KR102281606B1 - A multifunctional cosmetic composition for elasticity, anti-wrinkle, inhibiting tyrosinase comprising peptide complex - Google Patents

Abstract

The present invention relates to a multifunctional cosmetic composition comprising tetrapeptide-21 and palmitoyl pentapeptide-4 as active ingredients.

Description

Multifunctional cosmetic composition containing peptide complex for improving wrinkles and skin elasticity, inhibiting tyrosinase, etc. {A MULTIFUNCTIONAL COSMETIC COMPOSITION FOR ELASTICITY, ANTI-WRINKLE, INHIBITING TYROSINASE COMPRISING PEPTIDE COMPLEX}

The present invention relates to a multifunctional cosmetic composition comprising a low molecular weight peptide complex as an active ingredient.

The skin is a membrane that covers the outside of the body and performs various physiological functions to protect the body from environmental factors such as various external stimuli, obstacles, and dryness, thereby protecting and regulating internal organs and other internal organs. do.

However, due to aging in the living body, the secretion of various hormones that control metabolism internally is reduced, and the function of immune cells and the activity of cells are reduced.

In addition, externally, the UV content in the sun's rays increases due to the destruction of the ozone layer, and excessive physical and chemical stimulation and stress caused by the increase of free radicals and active harmful oxygen as the environmental pollution is further deepened is the normal function of the skin. It can damage the skin by promoting the generation of spots, freckles, etc. and skin aging caused by deterioration, melanin deposition.

Our body performs a skin regeneration process on damaged skin. Skin regeneration is a tissue response to damage and is a process of tissue repair, such as chemotaxis, cell differentiation and replication, matrix protein synthesis, vascularization, and wound reconstitution. It is a complex biological process involving

Therefore, in order to prevent skin aging and damage caused by internal and external factors to maintain healthy and beautiful skin, physiologically active substances obtained from various animals, plants, and microorganisms are added to cosmetics and used.

Efforts to maintain skin health by maintaining the unique function of the skin and activating skin cells through various physiologically active substances as described above and promoting skin metabolism are continuing, and cosmetics and external preparations for skin are being developed accordingly.

However, when these substances are applied to the skin, there is a limitation in the amount used due to safety problems such as irritation, erythema, redness, or the like, or the effect is insignificant, so it is difficult to actually expect an effect of improving skin function.

Therefore, there is a demand for the development of a new composition for external application for skin that is safer and more effective than the existing composition for external application for skin.

Recently, in the cosmetic industry, a number of products using natural products have been developed to reduce skin irritation caused by chemicals.

Natural ingredients have fewer side effects on the skin, and as consumers' response to cosmetics using natural ingredients increases, the value of development as a cosmetic raw material is increasing.

Therefore, the demand for cosmetics using natural ingredients with few side effects and high consumer response is increasing, and various studies on the development of multi-functional cosmetics with excellent biosafety and skin improvement effects are being actively conducted. there is.

An object of the present invention is to solve the problems of the prior art described above, and an object of the present invention is to provide a functional cosmetic composition that has little irritation to the skin and minimizes skin damage caused by external stimuli.

According to one aspect of the present invention, there is provided a cosmetic composition comprising tetrapeptide-21 (Tetrapeptide-21) and palmitoyl pentapeptide-4 (Palmitoyl Pentapeptide-4) as active ingredients.

In one embodiment, the cosmetic composition may further include one or more of hexapeptide-11 and carnosine.

In one embodiment, the cosmetic composition may further include one or more selected from the group consisting of adenosine, zymoroot extract, and glutathione.

In one embodiment, the cosmetic composition may be for wrinkle improvement and skin elasticity improvement.

In one embodiment, the cosmetic composition may be for skin whitening.

In one embodiment, the cosmetic composition may be for anti-inflammatory or antioxidant.

In one embodiment, the cosmetic composition may be for skin regeneration.

In one embodiment, the cosmetic composition is selected from the group consisting of softening lotion, nourishing lotion, nourishing cream, massage cream, essence, ampoule, gel, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray and powder. One or more may be formulated.

The cosmetic composition according to one aspect of the present invention has less skin irritation and is remarkably excellent in skin improvement effect due to the synergistic effect of each active ingredient.

It should be understood that the effects of the present invention are not limited to the above-described effects, and include all effects that can be inferred from the configuration of the invention described in the detailed description or claims of the present invention.

