KR101252149B1 - Cosmetic composition for anti-aging - Google Patents

Abstract

PURPOSE: A cosmetic composition containing a mixture extract of Caulerpa lentillifera and sea staghorn for anti-aging is provided to enhance anti-oxidation, anti-inflammation, moisturization, anti-wrinkling, and elasticity. CONSTITUTION: A cosmetic composition for anti-aging contains mixture extract of Caulerpa lentillifera and sea staghorn. The extract is prepared using water, C1-C4 alcohol, or a mixture thereof. The cosmetic composition is manufactured in the form of a skin softener, a lotion, a nutrition cream, an essence, a pack, an eye cream, a cleansing cream, a cleansing foam, a cleansing water, a spray, or a powder.

Description

Anti-aging cosmetic composition {Cosmetic composition for anti-aging}

The present invention relates to an anti-aging cosmetic composition comprising a sea grape and an auditory mixed extract.

The causes of wrinkles can be broadly divided into the generation of wrinkles by natural aging and the generation of wrinkles by UV exposure (photoaging), and the mechanism of wrinkle formation by two causes is not known yet. Common phenomena found in relation to skin changes due to natural aging and photoaging are the reduction and modification of collagen and elastin, the major constituent proteins in the dermis, and the reduction of skin elasticity due to the reduction of the hyaluronic acid, the substrate of the dermis. In particular, according to the results of the previous studies, it is known that the reduction and modification of the main proteins in the dermis, collagen and elastin, are directly involved in the formation of wrinkles and the deterioration of skin elasticity.

Collagen is a major substrate protein produced by skin fibroblasts and is present in extracellular epilepsy.Its important functions include mechanical firmness of skin, resistance of connective tissue and tissue binding, support of cell adhesion, cell division and differentiation. Induction of growth or wound healing) is known. This collagen is reduced by age and photoaging by ultraviolet irradiation, which is known to be closely associated with wrinkle formation of the skin. In recent years, extensive research on skin aging has been reported to reveal the important function of collagen in the skin. Active ingredients are known that promote collagen synthesis and have anti-wrinkle effects. For example, retinoic acid, transforming growth factor (TGF), protein from animal placenta, betulinic acid, chlorella extract and the like are known as collagen synthesis promoting substances.

The glycosylation or non-enzymatic glycation reaction refers to the free amino group of lysine or arginine, and the carbonyl group of the reducing sugar reacts to form the initial glycation product, the shiff base. It is a reaction that forms melanoidin through a series of complex reactions such as dislocation, oxidation, cleavage and cyclization to form irreversible advanced glycation end products (AGEs). Glycation reactions occur more rapidly in diabetic patients with hyperglycemia than in normal people, but normal blood glucose occurs in the body as aging progresses. Collagen, which has a relatively long half-life, is known to be easily saccharified and cross-linked, and the resultant glycosylated product accumulates in the skin through several stages of reaction, resulting in decreased elasticity and wrinkles of the skin. Many previous researchers report that glycosylation inhibitors, such as aminoguanidine (AG), have the effect of maintaining collagen safety.

Sea grapes ( Caulerpa lentillifera ) are algae that live in Okinawa, the Philippines, and Vietnam, and are called Sea Grape and Green Caviar. One of the five seaweed delicacies eaten by Okinawans, it is famous for its longevity. Sea grapes are alkaline foods, a treasure trove of various minerals, vitamins, and dietary fiber, alginic acid, which clears blood, inhibits the production of free radicals and lipids, and helps to excrete and excrete waste products. About 10% protein and about 30% to 40% sugar is known to contain.

Hearing Codium fragile ) grows on rocks in the calm waters of the seas of northern, central and southern Korea. "Janjabo" has a smooth texture, dark blue color, and a light taste that makes kimchi arouse the taste of kimchi. In Japan, the sonar lives in the sea and is called Miru. The height is 10-30cm, the thickness is 1.5-3mm, and the lower side is thicker. The branches are split into deer horns, grow straight, and reach the same height, forming a fan shape. The surface is smooth like a velvety, and under a microscope there is an irregular tangled texture of colorless transparent threads inside the branches. Young ones have hairs on the front, especially on top, but fall later and leave hairless marks. The whole body is a structure in which several cell nuclei are contained in one cell because they are all connected without a membrane between cells. Spore containing pouches are club-shaped, 5-7 times the thickness, the top is sharp. It grows in the depths of 1 ~ 20m deep and less affected by the waves. In particular, it is not affected by the water flowing from the land, and is affected by the seawater flowing from the distant sea, and lives on rocks, shells or other algae. Common in temperate and subtropical waters. Dip water kimchi with Chinese cabbage, or eat it with herbs. When you add kimchi, it neutralizes the smell of salted fish, fish, and garlic to make the aftertaste refresh. In the past, it was used as a roundworm medicine because of the antiparasitic component, and the water-soluble extract is known to have strong antibiotic action against bacteria.

