KR101978570B1 - Composition for anti-aging - Google Patents

Abstract

The present invention relates to an anti-aging cosmetic composition comprising extracts of L. codica, L. chinensis, L. japonica, Rhizophora, and meteorite. In particular, the composition according to the present invention exhibits an effect of protecting skin from aging factors such as wrinkle prevention or improvement, skin moisturizing power, skin elasticity improvement, skin barrier improvement, antioxidative effect, various oxidative toxicity and phototoxicity.

Description

Anti-aging composition {COMPOSITION FOR ANTI-AGING}

The present invention relates to an anti-aging cosmetic composition comprising extracts of L. codica, L. chinensis, L. japonica, Rhizophora, and meteorite.

Skin aging has aging due to internal and external factors. The aging due to the internal factors is a natural phenomenon that occurs with aging without special environmental factors. On the other hand, aging caused by external factors is aging due to environmental factors including ultraviolet rays, smoking, stress, chemical substances, pollution, etc., and is mainly caused by aging due to ultraviolet rays.

In the aging process, active oxygen corresponding to free radicals is involved. The active oxygen plays an important role in removing or suppressing intracorporeal substances such as pathogens or viruses. On the other hand, active oxygen species are energetically large and highly reactive, and thus, when overproduced, enzymatic and non-enzymatic antioxidant defense It is known that the system breaks down and accelerates skin aging (Yang et al., J. Soc. Cosmet Sci. Korea, 34: 275-286, 2008). Active oxygen and free radicals have been reported to occur when the skin is exposed to ultraviolet light, resulting in phototoxicity, photosensitivity, skin aging and various immune related diseases (Norins, J. Invest. Dermatol., 39: 445, 1962; Cadenas, Ann. Rev. Biochem., 58: 79, 1989).

In the skin aging process, lipids, proteins, polysaccharides and nucleic acids, which are major constituents of the skin, are oxidized, thereby destroying skin cells and tissues. In particular, oxidation of proteins damages skin connective tissues such as collagen, elastin, proteoglycans, fibronectin, and hyaluronic acid, and reduces the inflammatory response and skin elasticity. Furthermore, when the damage becomes worse, DNA mutation leads to mutation, cancer induction, and immune function depression.

Therefore, in order to quickly recover aged skin and prevent aging of skin, it is necessary to protect the cell membrane by destroying the active oxygen that is mediated by the metabolic process and ultraviolet irradiation of the body, There is a need to do.

Examples of free radicals and reactive oxygen species (ROS) are O ₂ ^- (superoxide anion radical), OH (hydroxyl radical), ¹ O ₂ (singlet oxygen) and H ₂ O ₂ . Antioxidants have been developed to prevent skin aging by inhibiting enzymatic and nonenzymatic oxidation by these reactive oxygen species, and can be broadly divided into synthetic antioxidants and natural antioxidants.

Representative examples of the synthetic antioxidants include butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), etc. These antioxidative substances are excellent in antioxidative effect but have a high carcinogenicity and can adversely affect the human body. It is known that there is a disadvantage that the enzymatic antioxidant effect is not good compared with the effect.

Recently, there have been many studies on natural antioxidants that can overcome the problems of synthetic antioxidants. The natural antioxidants are still less effective than synthetic antioxidants in terms of side effects such as carcinogenicity, but still have limitations in meeting both enzymatic and non-enzymatic antioxidant effects.

On the other hand, in addition to active oxygen, senescence involves an enzyme called MMP (Matrix metalloproteinase). In healthy skin, the synthesis and degradation of extracellular matrix such as collagen are properly controlled in vivo, while as aging proceeds, the synthesis of collagen is reduced and the elasticity of the skin is lowered and wrinkles are formed. At this time, since the MMP enzyme exhibits an action of decomposing collagen, when the expression of MMP is promoted by ultraviolet irradiation or the like, aging of the skin is accelerated.

It is possible to effectively suppress the expression of MMP that affects skin aging and to exhibit both an increase in both of the enzymatic antioxidative effect and the non-enzymatic antioxidative effect that the conventional natural antioxidant has not attained even without the side effects exhibited by the conventional synthetic antioxidants The need for anti-aging cosmetic compositions remains largely in the art.

The inventors of the present invention recognized the problems of the prior art and participated in various mechanisms related to aging of the skin to thereby exhibit an excellent anti-aging effect.

According to one embodiment of the present invention, there is provided an anti-aging cosmetic composition comprising extracts of Sweetpotato, Lycopodium, Lilium, Rhodiola, and meteorite.

As used herein, the term " Wa pigweed " refers to a perennial herb belonging to the Lycopersicon esculentum and its scientific name is Chenopodium album.