The terms used in this specification have been selected as currently widely used general terms as possible while considering the functions in the present invention, which may vary depending on the intention or precedent of a person skilled in the art, the emergence of new technology, and the like. In addition, in a specific case, there is a term arbitrarily selected by the applicant, and in this case, the meaning will be described in detail in the description of the corresponding invention. Therefore, the term used in the present invention should be defined based on the meaning of the term and the overall content of the present invention, rather than the name of a simple term.

Unless defined otherwise, all terms used herein, including technical and scientific terms, have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Terms such as those defined in a commonly used dictionary should be interpreted as having a meaning consistent with the meaning in the context of the related art, and should not be interpreted in an ideal or excessively formal meaning unless explicitly defined in the present application. does not

Numerical ranges are inclusive of the values defined in that range. Every maximum numerical limitation given throughout this specification includes all lower numerical limitations as if the lower numerical limitation were expressly written. Every minimum numerical limitation given throughout this specification includes all higher numerical limitations as if the higher numerical limitation were expressly written. All numerical limitations given throughout this specification shall include all numerical ranges that are better within the broader numerical limits, as if the narrower numerical limitations were expressly written.

Hereinafter, examples of the present invention will be described in detail, but it is obvious that the present invention is not limited by the following examples.

One aspect of the present invention provides a cosmetic composition comprising tetrapeptide-21 (Tetrapeptide-21) and palmitoyl pentapeptide-4 (Palmitoyl Pentapeptide-4) as active ingredients.

The cosmetic composition contains a low-molecular complex peptide component as an active ingredient, so that it has excellent biosafety and skin improvement activity as well as less skin irritation.

The peptide refers to a polymer composed of one or more amino acids linked by amide bonds (or peptide bonds). For the purpose of the present invention, the peptide may refer to a peptide or a fragment thereof that exhibits wrinkle improvement and skin elasticity improvement effects.

A general rule for naming such peptides may be based on a three-letter or one-letter amino acid code, unless specifically indicated otherwise. For example, the central part of the amino acid structure is indicated by a three-letter code (eg, Ala, Lys), and a D-conformation (eg, D-Ala, D-Lys) is specified by writing "D-" in front of the three-letter code. If not indicated, it can be assumed to have an L-stereoscopic shape. The amino acid residues constituting the peptide may be natural or non-natural amino acid residues.

The present inventors have identified that the peptide can effectively inhibit skin damage and skin aging that may be caused by various factors by interacting with other components, and various uses have been derived through this.

The peptide can be prepared according to chemical synthesis methods known in the art, particularly solid-phase synthesis techniques (Merrifield, J. Amer. Chem. 1963, Soc. 85:2149-54; Stewart, et al. Solid Phase Peptide Synthesis, 2nd. ed., Pierce Chem. 1984, Co.: Rockford, 111).

The "comprising as an active ingredient" may mean containing an effective amount that can exhibit a skin improvement effect, for example, wrinkle improvement, skin elasticity improvement, skin whitening, antioxidant effect, and the like.

The “Tetrapeptide-21” is a peptide in which glycine, glutamic acid, lysine, and glycine are sequentially linked, and may be a compound of Formula 1 below.

[Formula 1]

The "Palmitoyl Pentapeptide-4" is a lipophilic fatty acid palmitate at the amino terminal site of a peptide in which lysine, threonine, threonine, lysine, serine, and serine are sequentially linked. It may be a compound of Formula 2 below.

[Formula 2]

The cosmetic composition may further include one or more of hexapeptide-11 and carnosine.

The “hexapeptide-11” is a synthetic peptide in which alanine, phenylalanine, proline, and valine are sequentially linked, and may be a compound of Formula 3 below.

[Formula 3]

The carnosine is a dipeptide of β-alanyl-L-histidine composed of β-alanin and histidine, and is found in a fairly large amount in long-lived tissues, particularly in the spine. It is abundantly present in the non-protein fraction of animal muscle cells, and its amount ranges from about 1 to 20 mM.

The carnosine is known as an antioxidant that protects cells from oxidative stress, and is a free radical scavenger having an excellent effect in removing reactive oxygen species.

The protective action of canosine is achieved by inhibiting the transformation of biopolymers in an oxidative stress environment through proton buffering, heavy metal chelating, and removal of free radicals and active sugar molecules and maintaining their intrinsic function. and the canosine acts together with the low molecular weight complex peptide, thereby exhibiting an excellent synergistic effect.

The cosmetic composition may further include one or more selected from the group consisting of adenosine, zymoroot extract, and glutathione.

The “Adenosine” is a nucleotide based on adenine and is declared as a functional raw material. The adenosine is a nucleoside in which adenine is connected to a part of ribose (ribofuranose) by a β-glycosidic bond. The adenosine may contribute to biochemical action in the form of triphosphate and diphosphate.

