37℃,5%CO2)でオーバーナイト培養した。 Thereafter, the cells were washed with phosphate buffered saline (PBS), resuspended in medium, seeded in a 24-well plate at a concentration of 1.0×10 5 cells/well, and placed in a CO 2 incubator (37°C, 5% CO 2 ). was cultured overnight.
　HaCaT細胞播種から24時間後、所定終濃度となる量のGly-Ile、Ile-Gly、Gly-Leu、Leu-Glyの何れかを含む無血清のDMEM(高グルコース)(含1%ペニシリン-ストレプトマイシン)を各ウェル1mLずつ加え、CO2インキュベーター(37℃,5%CO2)で48時間培養した。 24 hours after seeding HaCaT cells, serum-free DMEM (high glucose) (containing 1% penicillin-streptomycin) containing either Gly-Ile, Ile-Gly, Gly-Leu, or Leu-Gly at a predetermined final concentration ) was added to each well and cultured for 48 hours in a CO 2 incubator (37°C, 5% CO 2 ).
　次に、リアルタイムPCRによる遺伝子の転写発現量の変化について試験を行った。まず、各ジペプチドを添加して48時間が経過した後のHaCaT細胞を回収し、RNeasy Mini kit(Qiagen)を用いてtotal RNAを抽出した。 Next, we tested changes in gene transcription expression levels by real-time PCR. First, HaCaT cells were collected 48 hours after adding each dipeptide, and total RNA was extracted using RNeasy Mini kit (Qiagen).
　次に、ReverTra Ace qPCR RT Master Mix with gDNA Remover(TOYOBO)にて、抽出したtotal RNAからcDNAを合成した。合成したcDNAを鋳型としてAriaMX(Agilent)装置でリアルタイムPCRを行った。リアルタイムPCR反応には、THUNDERBIRD SYBR qPCR Mix(TOYOBO)を用いた。 Next, cDNA was synthesized from the extracted total RNA using ReverTra Ace qPCR RT Master Mix with gDNA Remover (TOYOBO). Using the synthesized cDNA as a template, real-time PCR was performed using an AriaMX (Agilent) apparatus. THUNDERBIRD SYBR qPCR Mix (TOYOBO) was used for the real-time PCR reaction.
　また、リアルタイムPCR反応には、セラミド合成酵素遺伝子CERS2用プライマーとして5’-CCGATTACCTGCTGGAGTCAG-3’（配列番号１）および5’-GGCGAAGACGATGAAGATGTTG-3’（配列番号２）を、セラミド合成酵素遺伝子CERS3用プライマーとして5’-ACATTCCACAAGGCAACCATTG-3’（配列番号３）および5’-CTCTTGATTCCGCCGACTCC-3’（配列番号４）を、セラミド合成酵素遺伝子CERS4用プライマーとして5’-GGAGGCCTGTAAGATGGTCA-3’（配列番号５）および5’-GAGGACCAGTCGGGTGTAGA-3’（配列番号６）を、内部標準β-アクチン用プライマーとして5’-GGGTCAGAAGGACTCCTATG-3’（配列番号７）および5’-GTAACAATGCCATGTTCAAT-3’（配列番号８）を使用した。リアルタイムPCR反応条件として、95℃、60秒の初期変性後、95℃、15秒での変性、60℃、60秒のアニーリング/伸長という2ステップのPCR反応を40サイクル行った。 In the real-time PCR reaction, 5'-CCGATTACCTGCTGGAGTCAG-3' (SEQ ID NO: 1) and 5'-GGCGAAGACGATGAAGATGTTG-3' (SEQ ID NO: 2) were used as primers for the ceramide synthase gene CERS2, and a primer for the ceramide synthase gene CERS3. 5'-ACATTCCACAAGGCAACCATTG-3' (SEQ ID NO: 3) and 5'-CTCTTGATTCCGCCGACTCC-3' (SEQ ID NO: 4) as primers for the ceramide synthase gene CERS4 5'-GGAGGCCTGTAAGATGGTCA-3' (SEQ ID NO: 5) and 5 '-GAGGACCAGTCGGGTGTAGA-3' (SEQ ID NO: 6) was used, and 5'-GGGTCAGAAGGACTCCTATG-3' (SEQ ID NO: 7) and 5'-GTAACAATGCCATGTTCAAT-3' (SEQ ID NO: 8) were used as primers for internal standard β-actin. As real-time PCR reaction conditions, 40 cycles of a two-step PCR reaction consisting of initial denaturation at 95°C for 60 seconds, denaturation at 95°C for 15 seconds, and annealing/extension at 60°C for 60 seconds were performed.
　本願ジペプチドであるIle-GlyやGly-Ile、比較ジペプチドであるLeu-GlyやGly-Leuをヒト表皮角化細胞株の1つであるHaCaT細胞に添加し、セラミド合成酵素遺伝子の発現変化を検討した。図1に、各ジペプチドの添加によるHaCaT細胞の細胞生存率を示す。 Ile-Gly and Gly-Ile, which are the dipeptides of the present application, and Leu-Gly and Gly-Leu, which are the comparative dipeptides, were added to HaCaT cells, one of the human epidermal keratinocyte lines, and the changes in the expression of the ceramide synthase gene were examined. did. FIG. 1 shows the cell viability of HaCaT cells with the addition of each dipeptide.
　図１に示すように本願ジペプチドや比較用ジペプチドを添加すると、コントロールと比較してヒト角化細胞HaCaTの細胞生存率が上昇する傾向が認められた。特に、本願ジペプチドを添加した場合は、0.1μg/mL、10μg/mL、200μg/mLのいずれの濃度においてもコントロールに対する細胞生存率の上昇が確認された。また、Gly-Leu 10μg/mL、Ile-Gly 0.1および10μg/mL、Gly-Ile 200μg/mLでは、コントロールと比較して有意に細胞生存率が上昇することが確認できた。これらのことから、本願ジペプチドであるIle-GlyやGly-Ileには角化細胞賦活化作用が期待できることが示唆された。 As shown in Figure 1, addition of the dipeptide of the present application and the dipeptide for comparison tended to increase the cell viability of human keratinocytes HaCaT compared to the control. In particular, when the dipeptide of the present application was added, an increase in cell viability was confirmed as compared to the control at any concentration of 0.1 μg/mL, 10 μg/mL and 200 μg/mL. It was also confirmed that Gly-Leu 10 μg/mL, Ile-Gly 0.1 and 10 μg/mL, and Gly-Ile 200 μg/mL significantly increased the cell viability compared to the control. These findings suggest that the dipeptides of the present invention, Ile-Gly and Gly-Ile, are expected to have a keratinocyte activating effect.
　図２には、本願ジペプチドであるIle-GlyやGly-Ile、比較ジペプチドであるLeu-GlyやGly-Leuを添加しHaCaT細胞のセラミド合成酵素遺伝子の発現変化を示している。図２（ａ）はセラミド合成酵素CERS2遺伝子の発現量のコントロールに対する相対値を示し、同様に図２（ｂ）はセラミド合成酵素CERS3遺伝子、図２（ｃ）はセラミド合成酵素CERS4遺伝子の発現量を相対値で示している。 Fig. 2 shows changes in the expression of the ceramide synthase gene in HaCaT cells with the addition of the dipeptides of the present application, Ile-Gly and Gly-Ile, and the comparative dipeptides, Leu-Gly and Gly-Leu. FIG. 2(a) shows the expression level of the ceramide synthase CERS2 gene relative to the control, similarly FIG. 2(b) shows the expression level of the ceramide synthase CERS3 gene, and FIG. is shown as a relative value.