Korean Patent Publication No. 2012-0056594, 2012. 06. 04

An object of the present invention is to provide a cosmetic composition having anti-aging activity by the antioxidant, anti-inflammatory, moisturizing, anti-glycosylation, wrinkle improvement and elasticity enhancing effect.

The present inventors earnestly researched to develop a cosmetic composition having anti-aging activity such as wrinkle improvement and elasticity enhancement, and completed the present invention by confirming that sea grape and auditory mixed extracts exhibited excellent activity in promoting antioxidant and collagen production. .

In order to achieve the above object, the present invention provides an anti-aging cosmetic composition comprising a sea grape and auditory mixed extract.

Sea grape and auditory mixed extract of the present invention, anti-wrinkle, anti-collagen production, collagenase expression inhibition, elastase expression inhibition, NO production inhibition, prostaglandin production inhibition and TNF-α production to improve wrinkles, produce the final glycation product It has an inhibitory, elasticity and moisturizing effect to prevent skin aging.

Using a mixture of sea grapes and hearing has better effects in antioxidant, promoting collagen production, inhibiting collagenase expression, and the like than using sea grape extract or auditory extract, respectively. That is, the sea grapes and the auditory mixed extract, has an anti-aging effect superior to each extract.

Sea grapes ( Caulerpa lentillifera ) in the present invention can be purchased from Japan or the Philippines and used.

In the present invention hearing ( Codium fragile ) can be purchased from domestic market, online market and Japan.

Sea grapes and hearing in the present invention is used to cleanly wash with purified water, it is preferable to use the raw as it is or crushed according to the extraction method.

In the present invention, the sea grapes and the auditory mixed extract may be an extract obtained by first mixing the sea grapes and the dry material of the hearing, or may be a mixed extract of sea grapes and auditory extracts respectively. Sea grape and auditory mixed extract in the present invention may be a concentrated liquid, or may be in the form of freeze-dried powder.

Sea grape and auditory mixed extract in the present invention is prepared by known extraction methods known to those skilled in the art. For example, the sea grapes and the acoustic dry matter are mixed and chopped, and the extraction solvent selected from 1 to 50 times the volume of the dry weight of water, an alcohol having 1 to 4 carbon atoms, or a hydrous alcohol thereof is added, and then 4 to 30 ° C. After immersion for 3 to 20 days to extract the active ingredient, the extraction solvent may be obtained by concentrating with a vacuum condenser.

In addition, the sea grapes and the auditory mixed extract of the present invention is pulverized and mixed dry matter, 1 to 30 times the volume of distilled water, alcohol having 1 to 4 carbon atoms, ethyl acetate, glycerin, ethylene glycol, propylene mixed dry weight of these Glycol, butylene glycol and mixed solvents thereof (e.g., hydrous alcohol, hydrous glycerin, hydrous ethylene glycol, hydrous propylene glycol, hydrous butylene glycol) can be extracted with a solvent, preferably water, carbon 1 It may be extracted with a solvent selected from anhydrous alcohols or hydrous alcohols of 4 to 4. Here, the alcohol having 1 to 4 carbon atoms may be methanol, ethanol, n-propanol, iso-propanol, n-butanol, iso-butanol, tert-butanol, more preferably ethanol, the hydrous alcohol is water 0.3 to 90 (wt / wt)%. Sea grape and auditory mixed extract can be extracted by heating for 1 to 48 hours at 30 to 100 ° C. or by dipping at 5 to 30 ° C. for 1 to 5 days, and preferably cooled to prevent evaporation of the solvent. It can extract with the extractor provided with a condenser. Finally, the extraction solvent may be obtained by concentrating with a vacuum concentrator.

In addition, the sea grapes and the auditory mixed extract of the present invention may be obtained by preparing each of the sea grapes and the auditory, respectively, after mixing them.

In addition, it can be prepared using the ultrasonic extraction method and the like known in the extraction solvents listed above.

In the present invention, the mixed weight ratio of the sea grapes and the auditory mixed extract may vary, but is preferably mixed in a dry weight ratio of 1: 2 to 2: 1, and the mixed dry weight ratio of the sea grape extract and the auditory extract is 1: 1. Even more preferably.

Sea grape and auditory mixed extract in the present invention may be added to the cosmetic in an amount of 0.001% to 10% by weight in powder form or liquid form with respect to the total weight of the cosmetic composition, preferably in powder form relative to the total weight of the cosmetic composition Or in an amount of 0.01 to 5% by weight in liquid form.