The above "Zemo (知母, Anemarrhenaasphodeloides Bunge)" is a perennial plant of the monocotyledonous Liliaceae family, and uses the rhizome of the Zymoaceae family, the origin is China, and is distributed evenly in Hwanghae Province in Korea.

The leaves are gathered and grown at the root of the genus, and the leaves are divided into exogenous, normal, and endogenous from the outer part of the leaf. The normal leaf in the middle is the largest, and the shape of the leaf is long narrow lanceolate, and the tip is quite sharp.

The roots and stems of the jimo contain timosaponin, chimonin, prococathechi acid, pantothenic acid, etc. It is reported to have antibacterial, expectorant, sedative, hypoglycemic, and anticancer effects.

The “glutathione” is a compound having the structure of Formula 4 below, and may have a Cas number of 70-18-8 and a molecular weight of 307.32.

[Formula 4]

The glutathione as an antioxidant present in the living body can effectively remove radicals in the living body by reversibly reacting with in vivo antioxidants such as ascorbic acid (vitamin C) and tocopherol (vitamin E).

The glutathione may play a very important role as a final radical receptor in a biological system related to the radical removal.

The cosmetic composition may be for wrinkle improvement and skin elasticity improvement.

The "improvement of skin elasticity" refers to a phenomenon in which elastic fibers composed of elastin exist together with collagen, and skin elasticity is maintained or improved in a state in which the elastin and collagen are sufficiently present.

In addition, the skin elasticity improvement effect may be accompanied by a skin wrinkle improvement effect.

The "wrinkle improvement" refers to a phenomenon in which wrinkles are inhibited or inhibited from being generated on the skin, or wrinkles that have already been created are alleviated.

In addition, the cosmetic composition may be for skin whitening.

The "skin whitening" suppresses the production of various biomolecules such as melanin, oxidation-reduction hemoglobin, carotene, and melanoids that affect skin color, thereby alleviating skin pigmentation phenomena such as blemishes, freckles, and blemishes, skin yellowness, It refers to the effect of improving skin brightness and uniformity by alleviating redness.

The cosmetic composition may be for anti-inflammatory or antioxidant.

The “antioxidation” refers to the action of inhibiting oxidation, and the human body has a balance of oxidative and antioxidant substances. However, due to various factors, this balance is lost and when the body tilts toward accelerating oxidation, , oxidative stress is induced in the living body, which may cause cell damage and pathological diseases.

Reactive oxygen species (ROS), which is a direct cause of oxidative stress, is chemically unstable and highly reactive, so it can easily react with various biomaterials such as DNA, proteins, lipids and carbohydrates, It attacks and causes irreversible damage to cells and tissues or causes mutations, cytotoxicity and cancer, and is also a direct cause of aging.

The cosmetic composition can prevent aging and maintain health by obtaining an antioxidant effect by removing or reducing the active oxygen species.

The “anti-inflammatory” refers to the action of inhibiting inflammation, and the regulation of the inflammatory response is known to be very complex, which reduces the enhancement and damage of the repair system in vivo.

If the inflammatory response continues due to repeated tissue damage or regeneration, ROS and RNS are overproduced in inflammation-related cells, and permanent gene modification may be caused. That is, ROS and RNS are deeply related to the inflammatory response that regulates the actions of various cells in vivo.

The cosmetic composition may exhibit excellent anti-inflammatory activity by regulating the expression of inflammatory mediators generated due to the activation of NF-kB.

In addition, the cosmetic composition may be for skin regeneration.

The “skin regeneration” refers to a process of restoring skin tissue from damage caused by external and internal causes of the skin.

The damage caused by the external cause may include ultraviolet rays, external pollutants, wounds, trauma, and the like, and the damage caused by the internal cause may include stress.

The cosmetic composition can restore the skin barrier, promote skin regeneration, and increase moisture to protect the skin from UV rays or pollution, increase immunity, and inhibit skin aging.

The cosmetic composition may be formulated with one or more selected from the group consisting of softening lotion, nutrient lotion, nutrient cream, massage cream, essence, ampoule, gel, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray and powder. can

The cosmetic composition may be formulated by a conventional method. In the formulation of external skin preparations, reference may be made to the contents disclosed in Remington's Pharmaceutical Science, Mack Publishing Company, Easton PA, and in the formulation of cosmetic compositions, the International cosmetic ingredient dictionary, 6th ed (The cosmetic, Toiletry and Fragrance Association) , Inc., Washington, 1995) may be referred to.