　セラミド合成酵素CERS2遺伝子は、腎臓、肝臓および腸など多くの細胞組織で大量に発現していることが報告されている。また、セラミド合成酵素CERS3遺伝子は、主に皮膚、ケラチノサイトで発現している。さらに、セラミド合成酵素CERS4遺伝子は、皮膚に限らず白血球、心臓および肝臓でも発現していることが知られている。 It has been reported that the ceramide synthase CERS2 gene is highly expressed in many cell tissues such as kidney, liver and intestine. In addition, the ceramide synthase CERS3 gene is mainly expressed in skin and keratinocytes. Furthermore, it is known that the ceramide synthase CERS4 gene is expressed not only in skin but also in leukocytes, heart and liver.
　本検討では、各ジペプチドをHaCaT細胞へ添加後、リアルタイムPCRを用いてセラミド合成酵素CERS2、CERS3およびCERS4遺伝子の発現解析をした。その結果、図２（ａ）に示す通り、CERS2遺伝子の発現は、コントロールと比較していずれの濃度でも有意な増加は確認出来なかった。 In this study, after each dipeptide was added to HaCaT cells, real-time PCR was used to analyze the expression of the ceramide synthase CERS2, CERS3, and CERS4 genes. As a result, as shown in FIG. 2(a), no significant increase in the expression of the CERS2 gene could be confirmed at any concentration compared to the control.
　これに対し、セラミド合成酵素CERS3遺伝子は、図２（ｂ）に示すように、本願ジペプチドであるIle-GlyやGly-Ileの添加により、コントロールと比較していずれの濃度においても発現量の上昇が確認された。 In contrast, as shown in Fig. 2(b), the expression level of the ceramide synthase CERS3 gene increased at any concentration compared to the control by the addition of Ile-Gly and Gly-Ile, which are the dipeptides of the present application. was confirmed.
　特に、Gly-Ileの添加は、濃度に依存した発現量の有意な増加が認められ、200μg/mLの濃度ではコントロールに比して発現量が３倍にも増加している点は極めて興味深い。 In particular, the addition of Gly-Ile showed a significant concentration-dependent increase in the expression level, and it is extremely interesting that at a concentration of 200 μg/mL, the expression level increased 3-fold compared to the control.
　また、Ile-Glyの添加は、CERS3遺伝子の発現量の増加をもたらし、また、0.1μg/mLの濃度での使用では有意差が認められるのであるが、濃度上昇に伴い発現量が低下している点で特徴的である。 In addition, the addition of Ile-Gly increased the expression level of the CERS3 gene, and although a significant difference was observed when used at a concentration of 0.1 μg/mL, the expression level decreased as the concentration increased. It is characteristic that
　なお、比較ジペプチドであるLeu-GlyやGly-Leuの添加によってもコントロールと比較して最大で約1.6倍の発現量の上昇が確認された。但し、本願ジペプチドほどの顕著な発現量の増加は見られなかった。 The addition of the comparative dipeptides Leu-Gly and Gly-Leu also confirmed an increase in the expression level of up to about 1.6 times compared to the control. However, no significant increase in the expression level was observed as compared with the dipeptide of the present application.
　また、データは割愛するが、発明者らが行った研究の経験上、0.03μg/g程度の濃度でのIle-GlyやGly-Ileの添加であっても、CERS3遺伝子の発現量がコントロールに比して有意に増加することが明らかとなっている。 In addition, although the data are omitted, in the experience of the research conducted by the inventors, even with the addition of Ile-Gly or Gly-Ile at a concentration of about 0.03 μg / g, the expression level of the CERS3 gene was lower than that of the control. It has been clarified that it significantly increases compared to the
　セラミド合成酵素CERS4遺伝子は、図２（ｃ）に示すように、Gly-Ile添加で発現量の顕著な上昇が確認された。 As for the ceramide synthase CERS4 gene, as shown in Fig. 2(c), a marked increase in the expression level was confirmed with the addition of Gly-Ile.
　これらの結果から、本願ジペプチドには細胞賦活化作用が期待でき、セラミド合成酵素CERS3およびCERS4遺伝子の発現を亢進する作用を有することが示唆された。この作用は、特に本願ジペプチドの中でもGly-Ileで高いことも示唆された。このように、本願ジペプチドは、セラミド合成酵素遺伝子の発現促進効果が認められ、皮膚のバリア機能に重要な役割を果たすセラミド合成促進効果が期待できる。 These results suggest that the dipeptide of the present application can be expected to have a cell activating effect, and that it has the effect of enhancing the expression of the ceramide synthase CERS3 and CERS4 genes. It was also suggested that this effect is particularly high with Gly-Ile among the dipeptides of the present application. Thus, the dipeptide of the present application is found to have the effect of promoting the expression of the ceramide synthase gene, and is expected to have the effect of promoting ceramide synthesis, which plays an important role in the skin barrier function.
　また、Gly-Ile及び／又はIle-Glyで表されるジペプチドは、肌質改善剤における機能惹起のための主要成分であったり、機能食品における肌質改善のための関与成分であったり、化粧料における肌質改善のための有効成分として利用可能であることが示された。 In addition, the dipeptide represented by Gly-Ile and/or Ile-Gly is a main component for inducing function in skin quality improving agents, a component involved in improving skin quality in functional foods, and a cosmetic component. It was shown that it can be used as an active ingredient for improving skin quality in cosmetics.
〔２．セラミド合成酵素遺伝子の発現試験（低用量）〕
　次に、本願ジペプチドであるGly-IleやIle-Glyによるセラミド合成酵素遺伝子の発現について、上述した濃度よりも低い濃度での発現誘導が可能であるかについて、同様の手法により確認を行った。 [2. Expression test of ceramide synthase gene (low dose)]
Next, whether the expression of the ceramide synthase gene by Gly-Ile or Ile-Gly, which is the dipeptide of the present application, can be induced at a concentration lower than the concentration described above was confirmed by a similar method.
　本検討では、少なくとも0.00038μg/mLのGly-Ileと、少なくとも0.00097μg/mLのIle-Glyとを含有する、両ジペプチドの合計濃度が0.00135μg/mLのサンプル溶液Ｓ１と、少なくとも0.00076μg/mLのGly-Ileと、少なくとも0.00195μg/mLのIle-Glyとを含有する、両ジペプチドの合計濃度が0.00271μg/mLのサンプル溶液Ｓ２（サンプル溶液Ｓ１の２倍濃度溶液）と、少なくとも0.00152μg/mLのGly-Ileと、少なくとも0.0039μg/mLのIle-Glyとを含有する、両ジペプチドの合計濃度が0.00542μg/mLのサンプル溶液Ｓ３（サンプル溶液Ｓ２の２倍濃度溶液）とを被験液とし、HaCaT細胞播種から24時間後、上記何れかの被験液を構成する終濃の無血清のDMEM(高グルコース)(含1%ペニシリン-ストレプトマイシン)を各ウェル1mLずつ加え、CO2インキュベーター(37℃,5%CO2)で48時間培養した。 In this study, sample solution S1 containing at least 0.00038 μg/mL of Gly-Ile and at least 0.00097 μg/mL of Ile-Gly with a total concentration of both dipeptides of 0.00135 μg/mL and of Gly-Ile and at least 0.00195 μg/mL of Ile-Gly, with a total concentration of both dipeptides of 0.00271 μg/mL (double concentration solution of sample solution S1) and at least 0.00152 μg/mL of Ile-Gly. Sample solution S3 containing 1 mL of Gly-Ile and at least 0.0039 μg/mL of Ile-Gly and having a total concentration of 0.00542 μg/mL of both dipeptides (double concentration solution of sample solution S2) was used as the test solution. , 24 hours after the seeding of HaCaT cells, 1 mL of final serum-free DMEM (high glucose) (containing 1% penicillin-streptomycin) constituting any of the above test solutions was added to each well, and placed in a CO 2 incubator (37°C). , 5% CO 2 ) for 48 hours.