In the present invention, the components included in the cosmetic composition include components commonly used in cosmetic compositions in addition to sea grapes and auditory mixed extracts as active ingredients, and include, for example, antioxidants, stabilizers, solubilizers, vitamins, pigments, pigments and flavors. Such conventional adjuvants, and carriers.

The cosmetic composition of the present invention can be prepared into any of the formulations conventionally produced in the art and can be used as a solution, a suspension, an emulsion, a paste, a gel, a cream, a lotion, a powder, a soap, , Oil, powder foundation, emulsion foundation, wax foundation and spray, but is not limited thereto. More specifically, it may be prepared in the form of a flexible lotion, nutrition lotion, nutrition cream, massage cream, essence, pack, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray or powder.

When the formulation of the present invention is a paste, cream or gel, an animal oil, vegetable oil, wax, paraffin, starch, tracant, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide may be used as the carrier component .

In the case where the formulation of the present invention is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component. Especially, in the case of a spray, a mixture of chlorofluorohydrocarbons, propane / Propane or dimethyl ether.

When the formulation of the present invention is a solution or emulsion, a solvent, solubilizer or emulsifier is used as the carrier component, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, butyl Ethylene glycol, 1,3-butylglycol oil, polyoxyethylene hardened castor oil, glycerol, glycerin, aliphatic ester, phenoxyethanol, triethanolamine, polyethylene glycol, beeswax, polysorbate 60, sorbitan sesquiolade, paraffin, Sorbitan stearate, lipophilic monostearic acid glycerine, stearic acid, glyceryl stearate / fiji-400 stearate, carboxypolymer, cytososterol, polyglyceryl 2-oleate, ceramide, cholesterol, steares-4, dicetylphosphate , Fatty acids of macadamia oil, carboxyvinyl polymer, xanthan gum or sorbitan Esters and the like.

When the formulation of the present invention is a suspension, suspensions such as liquid diluents such as water, ethanol, butylene glycol or propylene glycol, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol esters and polyoxyethylene sorbitan esters as carrier components Secondly, microcrystalline cellulose, hydroxyethyl cellulose, sodium hyaluronate, phenoxyethanol, aluminum meta hydroxide, bentonite, agar or tracant and the like can be used.

When the formulation of the present invention is an interfacial active agent-containing cleansing, the carrier component may include aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyltaurate, sarcosinate, fatty acid amide Ether sulfates, alkylamidobetaines, aliphatic alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, lanolin derivatives, or ethoxylated glycerol fatty acid esters.

Anti-aging cosmetic composition comprising sea grapes and auditory mixed extract according to the present invention is antioxidant, promote collagen production, collagenase expression inhibition, elastase expression inhibition, NO production inhibition, final glycation product production inhibition, prostaglandin production inhibition and It has excellent anti-aging effect by having wrinkle improvement, elasticity enhancement and moisturizing effect through inhibition of TNF-α production.

Hereinafter, the present invention will be described in more detail with reference to the following examples and experimental examples. However, the following examples are not intended to limit the scope of the present invention, which will be construed as to aid the understanding of the present invention.

Example  One. Sea grapes  And preparation of auditory mixed extract

After washing sea grapes imported and purchased from Southeast Asia (Philippines) with purified water, put 100g of dried sea grapes into 2kg of distilled water, and extract them with a cooling condenser (Product name: Cosmos-660, manufacturer: Gyeongseo Machinery). The mixture was heated to 100 ° C. and extracted three times for 2 hours.

After washing the hearing (origin: Sando) purchased from the online market with purified water, put 100 g of dried hearing in 2 kg of distilled water, and extract it with a cooling condenser (Product name: Cosmos-660, manufacturer: Gyeongseo Machinery). The mixture was heated to ℃ and extracted three times for 2 hours each.

After extracting the sea grapes and the hearing by the above method, 50 g of sea grape extract and 50 g of auditory extract were mixed at the same ratio and filtered through 300 mesh filter paper, and the precipitate was filtered twice using Edventech No. 5 filter paper and Whatman GFC 150 mm filter paper. Filtered. After concentrating at a temperature of 40 ° C. to 50 ° C. using a reduced pressure concentrator (product name: Coolace CCA-1100, manufacturer: EYELA), an inlet was used by using a spray dryer (model name: B-290, manufactured by BUCHI). ) Temperature: 180 ℃, Aspirator Efficiency: 100% (35cm3 / hr), Pump Efficiency: 25% (7.5ml / min), Nozzle Cleaner 4 and Rotameter: Drying under the condition of 30 mm (357 liters / hour) gave 10 g of the title extract powder.