As used herein, the term " herbaceous plant " refers to a perennial plant belonging to the plantain tree family, and its scientific name is Plantago asiatica.

As used herein, the term " perilla " refers to a thin bark wrapped around a chestnut tree of the chestnut (Castanea crenata) belonging to the oakwood family.

As used herein, the term " rhubarb, " refers to a perennial plant belonging to the monocotyledonous plant family, and its scientific name is Rehmannia glutinosa.

As used herein, the term " meteorite " refers to rocks that have fallen from space to surface, with meaning generally understood and used in the art.

The meteorites can be classified into stony meteorite, stony iron meteorite and iron meteorite depending on their constituents.

The above-mentioned meteorite is composed mainly of silicate minerals. Most of the meteorites recovered from witnessing fall are seismic meteorites. The rocky meteorite can be divided into chondrite with chondrule and Achondrite with no chondrite. It is known that the acondrite has more silica (SiO ₂ ) than the chondrite, has almost no nickel-iron (Ni-Fe) metal phase, is generally more assembled than the chondrite, and chemical coordination or mineral control is close to the earth rock.

The meteorite includes more than 25% of nickel-iron (Ni-Fe) and silicate minerals, respectively, and accounts for about 4% of all existing meteorites. According to the composition of the silicate minerals, the meteorite meteorite is composed of phallasite (olivine stone meteorite), mesosideite (pyroxene plagioclase), rhodanite (olivine) . Most of the stalagmite is palaceite and mesosideite, while rodaneite and seaerfire are similar to palaceite.

The iron meteorite is mainly composed of iron and nickel, and is also referred to as iron. It accounts for about 6% of the meteorites falling on Earth.

The meteorite used in the composition according to the present invention may be a meteorite meteorite, a meteorite meteorite, an iron meteorite, or any mixture thereof.

The composition according to the present invention includes extracts of Sweetpotato, Lycoris spp., Lycopodium, Rhizophora and meteorite. Here, the cultivated, harvested, or marketed products such as Sweetpotato, Chrysoprasea, Rhododendron and Rhododendron can be used without limitation. Sweetpotato, Chaetodon, and Rhubarb can be flowers, leaves, seeds, fruits, roots and / or stems.

As used herein, the term " extract " means a product obtained by extracting with an appropriate extraction solvent, including an extract obtained by extraction, a dilute or concentrated solution of the extract, a dried product obtained by drying the extract, Purified water, or fraction thereof. Preferably, the extract according to the present invention is obtained by extracting a complex of starch, primrose, clover, rhizomes and meteorites (for example, in the form of a combination of starch, primrose, clove, rhizosphere and meteorite in dry form).

1 to 96% by weight of rhizome, 1 to 96% by weight of rhizome, 1 - 96% by weight of rhizome, 1 - 96% by weight of rhizome, 1 - 96% by weight.

The extract according to the present invention can be prepared by using a general extraction method known in the art. For example, a method such as hot water extraction, hot water extraction, cold extraction, reflux cooling extraction, ultrasonic extraction, have. Particularly, the hot pressurization extraction method is preferable, and it is more preferable to perform the roasting pretreatment before the hot pressurization extraction.

The extraction solvent used for preparing the extract according to the present invention includes water; Lower alcohols having 1 to 4 carbon atoms such as methanol, ethanol, propyl alcohol and butyl alcohol; Polyhydric alcohols such as glycerin, butylene glycol and propylene glycol; Hydrocarbon solvents such as chloroform, methyl acetate, ethyl acetate, benzene, hexane, diethyl ether, petroleum ether, and dichloromethane; Or mixed forms thereof, but are not limited thereto.

The cosmetic composition according to the present invention contains extracts of Sweetpotato, Lycoris, Rhizopus, Rhizopus, and meteorites as active ingredients. The content of the extract in the cosmetic composition may be suitably determined in consideration of various factors such as the purpose of use, the period of use, the type of formulation, the route of administration, and the degree of aging of the subject. For example, the content of the extract may be 0.0001 to 20% by weight based on the total weight of the cosmetic composition.

The cosmetic composition according to the present invention excellently exhibits an effect of protecting the skin from aging factors such as wrinkle prevention or improvement, skin moisturizing power, skin elasticity improvement, skin barrier improvement, antioxidative effect, various oxidative toxicity and phototoxicity. As such, it influences the various mechanisms of skin aging, resulting in an excellent anti-aging effect.

The term " improvement " as used herein may be interpreted to include not only all actions that reduce, retard, alleviate or inhibit the parameters associated with an aging-related condition, such as symptoms or progression, but also reverse actions.