　まず図３（ａ）に、各サンプル溶液Ｓ１～Ｓ３の添加によるHaCaT細胞の細胞生存率を示す。図３（ａ）に示すように、各サンプル溶液Ｓ１～Ｓ３は、コントロールとの比較において、特に有意な差は確認されなかった。 First, Fig. 3(a) shows the cell viability of HaCaT cells with the addition of each sample solution S1 to S3. As shown in FIG. 3(a), no significant difference was observed between the sample solutions S1 to S3 and the control.
　図３（ｂ）には、各サンプル溶液Ｓ１～Ｓ３を添加した後のHaCaT細胞のセラミド合成酵素遺伝子の発現変化を示している。図３（ｂ）に示すように、CERS2遺伝子の発現は、コントロールと比較していずれの濃度でも有意な差は確認出来なかった。一方、皮膚での発現が報告されているCERS3遺伝子やCERS4遺伝子は、コントロールと比較して約２～４倍程度に発現上昇していた。また、CERS3遺伝子は、CERS4遺伝子と比較すると、より顕著な発現の上昇が見られた。 Fig. 3(b) shows changes in the expression of the ceramide synthase gene in HaCaT cells after the addition of each sample solution S1 to S3. As shown in FIG. 3(b), no significant difference in the expression of the CERS2 gene was observed at any concentration compared to the control. On the other hand, the CERS3 and CERS4 genes, which have been reported to be expressed in the skin, were upregulated by about 2 to 4 times compared to controls. In addition, the CERS3 gene showed a more pronounced increase in expression compared to the CERS4 gene.
　以上の結果から、本願ジペプチドであるGly-IleやIle-Glyには細胞毒性は認められず、セラミド合成酵素CERS3およびCERS4遺伝子の発現を冗進する作用を有することが示唆された。したがって、本願ジペプチドであるGly-IleやIle-Glyは、両ジペプチドの合計濃度が0.00135μg/mL以上において、皮膚のバリア機能に重要な役割を果たすセラミド合成促進効果が期待出来る。付言すれば、好ましくは例えば、少なくとも0.00038μg/mLのGly-Ileと、少なくとも0.00097μg/mLのIle-Glyとを含有する、両ジペプチドの合計濃度が0.03μg/mLとすることで、より堅実に効果を発揮させることができる。 From the above results, it was suggested that Gly-Ile and Ile-Gly, which are the dipeptides of the present application, have no cytotoxicity and have the effect of enhancing the expression of the ceramide synthase CERS3 and CERS4 genes. Therefore, Gly-Ile and Ile-Gly, which are the dipeptides of the present application, can be expected to promote ceramide synthesis, which plays an important role in skin barrier function, at a total concentration of 0.00135 μg/mL or higher. In addition, preferably, for example, containing at least 0.00038 μg/mL Gly-Ile and at least 0.00097 μg/mL Ile-Gly, the total concentration of both dipeptides is 0.03 μg/mL. can be effective.
〔３．機能食品の製造〕
　次に、Gly-Ile及びIle-Glyで表されるジペプチドを肌質改善のための機能惹起成分、例えば機能性食品の場合における関与成分として含有させた機能食品の製造を行った。 [3. Manufacture of functional foods]
Next, a functional food containing dipeptides represented by Gly-Ile and Ile-Gly as a function-inducing ingredient for improving skin quality, eg, a participating ingredient in the case of a functional food, was produced.
　試験に供するための機能食品の剤形は、ハードカプセルとした。具体的には、200mgの粉末デキストリンを基材とし、ペプチド合成により得たGly-Ile及び／又はIle-Glyを添加し混合を行い、得られた混合粉体をゼラチンを皮膜原料としたカプセルのボディに充填しキャップを施しハードカプセル状の機能食品Ａを得た。 The dosage form of the functional food for testing was a hard capsule. Specifically, 200 mg of powdered dextrin is used as a base material, Gly-Ile and/or Ile-Gly obtained by peptide synthesis are added and mixed, and the obtained mixed powder is used as a capsule with gelatin as a shell material. The body was filled and capped to obtain a functional food A in the form of a hard capsule.
〔４．機能食品中のGly-Ile及びIle-Glyの測定〕
　次に、得られた機能食品Ａの錠剤１粒中に含まれる本願ジペプチドの含有量を測定した。 [4. Measurement of Gly-Ile and Ile-Gly in functional foods]
Next, the content of the dipeptide of the present application contained in one tablet of functional food A obtained was measured.
（４－１．２種のジペプチドの回収試験）
　まず、今回実施する試験方法が機能食品ＡのGly-IleやIle-Glyの含量を測定する上で妥当であるかを検討すべく、機能食品Ａを模して調製したサンプル食品中に予め添加した既知量のGly-Ile及びIle-Glyの回収率を確認する試験を行った。 (4-1.2 types of dipeptide recovery test)
First, in order to examine whether the test method to be carried out this time is appropriate for measuring the content of Gly-Ile and Ile-Gly in functional food A, it was added in advance to a sample food prepared to simulate functional food A. A test was performed to confirm the recovery of known amounts of Gly-Ile and Ile-Gly.
　混合粉体を含まないカプセル１粒に、２種類のペプチド標準品(Gly-Ile， Ile-Gly)混合溶液(500μg/mL)をそれぞれ20, 50, 100, 150μL(測定溶液中の終濃度：20, 50, 100, 150ng/mL)を加えた。その後、超純水を10mL加え、40分間、超音波抽出(38kHz、200W)を行った。なお、抽出操作中には抽出溶媒が均一分散するよう３～４回激しく振動させた。抽出後、遠心分離(3500rpm、10min)を行い、上清を25mLのメスフラスコに入れた。残渣に更に8mL、5mL超純水で上記と同様に２回超音波抽出、遠心分離を行った。３回の抽出液を合わせ、超純水で定容し、抽出溶液とした(n=3)。上記抽出溶液100μLに、内部標準溶液(2μg/mL)を20μL添加し、更に380μLのアセトニトリルを加え、撹拌し、遠心分離(3500rpm、10min)後、上清を0.20μm PTFE膜にてろ過した。ろ液は超純水で４倍希釈し、LC-MS分析に供した。添加した標準品の回収率の計算式を式1に示し、分析条件は表1に示す。
Figure JPOXMLDOC01-appb-M000001
Figure JPOXMLDOC01-appb-T000002