Comparative example  One. Sea grapes  Preparation of extract

After washing sea grapes imported and purchased from Southeast Asia (Philippines) with purified water, add 100g of dried sea grapes to 2kg of distilled water and add 80kg ~ in an extractor equipped with a cooling condenser The mixture was heated to 100 ° C. and extracted three times for 2 hours. 50 g of sea grape extract was filtered through a 300 mesh filter paper, and the precipitate was filtered twice using an Edbentech No. 5 filter paper and a Whatman GFC 150 mm filter paper. After concentrating at a temperature of 40 ° C. to 50 ° C. using a reduced pressure concentrator (product name: Coolace CCA-1100, manufacturer: EYELA), an inlet was used by using a spray dryer (model name: B-290, manufactured by BUCHI). ) Temperature: 180 ℃, Aspirator Efficiency: 100% (35cm3 / hr), Pump Efficiency: 25% (7.5ml / min), Nozzle Cleaner 4 and Rotameter: Drying under the condition of 30 mm (357 liters / hour) gave 5 g of the title extract powder.

Comparative example  2. Preparation of Auditory Extracts

After washing the hearing (origin: Sando) purchased from the online market with purified water, put 100 grams of the dried hearing into 2 kg of distilled water and put it in an extractor equipped with a cooling condenser (Product name: Cosmos-660, manufacturer: Gyeongseo Machinery). The mixture was heated to ℃ and extracted three times for 2 hours each.

50 g of the auditory extracts were filtered through 300 mesh filter paper, and the precipitate was filtered twice with Edbentech No. 5 filter paper and Whatman GFC 150 mm filter paper. After concentrating at a temperature of 40 ° C. to 50 ° C. using a reduced pressure concentrator (product name: Coolace CCA-1100, manufacturer: EYELA), an inlet was used by using a spray dryer (model name: B-290, manufactured by BUCHI). ) Temperature: 180 ℃, Aspirator Efficiency: 100% (35cm3 / hr), Pump Efficiency: 25% (7.5ml / min), Nozzle Cleaner 4 and Rotameter: Drying under the condition of 30 mm (357 liters / hour) gave 5 g of the title extract powder.

Experimental Example  1. Confirm antioxidant effect

The scavenging activity of the superoxide radicals produced by Xanthine oxidase was measured by NBT (Nitro-Blue Tetrazolium) reduction method.

1 ml of 0.1 M potassium phosphate buffer (pH 7.5) containing 0.4 mM xanthine and 0.24 mM NBT and 0.6 ml of 0.1 M potassium phosphate buffer (pH 7.5) were mixed with 0.1 ml of sample and then xanthine oxy 1 ml of a multidose (0.05 U / ml) was added and reacted at 37 ° C. for 20 minutes. After the reaction was terminated by adding 1 ml of 1 M HCl, the absorbance at 520 nm was measured. Sample solutions of Comparative Example 1, Comparative Example 2 and Example 1 were dissolved in DMSO (Dimethyl Sulfoxide) solvent and prepared at a concentration of 0.1, 0.5, 1.0, 2.0, 5.0 (wt / v)%. NBT reduction inhibitory ability was calculated by the following [Formula 1], the results are shown in the following [Table 1].

[Formula 1]

In [Formula 1], B (Blank) is the absorbance of 0.1 M potassium phosphate buffer (pH7.5) containing 0.4 mM xanthine and 1 M HCl, and C (Control) is 0.24 mM NBT instead of the sample. The absorbance of one reaction solution, S (Sample), represents the absorbance of the reaction solution to which the sample was added.

[Table 1] Antioxidant Effect of Marine Grape and Hearing Mixture Extracts

As shown in [Table 1], when the same concentration treatment, it was confirmed that the sea grapes and the auditory mixed extract of the sea grape extract and the auditory extract of Comparative Example 1, Comparative Example 2, respectively, showed an excellent antioxidant effect. That is, it can be seen that the sea grapes and the auditory mixed extract have an increased antioxidant effect than the respective extracts.

Experimental Example  2. Check for cytotoxicity

MTT assay was used to determine the cytotoxicity of sea grapes and auditory mixed extracts. After culturing melanocytes in a T-75 culture flask, the culture medium was discarded and treated with 5 ml of Trypsin-EDTA to collect all melanocytes. The number of harvested cells was measured using a microscope and the culture solution was diluted to make a cell suspension. Next, 1 ml of the cell suspension was dispensed to a number of 30,000 cells / well in a 12 well plate, and a natural product sample was administered after 24 hours. Dosing concentration was prepared according to the test sample as a final concentration, and treated with MTT reagent after 48 hours incubation. After 4 hours of reagent treatment, the culture medium was removed, dried well, and then dissolved in cells in DMSO solution. The concentration of formazan salt appeared at this time was measured by absorbance at 560 nm.