Specifically, the cosmetic composition can be prepared in general emulsified formulations and solubilized formulations. For example, lotion, such as a softening lotion or a nourishing lotion; emulsions such as facial lotions and body lotions; creams such as nourishing creams, moisturizing creams, and eye creams; essence; makeup ointment; spray; gel; pack; sunscreen; makeup base; foundations such as liquid type, solid type or spray type; powder; makeup removers such as cleansing creams, cleansing lotions, and cleansing oils; Or it may be formulated as a cleaning agent such as a cleansing foam, soap, body wash, but is not limited thereto. In addition, the external preparation for skin may be formulated as an ointment, patch, gel, cream or spray, but is not limited thereto.

In the cosmetic composition, in addition to the essential components in each formulation, other components may be appropriately blended within the range that does not impair the purpose according to the present invention, depending on the type or purpose of use, etc. of the formulation.

The cosmetic composition may include a conventionally acceptable carrier, for example, oil, water, surfactant, humectant, lower alcohol, thickener, chelating agent, colorant, preservative, fragrance, etc. may be appropriately mixed, but is limited thereto no.

The acceptable carrier may vary depending on the formulation. For example, when formulated into an ointment, paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tracanth, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide or their Mixtures can be used

When the cosmetic composition is formulated as a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate, polyamide powder, or mixtures thereof may be used as a carrier component, and in the case of a spray, chlorofluorohydro It may further contain a propellant such as carbon, propane, butane or dimethyl ether.

When the cosmetic composition is formulated as a solution or emulsion, a solvent, solubilizer, or emulsifier may be used as a carrier component, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl benzoate, propylene glycol, 1,3-butylglycol oil may be used.

When the cosmetic composition is formulated as a suspension, a liquid diluent such as water, ethanol or propylene glycol as a carrier component, a suspending agent such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester; Microcrystalline cellulose, aluminum metahydroxide, bentonite, agar or tracanth and the like can be used.

The cosmetic composition may contain fatty substances, organic solvents, solubilizers, thickeners, gelling agents, softeners, antioxidants, suspending agents, stabilizers, foaming agents, and fragrances commonly used in the industry depending on the quality or function of the final product. , surfactants, water, ionic or nonionic emulsifiers, fillers, sequestering agents, chelating agents, preservatives, blocking agents, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic active agents, commonly used in cosmetics It may additionally contain adjuvants commonly used in cosmetic or dermatological fields, such as any other ingredients.

However, the adjuvant and its mixing ratio may be appropriately selected so as not to affect the desirable properties of the cosmetic composition according to the present invention.

Through the following examples, the present invention will be described in more detail, but it is obvious that the present invention is not limited by the following examples.

Examples and Comparative Examples

In order to verify the stability and functionality of the peptide complex of the present invention, samples were mixed as shown in Tables 1 and 2 below to set Examples and Comparative Examples. Each sample was obtained from the market and used.

[Content (parts by weight)] division Example One 2 3 4 5 6 7 8 tetrapeptide-21 60 40 40 30 24 24 24 24 Palmitoyl Pentapeptide-4 60 40 40 30 24 24 24 24 Hexapeptide-11 - 40 - 30 24 24 24 24 carnosine - - 40 30 24 24 24 24 adenosine - - - - 24 - - 8 Zymoji root extract - - - - - 24 - 8 glutathione - - - - - - 24 8

[Content (parts by weight)] division comparative example One 2 3 4 5 6 7 8 9 10 11 12 tetrapeptide-21 120 - - - - - - 60 60 40 30 - Palmitoyl Pentapeptide-4 - 120 - - - - - - - - - - Hexapeptide-11 - - 120 - - - - 60 - 40 - - carnosine - - - 120 - - - - 60 40 - -- adenosine - - - - 120 - - - - - 30 40 Zymoji root extract - - - - - 120 - - - - 30 40 glutathione - - - - - - 120 - - - 30 40

Experimental Example 1: Skin safety test

In order to verify the skin safety of the samples of Examples and Comparative Examples, an MTT assay experiment for fibroblasts (HDF) was performed.

After each of the samples of Examples and Comparative Examples were suspended in purified water at concentrations of 25, 50, 100, 200, and 400 μg/mL, cell viability was measured.

Cytotoxicity was performed by modifying Mosmann's method for measuring cell viability using MTT {3-(4,5-dimethylthiazol-2-yl)-2-5-diphenyltetrazolium bromide} reagent.

Fibroblasts (HDF) were each dispensed in a 96-well plate at ^{a concentration of 1×10 4} cells/well and cultured at 37° C., 5% CO ₂ for 48 hours.