 20, 50, 100, 150 μL of a mixed solution (500 μg/mL) of two kinds of standard peptides (Gly-Ile, Ile-Gly) were added to one capsule containing no mixed powder (final concentration in the measurement solution: 20, 50, 100, 150ng/mL) were added. After that, 10 mL of ultrapure water was added, and ultrasonic extraction (38 kHz, 200 W) was performed for 40 minutes. During the extraction operation, vigorous shaking was performed 3 to 4 times to uniformly disperse the extraction solvent. After extraction, centrifugation (3500 rpm, 10 min) was performed and the supernatant was placed in a 25 mL volumetric flask. The residue was further subjected to ultrasonic extraction and centrifugation twice in the same manner as above with 8 mL and 5 mL of ultrapure water. Three extracts were combined, and the volume was adjusted with ultrapure water to obtain an extract solution (n=3). To 100 μL of the above extraction solution, 20 μL of internal standard solution (2 μg/mL) was added, 380 μL of acetonitrile was added, and the mixture was stirred, centrifuged (3500 rpm, 10 min), and the supernatant was filtered through a 0.20 μm PTFE membrane. The filtrate was diluted 4-fold with ultrapure water and subjected to LC-MS analysis. Formula 1 shows the formula for calculating the recovery rate of the added standard, and Table 1 shows the analysis conditions.
Figure JPOXMLDOC01-appb-M000001
Figure JPOXMLDOC01-appb-T000002
　LC-MS分析の結果、混合粉体を含まない硬カプセル（カプセルボディ及びキャップ）からは、２種類のジペプチドは検出されなかった。従って、カプセルには２種のジペプチドは含まれていないことが確認できた。 As a result of LC-MS analysis, the two types of dipeptides were not detected from the hard capsules (capsule body and cap) that did not contain mixed powder. Therefore, it was confirmed that the capsule did not contain the two dipeptides.
　次に、サンプル溶液の前処理方法や定量分析値の真度を検証するため、カプセルに２種類の既知量のジペプチドを標準品として添加したサンプル食品について分析を行い、添加回収率を算出した。表２に添加回収率を示す。
Figure JPOXMLDOC01-appb-T000003
 Next, in order to verify the pretreatment method of the sample solution and the accuracy of the quantitative analysis value, we analyzed the sample food in which two types of known amounts of dipeptide were added to the capsule as standard products, and calculated the addition recovery rate. Table 2 shows the addition recovery rate.
Figure JPOXMLDOC01-appb-T000003
　表２からも分かるように、標準品添加のサンプル食品は、いずれの添加量においても、２種のペプチドの平均回収率は97.8%～104.2%の範囲内であった。従って本実験の前処理方法および定量分析方法は妥当であると考えられる。 As can be seen from Table 2, the average recovery rate of the two peptides was within the range of 97.8% to 104.2% for the sample food with the addition of the standard product, regardless of the amount of addition. Therefore, the pretreatment method and quantitative analysis method of this experiment are considered appropriate.
（４－２．機能食品Ａに含まれる２種のジペプチドの定量分析）
　次に、機能食品Ａに含まれる２種のジペプチドの定量分析を行った。１粒の機能食品Ａに対して超純水を10mL加え、40分間、超音波抽出(38kHz、200W)を行った。なお、抽出操作中には抽出溶媒が均一分散するよう3～4回激しく振動させた。　　　 (4-2. Quantitative analysis of two dipeptides contained in functional food A)
Next, quantitative analysis of two dipeptides contained in functional food A was performed. 10 mL of ultrapure water was added to one grain of functional food A, and ultrasonic extraction (38 kHz, 200 W) was performed for 40 minutes. During the extraction operation, vigorous shaking was performed 3 to 4 times to uniformly disperse the extraction solvent.
　抽出操作後、遠心分離(3500rpm、10min)を行い、上清を25mLのメスフラスコに入れた。残渣に更に8mL、5mL超純水で上記と同様に超音波抽出、遠心分離を２回行った。計３回の抽出液を合わせ、超純水でで定容し、抽出溶液とした(n=3)。 After the extraction operation, centrifugation (3500 rpm, 10 min) was performed, and the supernatant was placed in a 25 mL volumetric flask. The residue was further subjected to ultrasonic extraction and centrifugation twice in the same manner as above with 8 mL and 5 mL of ultrapure water. A total of three extracts were combined, and the volume was adjusted with ultrapure water to obtain an extract solution (n=3).
　上記抽出溶液100μLに、内部標準溶液(2μg/mL)を20μL添加し、更に380μLのアセトニトリルを加え、撹拌し、遠心分離(3500rpm、10min)後、上清を0.20μm PTFE膜にてろ過した。ろ液は超純水で４倍希釈し、LC-MS分析に供した。分析条件は表１に示した通りである。 To 100 μL of the above extraction solution, 20 μL of internal standard solution (2 μg/mL) was added, 380 μL of acetonitrile was added, stirred, and after centrifugation (3500 rpm, 10 min), the supernatant was filtered through a 0.20 μm PTFE membrane. The filtrate was diluted 4-fold with ultrapure water and subjected to LC-MS analysis. Analysis conditions are as shown in Table 1.
　まず、標準品を用いた分析結果によりGly-IleおよびIle-Glyの溶出時間を確認した。また、表３に示すように、２種類のジペプチド標準品は5ng/mLから200ng/mLの検量線範囲において、良好な直線性を示した(R2=0.999)。
Figure JPOXMLDOC01-appb-T000004
 First, the elution times of Gly-Ile and Ile-Gly were confirmed by analysis results using standard products. Also, as shown in Table 3, the two dipeptide standards showed good linearity in the calibration curve range from 5 ng/mL to 200 ng/mL (R 2 =0.999).
Figure JPOXMLDOC01-appb-T000004
　さらに、検量線を用いた機能食品Ａのジペプチド含量を表４に示す。１粒の機能食品ＡにはGly-Ileが26.25±0.38μｇ、Ile-Glyが22.57±0.48μｇ含まれていることが確認された。
Figure JPOXMLDOC01-appb-T000005
 Furthermore, Table 4 shows the dipeptide content of functional food A using a calibration curve. It was confirmed that one grain of functional food A contained 26.25±0.38 μg of Gly-Ile and 22.57±0.48 μg of Ile-Gly.
Figure JPOXMLDOC01-appb-T000005
〔５．摂取試験〕
　次に、前述の〔３．機能食品の製造〕にて得られたカプセル状の機能食品Ａについて、健常成人の肌質の改善効果の検証を行った。 [5. Intake test]
Next, the aforementioned [3. production of functional food], the effect of improving the skin quality of healthy adults was verified.