Cell viability was calculated by the following [Formula 2], the results are shown in the following [Table 2].

[Formula 2]

[Table 2] Confirmation of Cytotoxicity of Sea Grape and Hearing Mixture Extracts

As shown in Table 2 above, when the cell viability under the optimal conditions for cell growth was 100%, neither sea grape extract, auditory extract, and sea grape and auditory mixed extract showed cytotoxicity and cell proliferation effect. It could be confirmed.

Experimental Example  3. Confirmation of intracellular collagen production effect

Procollagen synthesis was measured using HDF-N (Normal Human Primary Dermal Fibroblasts-Neonatal) derived from human neonatal foreskin fibroblasts.

Fibroblast Basal Medium (FBM, Lonza, Inc.) containing 0.1% insulin, 0.1% rhFGF, 0.1% gentamicin, 2% FBS after inoculating HDF-N cell line (ATCC PCS-201-010) to the bottom of the culture dish. CC-3131) medium was added, and cultured in an incubator containing 37 ° C 5% CO ₂ .

In order to measure the procollagen (Procollagen) synthesis capacity, HDF-N cells were aliquoted into 1.4 wells in 48 well plates at 1.4 X 10 ⁴ cells / well and incubated for 24 hours in an incubator containing 5% CO ₂ at 37 ℃. Thereafter, the medium was removed, and the cells were starved for 24 hours. After that, the test substance was diluted in FBM (except for media supplement), which is a cell culture medium, and the cells were treated by concentration, followed by culturing at 37 ° C. for 24 hours. After 24 hours, the cultured cells were harvested and procollagen was measured using a procollagen type c-peptide (PIP) EIA kit (TAKARA, MK101). On the other hand, the cells attached to the bottom was washed with PBS (Phosphate Buffered Saline), and then lysed (lysed) with 1N NaOH to measure the total protein amount to measure the amount of procollagen synthesis per protein. Procollagen and protein amounts were calculated according to the calibration curve. The protein amount was measured using a Bio-rad DC protein kit (Bio-rad, 500-0001) based on the Lowry method.

The results are shown in [Table 3].

[Table 3] Collagen production effect of sea grapes and auditory mixed extract

As shown in [Table 3], it was confirmed that the sea grapes and the hearing mixed extracts exhibited superior procollagen synthetizing effects than the sea grape extracts and the auditory extracts of Comparative Examples 1 and 2 when the same concentration was treated. In other words, it can be seen that the sea grapes and the auditory mixed extract have an elevated collagen synthesis effect than the respective extracts.

Experimental Example  4. Collagenase ( MMP -1) Expression inhibition effect confirmed

The anti-wrinkle effect was confirmed by measuring the expression inhibitory effect of collagenase (MMP-1) using HDF-N (Normal Human Primary Dermal Fibroblasts-Neonatal) derived from human neonatal foreskin fibroblasts. .

Fibroblast Basal Medium (FBM, containing 0.1% insulin, 0.1% rhFGF, 0.1% gentamicin, 2% FBS) after inoculating HDF-N cell line (ATCC PCS-201-010) on the bottom of a 24-well plate Lonza, CC-3131) medium was added, and maintained at 37 ° C. and cultured in an incubator containing 5% CO ₂ . After irradiating UV-A (15J / ㎠) in a range that does not affect the fibroblasts cultured using the UV chamber, the sample was incubated for 48 hours by replacing it with the medium to which the sample was added, and then the medium was recovered to obtain a 96-well plate. Coated on. The blocking solution was treated for 1 hour to block sites where collagenase was not bound. After reacting the secondary antibody for about 90 minutes, the primary antibody was treated and reacted at 37 ° C. for 90 minutes, followed by color development by adding 100 µl of the substrate solution. Samples were incubated for 24 hours in DMEM medium containing no added serum. Absorbance was measured at 405 nm using an ELISA reader. Collagenase expression inhibition rate was calculated by the following [Formula 3], the results are shown in the following [Table 4].

[Equation 3]

[Table 4] Confirmation of the inhibitory effect of MMP-1 expression of sea grape and auditory mixed extract

As shown in [Table 4], it was confirmed that sea grape and auditory mixed extracts showed superior collagenase expression inhibitory effect than sea grape extract and auditory extract of Comparative Example 1 and Comparative Example 2 at the same concentration treatment. . In other words, it can be seen that the sea grapes and the auditory mixed extract have a higher collagenase expression inhibitory effect than the respective extracts.

Experimental Example  5. Elastase ( Elastase ) Expression inhibition effect confirmed

The anti-wrinkle effect was confirmed by measuring the expression inhibitory effect of Elastase using HDF-N (Normal Human Primary Dermal Fibroblasts-Neonatal) derived from human neonatal foreskin fibroblasts.