After removing the culture medium and culturing the sample in a medium treated for each concentration for 48 hours, the medium was removed and washed repeatedly with PBS (Phosphate buffered saline).

MTT was dissolved in PBS at 5 mg/mL, 50 μL was added, and incubated at 37° C., 5% CO ₂ for 48 hours. 100 L of DMSO (dimethyl sulfoxide) was added per well, and the absorbance was measured at 540 nm after stirring for 10 minutes.

As a result of the measurement, cytotoxicity was not confirmed at all concentrations of each sample. The above results suggest that the sample is not only harmless to the human body, but also has excellent human safety.

Experimental Example 2: Evaluation of free radical scavenging activity

The extracts of Examples and Comparative Examples were suspended in purified water to evaluate free radical scavenging activity.

The DPPH measurement method is a method of measuring the absorbance at 540 nm to determine the extent to which an inhibitor is decolorized by scavenging a stable radical DPPH (2,2-diphenyl-1-picrylhydrazyl radical).

Vitamin C was used as a control, and the same experiment was repeated 3 times for the accuracy of the experiment, and the results are shown in Table 3 below.

[Concentration that scavenges 50% of radical DPPH] division SC ₅₀ (μg/mL) Example 1 129.7 Example 2 119.2 Example 3 120.7 Example 4 114.9 Example 5 110.9 Example 6 110.3 Example 7 111.7 Example 8 106.8 Comparative Example 1 219.3 Comparative Example 2 190.5 Comparative Example 3 178.4 Comparative Example 4 177.2 Comparative Example 5 174.2 Comparative Example 6 178.4 Comparative Example 7 178.4 Comparative Example 8 174.2 Comparative Example 9 180.1 Comparative Example 10 174.2 Comparative Example 11 178.4 Comparative Example 12 178.4 Control (vitamin C) 124.5

The lower the SC ₅₀ value, the higher the scavenging activity of DPPH free radicals. Referring to Table 3, the samples of Examples showed significantly better skin elasticity improvement effect than those of Comparative Examples.

In particular, Example 1 containing tetrapeptide-21 and palmitoyl pentapeptide-4 at the same time had significantly superior free radical scavenging activity compared to Comparative Examples 8 to 11 lacking some, and the results were suggest a synergistic effect.

Experimental Example 3: Evaluation of anti-inflammatory activity

The anti-inflammatory activity of Examples and Comparative Examples was evaluated by analyzing the NO production inhibitory ability of EAME in RAW 264.7 cell, a mouse macrophage cell line induced by lipopolysaccharide (LPS) stimulation.

For the experiment, Raw 264.7 cells, a murine macrophage cell line, were distributed from the Korean cell line bank.

Using DMEM medium containing 100 units/mL penicillin-streptomycin and 10% fetal bovine serum (FBS), it _{was cultured in an incubator at 37° C., 5% CO 2} , and subcultured once every 2-3 days.

Raw 264.7 cells (3 x 10 ⁵ cells/mL) were cultured 18 hours before, and samples of Examples and Comparative Examples (concentration 1%) and LPS (1 μg/mL) were simultaneously treated and cultured for 24 hours.

The generated NO was measured in the form of _{NO 2} ⁻ present in the cell culture medium using Griess reagent.

The negative control group was not treated with LPS, and the positive control group was treated with only LPS to measure the amount of NO generated.

100 μL of cell culture supernatant and Griess reagent (1% (w/v) sulfanilamide, 0.1% (w/v) naphtylehtylenediamine in 2.5% (v/v) phosphoric acid) were mixed in equal amounts and reacted in the dark at room temperature for 10 minutes, followed by ELISA Absorbance (540 nm) was measured using a reader.

The standard concentration curve was _{calculated by serial dilution of sodium nitrite (NaNO 2} ) (10-100 μM). The measurement results are shown in Table 4 below.

division NO production rate (%) Example 1 47.1 Example 2 44.8 Example 3 43.2 Example 4 41.4 Example 5 37.2 Example 6 38.5 Example 7 39.1 Example 8 35.9 Comparative Example 1 73.0 Comparative Example 2 76.3 Comparative Example 3 72.7 Comparative Example 4 68.4 Comparative Example 5 83.0 Comparative Example 6 67.6 Comparative Example 7 73.0 Comparative Example 8 67.8 Comparative Example 9 68.6 Comparative Example 10 83.0 Comparative Example 11 74.0 Comparative Example 12 75.9 control 31.0 negative control (no treatment) 100.0

Referring to Table 4, the samples of Examples 1 to 8 had excellent NO (nitric oxide) production inhibitory activity compared to Comparative Examples 1 to 12.