　40～50歳の健康な一般女性20名を対象とし、試験品群10名、プラセボ群10名の２群に分けて試験を行った。摂取量は試験品群（機能食品Ａ）、プラセボ群（プラセボ食品）のいずれも１日２錠とした。なお、プラセボ食品にはGly-Ile及びIle-Glyは含まれていない。摂取期間はいずれも４週間とした。 A test was conducted on 20 healthy general women aged 40 to 50, divided into 2 groups: 10 test product groups and 10 placebo groups. The intake was 2 tablets per day for both the test product group (functional food A) and the placebo group (placebo food). The placebo food does not contain Gly-Ile and Ile-Gly. The ingestion period was 4 weeks.
　また、摂取期間経過後に肌質の測定を行った。具体的には、肌の水分量、水分蒸散量、粘弾性について測定を行った。なお、測定部位は左腕とした。 In addition, the skin quality was measured after the ingestion period. Specifically, the water content, water loss, and viscoelasticity of the skin were measured. The measurement site was the left arm.
　測定は、Cutometer DUAL MPA580（Courage + Khazaka electronic GmbH）を用い、水分、粘弾性は各５回ずつ測定し、最低値と最高値を除いた３回の平均を測定値とした。また、粘弾性の分析にはＲ２（総弾力）、Ｒ５（正味弾性）、Ｒ７（回復力）を使用した。 Measurements were made using a Cutometer DUAL MPA580 (Courage + Khazaka electronic GmbH), moisture content and viscoelasticity were measured five times each, and the average of three measurements excluding the lowest and highest values was taken as the measured value. R2 (total elasticity), R5 (net elasticity), and R7 (recovery force) were used for viscoelasticity analysis.
　標準誤差を算出し、事前測定と事後測定の比較（群内比較）及び群間比較のため、それぞれ対応あるｔ検定、対応のないｔ検定を行った。有意水準を５％とし、５％未満を有意差あり(p<0.05:**)、10％未満を有意傾向あり(p<0.1:*)と判断した。 　The standard error was calculated, and a paired t-test and an unpaired t-test were performed for comparison between pre-measurement and post-measurement (within-group comparison) and between-group comparisons. The significance level was set to 5%, and less than 5% was judged to have a significant difference (p<0.05:**), and less than 10% was judged to have a significant tendency (p<0.1:*).
　本試験に組み入れた20名中、事後測定に参加できなかった１名を除外した19名を解析対象者とした。具体的には、プラセボ群は女性10名で平均年齢は44.7歳、試験品群は女性9名で平均年齢は45.0歳であった。 Of the 20 people enrolled in this study, 19 people were included in the analysis after excluding one person who could not participate in the post-measurement. Specifically, there were 10 females with an average age of 44.7 years in the placebo group, and 9 females with an average age of 45.0 years in the test product group.
　図４は、水分量に関する試験結果を示している。水分量は角質層に含まれる水分量であり、肌の潤いの指標として参照される値である。図４（ａ）は、試験食品又はプラセボ食品の摂取前後における水分量測定値の変化を示しており、図４（ｂ）はプラセボ群と試験品群の水分量の変化量をそれぞれ示している。 Fig. 4 shows test results regarding water content. The water content is the amount of water contained in the stratum corneum, and is a value referred to as an index of skin moisture. FIG. 4(a) shows changes in water content measurements before and after ingestion of the test food or placebo food, and FIG. 4(b) shows changes in water content in the placebo group and the test product group, respectively. .
　図４（ａ）及び図４（ｂ）からも分かるように、試験品群には、摂取後の水分量の有意(p<0.05)な上昇が認められ、肌の乾燥しがちな季節環境においても、肌水分量が向上したことが確認された。 As can be seen from Figures 4(a) and 4(b), in the test product group, a significant (p<0.05) increase in water content after ingestion was observed, and in a seasonal environment where the skin tends to be dry. It was also confirmed that the skin moisture content was improved.
　また図示は割愛するが、同様の水分量測定を被験者の頬を対象に行ったところ、プラセボ群では、摂取前後で頬の水分量に有意な変化は認められなかったが、試験品群では有意な上昇が認められた(p<0.05)。 In addition, although the illustration is omitted, a similar water content measurement was performed on the subject's cheeks.In the placebo group, there was no significant change in the cheek water content before and after ingestion, but in the test product group, there was no significant change. significant increase was observed (p<0.05).
　図５は、経皮水分蒸散量に関する試験結果を示している。経皮水分蒸散量は、皮膚の表面から蒸発する水分量であり、バリア機能の指標となる値である。図５（ａ）は、試験食品又はプラセボ食品の摂取前後における経皮水分蒸散量の変化を示しており、図５（ｂ）はプラセボ群と試験品群の経皮水分蒸散量の変化量をそれぞれ示している。 Fig. 5 shows the test results regarding transepidermal water loss. The transepidermal water loss is the amount of water that evaporates from the surface of the skin, and is an indicator of the barrier function. Figure 5 (a) shows the change in transepidermal water loss before and after ingestion of the test food or placebo food, and Figure 5 (b) shows the change in the amount of transepidermal water loss between the placebo group and the test product group. each shown.
　図５（ａ）から分かるように、プラセボ群の経皮水分蒸散量は摂取後に有意に上昇したのに対し、試験品群では有意な変化は認められず、試験品の摂取により肌バリア機能が維持されていることが示された。また、図５（ｂ）からも分かるように、
試験品の摂取は、プラセボ品を摂取した場合に比して、経皮水分蒸散量の変化量を有意に抑制することが示された(p<0.05)。 As can be seen from Figure 5(a), the amount of percutaneous water loss in the placebo group increased significantly after ingestion, whereas no significant change was observed in the test product group. shown to be maintained. Also, as can be seen from FIG. 5(b),
Ingestion of the test product significantly suppressed changes in transepidermal water loss compared to ingestion of the placebo product (p<0.05).
　図６は、肌の総弾力（Ｒ２）に関する試験結果を示している。総弾力は、皮膚の全体の弾力を表す指標である。皮膚弾力性は、真皮層の繊維質や保水成分により維持されている。図６（ａ）は、試験食品又はプラセボ食品の摂取前後における総弾力の変化を示しており、図６（ｂ）はプラセボ群と試験品群の総弾力の変化量をそれぞれ示している。 Fig. 6 shows the test results regarding the total skin elasticity (R2). Total elasticity is an index representing the overall elasticity of the skin. Skin elasticity is maintained by the fiber and water-retaining components of the dermis layer. FIG. 6(a) shows changes in total elasticity before and after ingestion of the test food or placebo food, and FIG. 6(b) shows changes in total elasticity of the placebo group and the test article group, respectively.