Fibroblasts were placed in a 96 well plate containing DMEM medium containing 2.5% fetal calf serum and incubated until 90% growth. Thereafter, the cells were incubated for 24 hours in a serum-free medium, and the samples were incubated for 24 hours in a serum-free DMEM medium in which concentrations were added. Cell cultures were placed in 96 well plates uniformly coated with elastase antibodies, and antigen-antibody reactions were performed in a thermostat for 3 hours. After 3 hours, the chromophore-bound secondary elastin antibody was placed in a 96 well plate and reacted again for 15 minutes. After 15 minutes, a color-causing substance was added to cause color development at room temperature for 15 minutes. Then, 1M sulfuric acid was added to stop the reaction. The color of the reaction solution was yellow and the degree of yellow color was changed according to the progress of the reaction. Absorbance was measured at 405 nm using an ELISA reader. Elastase expression inhibition rate was calculated by the following [Formula 4], the results are shown in the following [Table 5].

[Formula 4]

[Table 5] Confirmation of Elastase Expression Inhibitory Effect of Sea Grape and Acoustic Mixed Extracts

As shown in [Table 5], it was confirmed that the sea grapes and the auditory mixed extracts showed superior elastase expression inhibitory effect than the sea grape extracts and the auditory extracts of Comparative Examples 1 and 2 when the same concentration treatment. .

In other words, it can be seen that the sea grapes and the auditory mixed extract have an elastase expression inhibitory effect higher than that of each extract.

Experimental Example  6. NO ( Nitric Oxide ) Confirmation of generation inhibitory effect

Wrinkle improvement effect was confirmed by measuring the NO production inhibitory effect according to the present invention extract in RAW264.7 cells.

RAW264.7 cells were adjusted to 5 × 10 5 cells / well using DMEM medium supplemented with 10% FBS, and then inoculated into 24 well plates, samples were treated by concentration, and cultured for 30 minutes, then LPS (lopopolysaccharide, 1 ㎍ / Ml) was added and incubated for 24 hours. The amount of generated NO was measured using a NO detection kit (iNtRON). The inhibition of NO production was calculated by the following [Formula 5], and the results are shown in the following [Table 6].

[Equation 5]

[Table 6] Confirmation of NO Production Inhibition Effect of Sea Grape and Hearing Mixture Extracts

As shown in [Table 6], it was confirmed that the sea grapes and the auditory mixed extracts showed superior NO production inhibitory effect than the sea grape extracts and the auditory extracts of Comparative Examples 1 and 2 when the same concentration treatment.

In other words, it can be seen that the sea grapes and the auditory mixed extract have a higher NO production inhibitory effect than the respective extracts.

Experimental Example  7. Prostaglandins ( PGE ₂ ) Confirmation of generation inhibitory effect

Wrinkle improvement effect was confirmed by measuring the inhibitory effect of prostaglandin production according to the present invention extract in RAW264.7 cells.

RAW264.7 cells were adjusted to 5 × 10 5 cells / well using DMEM medium supplemented with 10% FBS, and then inoculated into 24 well plates, samples were treated by concentration, and cultured for 30 minutes, then LPS (lopopolysaccharide, 1 ㎍ / Ml) was added and incubated for 24 hours. The amount of PGE ₂ produced was measured using a PGE ₂ ELISA kit (R & D System, Inc, USA). The inhibition of PGE ₂ production was calculated by the following [Formula 6], and the results are shown in the following [Table 7].

[Equation 6]

[Table 7] Confirmation of PGE ₂ Production Inhibition Effect of Sea Grape and Acoustic Mixed Extracts

As shown in [Table 7], it was confirmed that the sea grapes and the auditory mixed extracts showed superior PGE ₂ production inhibitory effect than the sea grape extracts and the auditory extracts of Comparative Examples 1 and 2 when the same concentration was treated.

In other words, it can be seen that the sea grapes and the auditory mixed extract have a higher PGE ₂ inhibitory effect than the respective extracts.

Experimental Example  8. TNF -α production inhibitory effect

Wrinkle improvement effect was confirmed by measuring the inhibitory effect of TNF-α production in accordance with the present invention extract in RAW264.7 cells.

RAW264.7 cells were adjusted to 5 × 10 5 cells / well using DMEM medium supplemented with 10% FBS, and then inoculated into 24 well plates, samples were treated by concentration, and cultured for 30 minutes, followed by LPS (lopopolysaccharide, 1㎍ /). Ml) was added and incubated for 24 hours. The amount of TNF-α produced was measured using a TNF-α ELISA kit (eBioscience INC, USA). Inhibition rate of TNF-α production was calculated by the following [Formula 7], and the results are shown in the following [Table 8].