In particular, Example 1 containing tetrapeptide-21 and palmitoyl pentapeptide-4 at the same time was significantly superior in anti-inflammatory activity compared to Comparative Examples 8 to 11 lacking a part, and the result was a synergistic effect according to the combination of each component. suggests

Experimental Example 4: Evaluation of melanin production inhibitory activity

To evaluate the melanogenesis inhibitory effect of Examples and Comparative Examples, melanocytes B16F10 were cultured.

B16F10 was aliquoted into each well of a 6-well plate to make 8 x 10 ⁴ cells, and then incubated for 18 hours in a cell incubator under _{5% CO 2 , 37°C conditions.}

After 18 hours, the medium of the B16F10 cells was removed, and the samples of Examples and Comparative Examples were treated at the same concentration and cultured for 2 days in a cell incubator at _{5% CO 2 , 37° C. conditions.}

After 2 days, cells were recovered from the plate using a scraper and centrifuged for 1 minute to obtain cells in the form of pellets.

1N NaOH was added to the obtained cells and reacted at 95° C. for 10 minutes to dissolve melanin.

After 10 minutes, the amount of melanin production was measured at a wavelength of 475 nm using a microplate reader, and the amount of protein was measured at a wavelength of 595 nm using a BCA assay.

Each experimental group was numerically expressed as “absolute amount of melanin / absolute amount of protein”, and the experimental group and control group were quantified relative to each other.

α-MSH was used as a melanogenesis inducer, and arbutin was used as a positive control.

division Melanin Relative Value (%) Example 1 51.4 Example 2 48.9 Example 3 48.5 Example 4 46.6 Example 5 44.5 Example 6 43.3 Example 7 45.1 Example 8 39.5 Comparative Example 1 77.8 Comparative Example 2 74.2 Comparative Example 3 69.5 Comparative Example 4 68.4 Comparative Example 5 74.2 Comparative Example 6 77.8 Comparative Example 7 68.4 Comparative Example 8 77.8 Comparative Example 9 67.5 Comparative Example 10 74.2 Comparative Example 11 68.4 Comparative Example 12 74.2 positive control 43.3 control 100.0

Referring to Table 5, the samples of Examples 1 to 8 were excellent in melanin production inhibitory activity compared to Comparative Examples 1 to 12.

In particular, Example 1 containing tetrapeptide-21 and palmitoyl pentapeptide-4 at the same time was significantly superior in melanogenesis inhibitory activity compared to Comparative Examples 8 to 11 lacking a part, and the results are based on the combination of each component of the present application. This suggests a synergistic effect.

Experimental Example 5: Tyrosinase (tyrosinase) inhibitory activity evaluation

Tyrosinase inhibitory activity of the Examples and Comparative Examples was evaluated.

Inhibitory activity was measured by treatment with the same concentration, and L-DOPA was used as a substrate.

Dissolve mushroom tyrosinase in 68mM sodium phosphate buffer (pH 6.8) to make 125U/ml, and store at -20℃ for use in the experiment.

In a 96-well plate, 40 μM of diluted sample and 120 μL of 8 mM L-DOPA were added.

After adding 40 μL of mushroom tyrosinase and reacting for 20 minutes, absorbance was measured at 490 nm with an ELISA reader.

Arbutin was used as a control. Tyrosinase inhibitory activity was determined by measuring the absorbance ratio of the sample-added group compared to the sample-free group.

division Tyrosinase activity relative value (%) Example 1 44.5 Example 2 39.8 Example 3 41.1 Example 4 36.1 Example 5 32.8 Example 6 33.2 Example 7 32.4 Example 8 29.0 Comparative Example 1 72.0 Comparative Example 2 70.1 Comparative Example 3 74.7 Comparative Example 4 75.9 Comparative Example 5 74.0 Comparative Example 6 82.2 Comparative Example 7 76.3 Comparative Example 8 74.8 Comparative Example 9 72.2 Comparative Example 10 74.8 Comparative Example 11 76.3 Comparative Example 12 74.0 positive control 44.5 control 100.0

Referring to Table 6, the samples of Examples 1 to 8 had excellent tyrosinase inhibitory activity compared to Comparative Examples 1 to 12.

In particular, Example 1 containing tetrapeptide-21 and palmitoyl pentapeptide-4 at the same time was significantly superior in tyrosinase inhibitory activity compared to Comparative Examples 8 to 11 lacking a part, and the result is an excellent result of the present invention. suggest whitening activity.