　図６（ａ）及び図６（ｂ）からも分かるように、総弾力を示すＲ２では、プラセボ群において減少傾向を示し、また、時間と群の二要因の有意な交互作用が認められ、プラセボ群と試験品群の変化量には有意な群間差が示された。このことより、Gly-Ile及びIle-Glyは、肌の総弾力（Ｒ２）に関し、真皮層の機能を活性化させ、肌のハリや弾力性を維持する効果があるといえる As can be seen from Figures 6 (a) and 6 (b), R2, which indicates total elasticity, showed a decreasing trend in the placebo group, and a significant interaction between the two factors of time and group was observed. A significant inter-group difference was shown in the amount of change between the group and the test article group. From this, it can be said that Gly-Ile and Ile-Gly have the effect of activating the function of the dermis layer and maintaining the firmness and elasticity of the skin with respect to the total elasticity (R2) of the skin.
　図７は、肌の粘弾性（Ｒ５（正味弾性））に関する試験結果を示している。粘弾性は、皮膚のハリや弾力性の指標となる値である。図７（ａ）は、試験食品又はプラセボ食品の摂取前後における粘弾性の変化を示しており、図７（ｂ）はプラセボ群と試験品群の粘弾性の変化量をそれぞれ示している。　　　 Fig. 7 shows the test results regarding skin viscoelasticity (R5 (net elasticity)). Viscoelasticity is a value that serves as an index of skin firmness and elasticity. FIG. 7(a) shows changes in viscoelasticity before and after ingestion of the test food or placebo food, and FIG. 7(b) shows changes in viscoelasticity between the placebo group and the test product group, respectively.　　　
　図７（ａ）及び図７（ｂ）からも分かるように、試験品群では正味弾力性を示すＲ５が摂取前と比較して有意(p<0.05)に上昇し、試験品の摂取は肌の弾力性を向上させることが示された。また、時間と群の二要因の有意傾向の交互作用が認められ、変化量においても有意傾向の群間差が示された。 As can be seen from Figures 7(a) and 7(b), in the test product group, R5, which indicates net elasticity, increased significantly (p<0.05) compared to before ingestion. was shown to improve the elasticity of In addition, a significant trend interaction between the two factors of time and group was observed, and a significant trend difference between the groups was also shown in the amount of change.
　図８は、肌の回復力Ｒ７（戻り率）に関する試験結果を示している。回復力は、吸引時の皮膚の伸びた長さに対する瞬間的な収縮率を表す値であり、肌のハリやたるみと高い相関を示す。図８（ａ）は、試験食品又はプラセボ食品の摂取前後における回復力の変化を示しており、図８（ｂ）はプラセボ群と試験品群の回復力の変化量をそれぞれ示している。 Fig. 8 shows test results regarding skin recovery power R7 (return rate). The resilience is a value representing the instantaneous contraction rate with respect to the stretched length of the skin at the time of suction, and shows a high correlation with firmness and sagging of the skin. FIG. 8(a) shows changes in resilience before and after ingestion of the test food or placebo food, and FIG. 8(b) shows changes in resilience between the placebo group and the test article group, respectively.
　図８（ａ）及び図８（ｂ）からも分かるように、試験品群では回復力Ｒ７が摂取前と比較して有意(p<0.05)に上昇し、試験品の摂取は肌の回復力を向上させることが示された。また、回復力Ｒ７においても時間と群の二要因の有意な交互作用が認められ、変化量においても有意傾向の群間差が示された。 As can be seen from FIGS. 8(a) and 8(b), in the test product group, the resilience R7 increased significantly (p<0.05) compared to before ingestion, and the ingestion of the test product increased skin resilience. was shown to improve In addition, a significant interaction between two factors, time and group, was observed in resilience R7, and a significant inter-group difference was shown in the amount of change.
　これら各摂取試験の結果を踏まえると、Gly-Ile及びIle-Glyで表されるジペプチドを肌質改善のための機能惹起成分、例えば機能性食品の場合における関与成分として含有させた機能食品は、肌水分の保持や肌水分の蒸散抑制、肌のハリや粘弾性の維持・向上に寄与し、肌質を改善できることが示された。 Based on the results of these intake tests, functional foods containing dipeptides represented by Gly-Ile and Ile-Gly as function-inducing ingredients for improving skin quality, for example, as participating ingredients in the case of functional foods, It was shown that it contributes to retaining skin moisture, suppressing evaporation of skin moisture, maintaining and improving skin firmness and viscoelasticity, and improving skin quality.
　上述してきたように、本実施形態に係るセラミド合成酵素遺伝子発現促進剤によれば、Gly-Ile及び／又はIle-Glyで表されるジペプチドを機能惹起のための主要成分として含有することとしたため、機能惹起のための主要成分として所定のジペプチドが採用され、セラミド合成酵素遺伝子の発現量を増大させることが可能なセラミド合成酵素遺伝子発現促進剤を提供することができる。 As described above, the ceramide synthase gene expression promoter according to the present embodiment contains a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function. It is possible to provide a ceramide synthase gene expression-promoting agent that employs a predetermined dipeptide as a main component for inducing function and that is capable of increasing the expression level of the ceramide synthase gene.
　また、本実施形態に係る肌質改善剤や肌質改善用機能食品、化粧料では、Gly-Ile及び／又はIle-Glyで表されるジペプチドを機能惹起のための主要成分として含有することとしたため、セラミド合成酵素遺伝子の発現量を増大させ、肌質改善を図ることが可能な肌質改善剤や肌質改善用機能食品、化粧料とすることができる。 In addition, the skin quality improving agent, skin quality improving functional food, and cosmetics according to the present embodiment contain a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function. Therefore, it is possible to increase the expression level of the ceramide synthase gene, and use it as a skin quality improving agent, a functional food for improving skin quality, or a cosmetic that can improve skin quality.
　最後に、上述した各実施の形態の説明は本発明の一例であり、本発明は上述の実施の形態に限定されることはない。このため、上述した各実施の形態以外であっても、本発明に係る技術的思想を逸脱しない範囲であれば、設計等に応じて種々の変更が可能であることは勿論である。  Finally, the description of each embodiment described above is an example of the present invention, and the present invention is not limited to the above-described embodiments. Therefore, it goes without saying that various modifications other than the above-described embodiments can be made in accordance with the design and the like within the scope not departing from the technical idea of the present invention. 