[Equation 7]

[Table 8] Confirmation of TNF-α Production Inhibitory Effect of Sea Grape and Acoustic Mixed Extracts

As shown in [Table 8], it was confirmed that the sea grape and auditory mixed extracts showed superior TNF-α production inhibitory effect than the sea grape extract and the auditory extract of Comparative Example 1 and Comparative Example 2 at the same concentration treatment. .

That is, it can be seen that the sea grapes and the auditory mixed extract have a higher TNF-α production inhibitory effect than the respective extracts.

Experimental Example  9. Check the moisturizing effect

As a clinical trial for skin moisturizing, skin moisturizing ability was evaluated by measuring skin conductivity using a skin moisture meter (Corneometer, Courage + Khazaka, GmbH, Germany). Skin moisture was measured inside the arm of five subjects using a skin moisture meter under constant temperature and humidity conditions maintained at room temperature 20-25 ° C and relative humidity 40-55%. Moisturizing power was calculated by the following [Equation 8], the results are shown in the following [Table 9].

[Equation 8]

[Table 9] Confirmation of moisturizing power of sea grapes and auditory mixed extracts

As shown in [Table 9], it was confirmed that the sea grapes and the auditory mixed extracts showed excellent moisturizing effects than the sea grape extracts and the auditory extracts of Comparative Examples 1 and 2 when the same concentration treatment.

In other words, it can be seen that the sea grapes and the auditory mixed extract have an increased moisturizing effect than the respective extracts.

Experimental Example  10. Antiglycosylation  Check the effect

As the glycosylation reaction proceeds, a series of complex processes such as oxidation and condensation produce final glycation products (AGEs) in which various compounds are mixed. Some of the final glycation products are fluorescence and are used as a measure to measure the degree of glycation reaction. Among them, the fluorescence at excitation 370 nm / emission 440 nm is generally known as a numerical value representing the degree of glycation reaction.

The experiment was conducted using the above principle. Preparation of the reaction was performed by partially modifying the method of Mcpherson et al. (Role of fructose in glycation and cross-linking of proteins. Biochemistry. 1998. 27. 1901-1907). Bovine serum albumin (BSA, 5 mg / ml), 25 mM ribose, 1 mM diethylene triamine pentaacetic acid (DTPA) and 0.02% sodium azide were mixed in 0.1 M phosphate buffer (pH 7.0) to 96 180 μl was dispensed into the well plate (Nunc, Denmark), and 20 μl of each sample (10 mg / ml) was tested. At this time, the control group (A), the glycosylation control group in which the glycosylation reaction occurred in the preparation, and each sample treatment group were prepared because the mixture was not mixed with ribose. In order to exclude interference of fluorescence by each sample, a sample treatment group not treated with ribose was also prepared. In order to prevent drying during the reaction, the reaction was carried out for 7 days in a 37 ° C. constant temperature and humidity incubator. After the reaction, the fluorescence was measured at ex = 370 nm / em = 440 nm using a spectrofluorometer (VICTOR3, Perkin Elmer, Waltham, MA, USA), and each sample was repeated three times. The final glycosylation product inhibitory activity was 100% in the control group (A) that did not cause glycation reaction and 0% of the glycation control group in which the glycosylation reaction occurred. In this case, a value (CD) was used to exclude fluorescence of each sample in which no glycosylation reaction occurred. Final glycation product production inhibition rate (%) was calculated by the following [Equation 9], the results are shown in the following [Table 10].

[Equation 9]

[Table 10] Anti-glycosylation effect of sea grapes and auditory mixed extracts

As shown in [Table 10], it was confirmed that the sea grapes and the auditory mixed extracts were significantly superior to the final glycation product production inhibitory effect than the sea grape extracts and the auditory extracts of Comparative Examples 1 and 2 when the same concentration was treated. there was.

That is, it can be seen that the sea grapes and the auditory mixed extract have a higher inhibitory effect on the production of the final glycated product than the respective extracts.

Formulation Example  One. Sea grapes  And preparation of a lotion containing an auditory mixed extract

1 kg of the skin lotion (skin lotion) containing the sea grapes and the auditory extract (Example 1) was prepared using the following preparation method in the following [Table 11].

[Table 11] Cosmetic lotion composition containing sea grapes and auditory mixed extracts

In Table 11, materials 2, 3, 4, and 8 were added to the reaction container in order, and dissolved by stirring. Subsequently, materials 5 and 6 were heated and dissolved at about 60 ° C. Thereafter, the material was added 10 times to dissolve and then added to material 11. Finally, 1, 6, 7, and 9 were added to the material and sufficiently stirred, followed by aging at 25 ° C. for 3 days to prepare the title lotion.