The cosmetic composition according to the present invention can be manufactured in the form of a general emulsified formulation, a solubilized formulation, or the like, using a generally known production method. At this time, the cosmetic composition of the present invention can be used as a cosmetic composition, such as patches, yogurts, skin adhesive gels, creams, packs, lotions, essences, sprays, masks, foundations, makeup bases, (W / S) type, water-in-silicone (S / W) type, solid type, water-in-oil type, Liquid form, and the like, and a commonly used cosmetic preparation method can be applied.

The cosmetic composition according to the present invention may further contain an additional component in addition to the above extract for enhancing the efficacy. For example, the additional ingredient may be a component or ingredient exhibiting a skin anti-aging effect, so long as it does not abate or reduce the efficacy of the extract. Optionally, it may further comprise adjuvants, carriers and the like commonly used in the cosmetics field. Alternatively, ingredients typically used to add or enhance the cosmetic function may also be added. For example, there may be mentioned a stabilizer, an emulsifier, a thickening agent, a moisturizer, a liquid crystal film stiffening agent, a pH adjusting agent, an antibacterial agent, a water-soluble polymer, a coating agent, a metal ion sequestering agent, an amino acid, an organic amine, a polymer emulsion, At least one aqueous additive selected from antioxidant auxiliaries, preservatives, perfumes and the like; And one or more oily additives selected from oils, waxes, hydrocarbonaceous oil, higher fatty acid oil, higher alcohol, synthetic ester oil and silicone oil.

The cosmetic composition according to the present invention is characterized by comprising the steps of: adding a solvent to a complex of starch, Chaetoche, zephyr, zucchini and meteorite; Performing an extraction process; Separating and filtering the extract; And lyophilizing or concentrating the resulting product.

Optionally, prior to the step of adding the solvent, a complex of Sweetpotato, Chrysoprasea, Pepi, Rhizophorus and meteorite may be pretreated. The fogue pretreatment may be roasting.

The composite may be in a dried form, a concentrated form, a solution form, or the like.

According to the present invention, the extracts of the complex consisting of Kwangju, Chuncheoncho, Yulpie, Rhizophorus, and meteorite have significant synergistic effects compared with the individual extracts.

Specifically, the extracts of the complex consisting of Kwangju, Chaesinjiae, Yulpie, Rhizophora, and meteoricum according to the present invention are superior in yield to individual extracts.

In addition, the content of representative antioxidant components such as phenolic compounds and flavonoids is much higher. In particular, the free radical scavenging test demonstrated excellent non-enzymatic antioxidant activity, and demonstrated excellent enzymatic antioxidant activity through reactive oxygen scavenging test. In addition, the extract according to the present invention does not cause adverse effects such as carcinogenicity as shown in the synthetic antioxidants, because all the constituents are composed of natural substances. Thus, the non-enzymatic and enzymatic antioxidative effects are excellent both in the absence of adverse effects on the human body and in the fact that the problems of conventional synthetic antioxidants and natural antioxidants are solved.

Furthermore, when ultraviolet light, which is one of the main causes of skin aging, is irradiated, it rapidly restores the cell viability of fibroblasts synthesizing extracellular matrix and collagen, and can mitigate cytotoxicity caused by ultraviolet rays.

In addition, although expression of collagenase MMP (matrix metalloproteinase) is promoted by ultraviolet irradiation, the extract according to the present invention can significantly inhibit the expression of MMP, thereby effectively preventing collagen degradation, .

In particular, the compositions of the present invention have been directly demonstrated in vivo experiments that reduce the depth of wrinkles of the subject, provide excellent damage recovery rates of skin barrier and minimize loss of transdermal water.

Accordingly, the composition according to the present invention has an effect of protecting the skin from aging factors such as wrinkle prevention or improvement, skin moisturizing power, skin elasticity improvement, skin barrier improvement, antioxidative effect, various oxidative toxicity and phototoxicity, And exhibits excellent anti-aging effect by acting in various ways against aging. The composition according to the present invention can be applied variously to cosmetics, external preparation for skin, cosmetics and the like.

FIG. 1 is a graph showing inhibition rate (%) of MMP-1 expression when human dermal fibroblasts are irradiated with ultraviolet light.

Hereinafter, a specific example will be described in order to facilitate understanding of the present invention. However, the following examples should not be construed as limiting the invention, but ordinary variations of the ordinary skill in the art are possible within the scope of the invention.

The composition of the dried complex natural material used in the following examples was constituted by a weight ratio of Starch: Chaesunju: Ujipi: Rhizo: Meteorite = 1: 1: 2: 1: 1. The meteorite used here is a meteorite in the northwest part of the Sahara Desert, estimated to be formed about 4.5 billion years ago, similar to the age of the Earth, according to carbon dating.