Claims (7)

  1. 　Gly-Ile及び／又はIle-Glyで表されるジペプチドを機能惹起のための主要成分として含有するセラミド合成酵素遺伝子発現促進剤。 A ceramide synthase gene expression promoter containing a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function.
  2. 　前記セラミド合成酵素遺伝子は、セラミド合成酵素３(CERS3)遺伝子であることを特徴とする請求項１に記載のセラミド合成酵素遺伝子発現促進剤。 The ceramide synthase gene expression promoter according to claim 1, wherein the ceramide synthase gene is a ceramide synthase 3 (CERS3) gene.
  3. 　Gly-Ile及び／又はIle-Glyで表されるジペプチドを機能惹起のための主要成分として含有する肌質改善剤。 A skin quality improving agent containing a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function.
  4. 　Gly-Ile及び／又はIle-Glyで表されるジペプチドを関与成分として含有する肌質改善用機能食品。 A functional food for improving skin quality containing dipeptides represented by Gly-Ile and/or Ile-Gly as participating ingredients.
  5. 　Gly-Ile及び／又はIle-Glyで表されるジペプチドを有効成分として含有する化粧料。 A cosmetic containing a dipeptide represented by Gly-Ile and/or Ile-Gly as an active ingredient.
  6. 　Gly-Ile及び／又はIle-Glyで表されるジペプチドのセラミド合成酵素遺伝子発現促進剤における機能惹起のための主要成分としての使用。 Use of a dipeptide represented by Gly-Ile and/or Ile-Gly as a main component for inducing function in a ceramide synthase gene expression promoter.
  7. 　Gly-Ile及び／又はIle-Glyで表されるジペプチドの機能食品における肌質改善を目的とした関与成分としての使用。 Use of dipeptides represented by Gly-Ile and/or Ile-Gly as a participating component in functional foods for the purpose of improving skin quality.
PCT/JP2022/004425 2021-02-05 2022-02-04 Ceramide synthase gene expression promoting agent, skin improving agent, functional food for skin improvement, cosmetic, and usage method for dipeptide WO2022168939A1 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP2021-017246 2021-02-05
JP2021017246A JP2022120386A (en) 2021-02-05 2021-02-05 Ceramide synthase gene expression promoting agent, skin improving agent, functional food for skin improvement, cosmetic, and usage method for dipeptide