Formulation Example  2. Sea grapes  And nutrition lotion containing hearing mixed extract

1 kg of the hydrosome nutrient lotion containing the sea grapes and the auditory mixed extract (Example 1) was prepared using the following preparation method with the content of the following [Table 12].

[Table 12] Nutritional Lotion Composition Containing Mixed Sea Grape and Hearing Extracts

In Table 12, the materials 10, 11, 13, and 16 are mixed and stirred and heated to 80 to 85 ° C., and then put into the manufacturing unit, and then the emulsifier is activated. 2, 3, 4, 5, 6, 7, 8, Material 9 and 12 were dissolved after heating to 80 ~ 85 ℃. After emulsification, cool down to 50 ℃ while stirring using a stirrer, and then add material 15 and cool to 45 ℃, add material 14 and add material 1 at 35 ℃ to cool to 25 ℃, and then 25 ℃. Aged 3 days in to prepare the title nutrition lotion.

Formulation Example  3. Sea grapes  And Acoustic Blend Extract Serum  Produce

1 kg of the hydrosome nutrient serum containing sea grapes and auditory mixed extract (Example 1) was prepared using the following preparation method.

Table 13: Nutritional Serum Composition Containing Sea Grape and Hearing Extracts

In Table 13, materials 2, 3, and 5 were mixed and stirred and heated to 40 to 45 ° C., and then introduced into the production unit, followed by an emulsifier, and materials 4, 6 and 7 were added to the production unit and emulsified. After emulsification, the mixture was cooled to 35 ° C. while stirring using a stirrer, and 1 substance was added thereto, cooled to 25 ° C., and aged at 25 ° C. for 3 days to prepare the title nutrition serum.

Formulation Example  4. Sea grapes  And essence containing hearing mixed extract

1 kg of the hydrosome essence containing the sea grapes and the auditory mixed extract (Example 1) was prepared using the following preparation method with the content of the following [Table 14].

[Table 14] Essence Composition Containing Sea Grape and Acoustic Mixed Extract

In Table 14, materials 2, 3, 4, 5 and 6 were homogenized at a constant temperature to prepare nonionic amphiphilic lipids. After mixing the nonionic amphiphilic lipids with materials 1, 7, 8 and 14 and homogenizing it at a constant temperature, it is passed through a microfluidizer, and then the material is heated to 50 to 60 ° C and slowly added to homogenize it. The microfluidizer was passed again. Thereafter, materials 10, 11, 12, and 13 were added to disperse, stabilize, and ripen to prepare the title essence.

Experimental Example  10. Confirmation of formulation stability

The formulations prepared in Formulation Examples 1 to 4 were stored and observed for 12 weeks in opaque glass containers in indoors, refrigerators and incubators kept constant at room temperature (25 ° C), refrigerated (4 ° C) and constant temperature (50 ° C). (Discoloration, deodorization and separation) and stability was confirmed. The results are shown in Table 14.

Table 15 Formulation Stability Check

<Formulation stability grade>

0: no change, 1: slight change, 2: change, 3: extreme change

As shown in Table 15, all of Formulation Examples 1 to 4 formulation, it was confirmed that the discoloration, odor and separation phenomenon is stable under 25 ℃, 4 ℃ and 50 ℃ temperature conditions, it is stable.

Claims (8)

Anti-aging cosmetic composition comprising sea grapes and hearing mixed extract. The anti-aging cosmetic composition according to claim 1, wherein the sea grape and auditory mixed extract is a mixed extract of sea grape extract and auditory extract. The anti-aging cosmetic composition of claim 1, wherein the sea grapes and the auditory mixed extract are extracted with one or more solvents selected from water, alcohols having 1 to 4 carbon atoms, or mixtures thereof. The anti-aging cosmetic composition according to claim 1, wherein the mixed dry weight ratio of the sea grapes and the auditory mixed extract is 1: 2 to 2: 1. The anti-aging cosmetic composition according to claim 4, wherein the mixed dry weight ratio of the sea grapes and the auditory mixed extract is 1: 1. The cosmetic composition according to claim 1, comprising 0.01 to 5% by weight of the sea grape and auditory mixed extract, based on the total dry weight. The method according to claim 1, wherein the anti-aging is antioxidation, collagen production, collagenase expression inhibition, elastase expression inhibition, NO production inhibition, prostaglandin production inhibition, TNF-α production inhibition, moisturizing or final glycation product inhibition Anti-aging cosmetic composition according to any one or more selected. According to claim 1, wherein the cosmetic composition has a softening lotion, nourishing lotion, nourishing cream, massage cream, essence, pack, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray or powder Cosmetic composition.