Example 1

30 g of the dried complex natural product was roasted with a roaster (FEC006, Biotech, Korea) at 100 ° C for 20 minutes, and 570 g of water was added to the reactor. Subsequently, the immersed sample was subjected to extraction at 80 DEG C for 3 hours under pressure using a reactor. After centrifugation at 8,000 rpm and 15 ° C for 20 minutes using a centrifuge (Supra 22K, Hanil, Korea), the separated extracts were filtered through 0.8 μm and 0.45 μm filters. As a result, the obtained filtrate was subjected to freeze-drying at -21 캜 and then freeze-dried at -80 캜 for 48 hours using a freeze dryer (Bondiro, Ilshin, Korea).

The resulting material was referred to as " sample 1 " and used in the following test examples.

Example 2

30 g of the dried complex natural material was charged to the reactor and 570 g of water was added. Subsequently, the immersed sample was subjected to extraction at 80 DEG C for 3 hours under pressure using a reactor. After centrifugation at 8,000 rpm and 15 ° C for 20 minutes using a centrifuge (Supra 22K, Hanil, Korea), the separated extracts were filtered through 0.8 μm and 0.45 μm filters. As a result, the obtained filtrate was subjected to freeze-drying at -21 캜 and then freeze-dried at -80 캜 for 48 hours using a freeze dryer (Bondiro, Ilshin, Korea).

The resulting material was referred to as " Sample 2 " and used in the following test examples.

Example 3

30 g of the dried complex natural product was subjected to a steam roasting treatment using a roaster (FEC006, Biotech, Korea) at 100 ° C for 20 minutes, and 570 g of a 70% aqueous ethanol solution was added to the reactor. Subsequently, the immersed sample was subjected to extraction at 80 DEG C for 3 hours under pressure using a reactor. After centrifugation at 8,000 rpm and 15 ° C for 20 minutes using a centrifuge (Supra 22K, Hanil, Korea), the separated extracts were filtered through 0.8 μm and 0.45 μm filters. The resulting filtrate was rotary evaporated to dryness, freeze-pretreated at -21 ° C, and freeze-dried at -80 ° C for 48 hours using a freeze dryer (Bondiro, Ilshin, Korea).

The resulting material was referred to as " sample 3 " and used in the following test examples.

Example 4

30 g of the dried complex natural material was placed in a reactor and 570 g of 70% aqueous ethanol solution was added. Subsequently, the immersed sample was subjected to extraction at 80 DEG C for 3 hours under pressure using a reactor. After centrifugation at 8,000 rpm and 15 ° C for 20 minutes using a centrifuge (Supra 22K, Hanil, Korea), the separated extracts were filtered through 0.8 μm and 0.45 μm filters. The resulting filtrate was rotary evaporated to dryness, freeze-pretreated at -21 ° C, and freeze-dried at -80 ° C for 48 hours using a freeze dryer (Bondiro, Ilshin, Korea).

The resulting material was referred to as " sample 4 " and used in the following test examples.

Example 5

30 g of the dried complex natural material was placed in a beaker and 570 g of water was added. Subsequently, hydrothermal treatment was performed at 80 캜 for 3 hours using a water bath (HB-205WP, Hanbaek, Korea). After centrifugation at 8,000 rpm and 15 ° C for 20 minutes using a centrifuge (Supra 22K, Hanil, Korea), the separated extracts were filtered through 0.8 μm and 0.45 μm filters. As a result, the obtained filtrate was subjected to freeze-drying at -21 캜 and then freeze-dried at -80 캜 for 48 hours using a freeze dryer (Bondiro, Ilshin, Korea).

The resulting material was referred to as " sample 5 " and used in the following test examples.

Example 6

30 g of the dried complex natural material was placed in a beaker and 570 g of water was added. Then, ultrasonication was performed for 1 hour at 20 kHz, 750 W, and 25 캜 using an ultrasonic device (5510R-DTH, BARANSON, USA). After centrifugation at 8,000 rpm and 15 ° C for 20 minutes using a centrifuge (Supra 22K, Hanil, Korea), the separated extracts were filtered through 0.8 μm and 0.45 μm filters. As a result, the obtained filtrate was subjected to freeze-drying at -21 캜 and then freeze-dried at -80 캜 for 48 hours using a freeze dryer (Bondiro, Ilshin, Korea).

The resulting material was referred to as " Sample 6 " and used in the following test examples.