Experimental Example 6: Elastase inhibitory effect evaluation

The wrinkle improvement effect of the samples of Examples and Comparative Examples was evaluated through the inhibitory activity of elastase. The samples of Examples and Comparative Examples were suspended in purified water, and elastase inhibitory activity was measured as follows.

1.6 mM elastase substrate (N-succinyl-(Ala)3-p-nitroanilide; SANA) and 0.2 mM Tris-HCl buffer (pH 8.0) were prepared. In a 96 well-plate, 0.165 mL of the Tris-HCl buffer, 0.005 mL of SANA and 0.01 mL of elastase were dispensed.

After putting the samples of Examples and Comparative Examples together with a positive control and reacting at 25° C. for 15 minutes, absorbance was measured at 405 nm.

The percentage (%) of elastase inhibitory activity was calculated as the ratio of the difference between the absorbance to which the sample was added to the absorbance to which the sample was not added.

Oleanolic acid was used as a positive control, and the concentration (IC ₅₀ ) that inhibited elastase by 50% of the samples of Examples and Comparative Examples was measured and shown in Table 7.

[Elastase 50% inhibitory concentration] division IC ₅₀ (μg/mL) Example 1 161.9 Example 2 152.5 Example 3 154.1 Example 4 146.2 Example 5 132.2 Example 6 132.4 Example 7 131.3 Example 8 114.2 Comparative Example 1 270.3 Comparative Example 2 277.5 Comparative Example 3 270.3 Comparative Example 4 263.2 Comparative Example 5 268.9 Comparative Example 6 280.4 Comparative Example 7 262.3 Comparative Example 8 266.2 Comparative Example 9 278.1 Comparative Example 10 268.9 Comparative Example 11 262.3 Comparative Example 12 265.9 positive control 98.8

The lower the IC ₅₀ value, the higher the elastase activity inhibition rate. Elastase is an enzyme that breaks down elastin, an important substrate for maintaining skin elasticity in the dermis. In addition, elastase is a non-specific hydrolase capable of decomposing collagen, which is another important matrix protein.

Therefore, the high elastase activity inhibition rate means that elastin and collagen, which are important for maintaining skin elasticity, are maintained for a long time, suggesting that the skin wrinkle improvement effect is excellent.

Referring to Table 7, the samples of Examples more effectively inhibited the elastase activity compared to Comparative Examples.

Experimental Example 7: Evaluation of wrinkle improvement effect

The skin wrinkle improvement effect of the cosmetic compositions formulated with the samples of Examples and Comparative Examples was evaluated.

Adult women over 30 years of age were divided into 20 groups, and the test subject was thoroughly absorbed by applying the cream-formed cosmetic to the face after cleansing every morning and evening for 4 weeks.

ANTERA 3D (Miravex, Ireland) was applied to evaluate wrinkle improvement, and device measurements were made before and 3 weeks after use of the test substance.

The same test person measured the left corner of the eye of all subjects, and for reproducibility of the measurement, the same part was measured by overlapping the image measured before the test substance was used.

The captured images were matched using ANTERA pro software, which is a dedicated software for ANTERA 3D, and then the matched measurement area was used for analysis.

Wrinkles small values representing skin wrinkles were analyzed using the indentation index, which is a measurement variable.

Table 8 shows the amount of change in Wrinkles small value after 4 weeks of the test subject according to the use of cosmetics formulated with samples of Examples and Comparative Examples. The greater the wrinkle improvement effect, the greater the change in the Wrinkles small value.

division Wrinkles small average Example 1 1.22 Example 2 1.27 Example 3 1.25 Example 4 1.32 Example 5 1.37 Example 6 1.39 Example 7 1.38 Example 8 1.51 Comparative Example 1 0.74 Comparative Example 2 0.73 Comparative Example 3 0.72 Comparative Example 4 0.81 Comparative Example 5 0.74 Comparative Example 6 0.74 Comparative Example 7 0.80 Comparative Example 8 0.75 Comparative Example 9 0.76 Comparative Example 10 0.81 Comparative Example 11 0.83 Comparative Example 12 0.82

Referring to Table 8, the cosmetic formulated with the sample of Example was significantly superior to the skin wrinkle improvement effect compared to Comparative Example.

Experimental Example 8: Evaluation of skin elasticity improvement effect

The skin elasticity improvement effect of the cosmetic compositions formulated with the samples of Examples and Comparative Examples was evaluated.

Each of the samples was formulated as a cream of the same composition, and only the samples of Examples and Comparative Examples were different.