Publications (1)

Publication Number Publication Date
WO2022168939A1 true WO2022168939A1 (en) 2022-08-11

Family

ID=82741576

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/JP2022/004425 WO2022168939A1 (en) 2021-02-05 2022-02-04 Ceramide synthase gene expression promoting agent, skin improving agent, functional food for skin improvement, cosmetic, and usage method for dipeptide

Country Status (3)

Country Link
JP (1) JP2022120386A (en)
TW (1) TW202246302A (en)
WO (1) WO2022168939A1 (en)

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2023046102A (en) * 2021-09-22 2023-04-03 国立大学法人九州大学 Expression promotor and functional food for mental health enhancement

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2011126163A1 (en) * 2010-04-08 2011-10-13 주식회사 웰스킨 Skin-whitening composition containing dipeptide
JP2012522043A (en) * 2009-03-31 2012-09-20 ウェルスキン カンパニー，リミテッド Composition for inhibiting erythema reaction by ultraviolet rays containing dipeptide as active ingredient

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2012522043A (en) * 2009-03-31 2012-09-20 ウェルスキン カンパニー，リミテッド Composition for inhibiting erythema reaction by ultraviolet rays containing dipeptide as active ingredient
WO2011126163A1 (en) * 2010-04-08 2011-10-13 주식회사 웰스킨 Skin-whitening composition containing dipeptide

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
NAGAE MASUMI, ET AL: "Effect of Porcine Placenta Extract Supplement on Skin Condition in Healthy Adult Women: A Randomized, Double-Blind Placebo-Controlled Study", NUTRIENTS, vol. 12, no. 6, pages 1671, XP055957223, DOI: 10.3390/nu12061671 *
PARK JONG IL, ET AL: "Oral Administration of Glycine and Leucine Dipeptides Improves Skin Hydration and Elasticity in UVB-Irradiated Hairless Mice", BIOMOLECULES & THERAPEUTICS, vol. 25, no. 5, 1 September 2017 (2017-09-01), pages 528 - 534, XP055957225, ISSN: 2005-4483, DOI: 10.4062/biomolther.2017.089 *

Also Published As

Publication number Publication date
JP2022120386A (en) 2022-08-18
TW202246302A (en) 2022-12-01

Similar Documents

Publication Publication Date Title
KR100952109B1 (en) Personal care compositions comprising a dipeptide
JPWO2004016236A1 (en) Cosmetics
WO2003072039A2 (en) Cosmetic formulations containing l-arginine oligomers
BRPI0816214B1 (en) cosmetic use, cosmetic and / or dermatological composition, and methods for the cosmetic treatment of the skin, for the cosmetic treatment of skin signs of aging and / or photoaging and for the cosmetic treatment to prevent and / or treat dry keratinous substances.
US20080153839A1 (en) Use of anisic acid for promoting desquamation
JP2022511023A (en) New cosmetic and dermatological uses of extracts of Cistus monsperiensis
MX2014009308A (en) Use of cpt-1 modulators and compositions thereof.
WO2022168939A1 (en) Ceramide synthase gene expression promoting agent, skin improving agent, functional food for skin improvement, cosmetic, and usage method for dipeptide
KR101929657B1 (en) Skin external agent for improving skin wrinkle comprising an extract of fermented wheat germ
US20100317736A1 (en) Composition of the skin external application or the food for accelerating proline recycling by containing theanine
JP2006290873A (en) Aquaporin expression promoter
JP4088497B2 (en) Cosmetics
TWI774046B (en) Use of solanum muricatum fermented liquid for preparing a composition for promoting skin anti-aging ability, promoting anti-glycation ability, reducing melanin content, increasing skin moisturization, reducing skin texture, reducing skin wrinkles, reducing skin redness and/or reducing fat
TW202118508A (en) Polypeptides having anti-senescent effects and uses thereof
CN105943414B (en) Cosmetic composition comprising cultured root oil of wild ginseng as active ingredient, method for preparing the same and cosmetic method using the same
WO2022094437A1 (en) Polypeptides having anti-inflammatory effects and uses thereof
WO2018197923A1 (en) Association and methods for fighting against the signs of ageing
JP6089163B1 (en) Beauty composition
JP2021187787A (en) Exosome secretagogue, exosome uptake promoter, exosome secretion-promoting beauty composition, and exosome secretion promoting method
JP2021004237A (en) Topical compositions containing n-acyl dipeptide derivatives and glycolic acid
EP3366273A1 (en) Moisturizer and cosmetic containing same
US20220378676A1 (en) Composition for enhancing skin elasticity or improving skin wrinkles
WO2023191015A1 (en) Collagen production promoter, cosmetic composition or external preparation for skin, and preparation for oral administration
JP5527917B2 (en) Arginase activity promoter and skin external preparation containing the same
KR20180108253A (en) Composition for skin improvement containing liquiritin apioside

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 22749810

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 22749810

Country of ref document: EP

Kind code of ref document: A1