The creams of Examples and Comparative Examples prepared by dividing into groups of healthy women (average age 39 years) over 20 years of age at a temperature of 24 to 26° C. and a humidity of 75% were applied, respectively.

The cream formulated for 12 weeks was applied twice a day (morning and evening) around the eyes, and skin elasticity was measured using a skin elasticity meter (Cutometer MPA580, Conrage + Khazaka, Germany).

The test results were described as the R8 (R8 (12 weeks)-R8 (0 weeks)) value of the cutometer MPA580, and the R8 value represents the property of skin viscoelasticity, and the results are shown in Table 9 below.

division Skin elasticity improvement effect Example 1 0.69 Example 2 0.77 Example 3 0.79 Example 4 0.78 Example 5 0.83 Example 6 0.84 Example 7 0.83 Example 8 0.91 Comparative Example 1 0.44 Comparative Example 2 0.37 Comparative Example 3 0.36 Comparative Example 4 0.41 Comparative Example 5 0.38 Comparative Example 6 0.42 Comparative Example 7 0.39 Comparative Example 8 0.45 Comparative Example 9 0.45 Comparative Example 10 0.47 Comparative Example 11 0.48 Comparative Example 12 0.44 Positive control (vitamin C) 0.91 negative control (no treatment) 0.31

Referring to Table 9, the sample of Example was significantly superior to the skin elasticity improvement effect compared to Comparative Example.

In particular, Example 1 containing tetrapeptide-21 and palmitoyl pentapeptide-4 at the same time was significantly superior to Comparative Examples 8 to 11 lacking some, and the results suggest a synergistic effect according to the combination of each component. .

Experimental Example 9: Evaluation of cell migration promoting activity

The skin regeneration and wound healing effects of the samples of Examples and Comparative Examples were confirmed through a wound healing assay.

Specifically, HaCat cells were seeded in a 96-well plate at ^{the number of 1×10 3} cells/well.

After 24 hours, the samples of Examples and Comparative Examples were treated at the same concentration (1%) in each well after replacing the DMEM medium with 5% FBS.

After 48 hours, a wound was made using a yellow tip, and the area was measured through the image J program by taking a picture of the degree of closure of the wound after 24 hours.

division Cell Mobility (%) Example 1 55.0 Example 2 60.6 Example 3 60.2 Example 4 64.0 Example 5 67.7 Example 6 68.4 Example 7 66.9 Example 8 74.5 Comparative Example 1 28.0 Comparative Example 2 27.1 Comparative Example 3 26.5 Comparative Example 4 25.8 Comparative Example 5 27.7 Comparative Example 6 26.4 Comparative Example 7 27.5 Comparative Example 8 32.1 Comparative Example 9 30.1 Comparative Example 10 31.2 Comparative Example 11 31.7 Comparative Example 12 31.0

Referring to Table 10, the sample of Example was remarkably excellent in cell migration ability compared to Comparative Example.

In particular, Example 1 containing tetrapeptide-21 and palmitoyl pentapeptide-4 simultaneously has significantly superior cell migration promoting activity compared to Comparative Examples 8 to 11 lacking some, and the results are according to the combination of each component. It suggests that skin regeneration and wound healing effects are improved by synergy.

The description of the present invention described above is for illustration, and those of ordinary skill in the art to which the present invention pertains can understand that it can be easily modified into other specific forms without changing the technical spirit or essential features of the present invention. will be. Therefore, it should be understood that the embodiments described above are illustrative in all respects and not restrictive. For example, each component described as a single type may be implemented in a distributed manner, and likewise components described as distributed may be implemented in a combined form.

The scope of the present invention is indicated by the following claims, and all changes or modifications derived from the meaning and scope of the claims and their equivalents should be construed as being included in the scope of the present invention.

Claims (8)

Anti-wrinkle containing Tetrapeptide-21, Palmitoyl Pentapeptide-4, Hexapeptide-11, Carnosine and Zymoji root extract as active ingredients and a cosmetic composition for skin whitening. delete According to claim 1,
A cosmetic composition further comprising at least one selected from the group consisting of adenosine and glutathione. 4. The method of claim 1 or 3,
A cosmetic composition further comprising the use of improving skin elasticity. delete 4. The method of claim 1 or 3,
A cosmetic composition further comprising an anti-inflammatory or antioxidant use. 4. The method of claim 1 or 3,
A cosmetic composition further comprising a skin regeneration use. 4. The method of claim 1 or 3,
A cosmetic composition formulated with one or more selected from the group consisting of softening lotion, nourishing lotion, nourishing cream, massage cream, essence, ampoule, gel, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray and powder.