Example 7

30 g of the dried complex natural product was placed in a beaker and 570 g of a 70% aqueous ethanol solution was added. Subsequently, hydrothermal treatment was performed at 80 캜 for 3 hours using a water bath (HB-205WP, Hanbaek, Korea). After centrifugation at 8,000 rpm and 15 ° C for 20 minutes using a centrifuge (Supra 22K, Hanil, Korea), the separated extracts were filtered through 0.8 μm and 0.45 μm filters. The resulting filtrate was rotary evaporated to dryness, freeze-pretreated at -21 ° C, and freeze-dried at -80 ° C for 48 hours using a freeze dryer (Bondiro, Ilshin, Korea).

The resulting material was referred to as " sample 7 " and was used in the following test examples.

Example 8

30 g of the dried complex natural product was placed in a beaker and 570 g of a 70% aqueous ethanol solution was added. Then, ultrasonication was performed for 1 hour at 20 kHz, 750 W, and 25 캜 using an ultrasonic device (5510R-DTH, BARANSON, USA). After centrifugation at 8,000 rpm and 15 ° C for 20 minutes using a centrifuge (Supra 22K, Hanil, Korea), the separated extracts were filtered through 0.8 μm and 0.45 μm filters. The resulting filtrate was rotary evaporated to dryness, freeze-pretreated at -21 ° C, and freeze-dried at -80 ° C for 48 hours using a freeze dryer (Bondiro, Ilshin, Korea).

The resulting material was referred to as " sample 8 " and used in the following test examples.

Example 9

30 g of dried sweetpotato, liquorice, persimmon, persimmon, and meteorite were placed in separate beakers and 570 g of water was added. Subsequently, hydrothermal treatment was performed in a water bath (HB-205WP, Hanbaek, Korea) at 80 ° C for 1 hour. After centrifugation at 8,000 rpm and 15 ° C for 20 minutes using a centrifuge (Supra 22K, Hanil, Korea), the separated extracts were filtered through 0.8 μm and 0.45 μm filters. As a result, the obtained filtrate was subjected to freeze-drying at -21 캜 and then freeze-dried at -80 캜 for 48 hours using a freeze dryer (Bondiro, Ilshin, Korea).

Samples 9-1 "," 9-2 "," 9-3 "," 9-4 "," 9- "and" 9-2 "were obtained in the order of Kwangju, 5 " and was used in the following test examples.

Example 10

Dried bean sprouts, spinach, yulpie, rhubarb, and 30 g of meteorite were placed in separate beakers, and 570 g of 70% ethanol aqueous solution was added. Subsequently, hydrothermal treatment was performed in a water bath (HB-205WP, Hanbaek, Korea) at 80 ° C for 1 hour. After centrifugation at 8,000 rpm and 15 ° C for 20 minutes using a centrifuge (Supra 22K, Hanil, Korea), the separated extracts were filtered through 0.8 μm and 0.45 μm filters. The resulting filtrate was rotary evaporated to dryness, freeze-pretreated at -21 ° C, and freeze-dried at -80 ° C for 48 hours using a freeze dryer (Bondiro, Ilshin, Korea).

10-2 "," 10-3 "," 10-4 "," 10-2 ", and" 10-2 "were obtained in the order of Kwangju, 5 " and was used in the following test examples.

Test Example 1. Extraction yield

In order to compare the yield of the extract prepared in the above examples, the extract yield relative to the input of the dry matter was calculated. The results are shown in Table 1 below.

Yield (%)
Sample 1 15.7 Sample 3 18.4 Sample 2 12.7 Sample 4 11.9 Sample 5 9.3 Sample 7 10.2 Sample 6 8.4 Sample 8 9.3 Sample 9-1 6.4 Sample 10-1 8.1 Sample 9-2 7.1 Sample 10-2 8.7 Sample 9-3 6.5 Sample 10-3 6.8 Samples 9-4 6.9 Samples 10-4 7.3 Sample 9-5 0.1 Sample 10-5 0.1

As shown in Table 1 above, the extract of the composite showed a higher yield than the extract of the single regardless of the extraction method and the extraction solvent.

In addition, hot press extraction showed higher yield than hot water (reflux) extraction and ultrasonic extraction. In particular, it was confirmed that the yields of samples 1 and 3 pretreated prior to high temperature pressure extraction were the highest.

Test Example 2. Contents of antioxidant component

The contents of antioxidant components in the extracts prepared in the above Examples were confirmed. For this purpose, the contents of phenolic compounds and flavonoids, which are well known in the art, are shown in each sample to show antioxidative effects.

First, the total content of phenolic compounds was determined as follows.

After adding 10 mL of distilled water to 1 mL of each sample, add 2 mL of Folin-Ciocalteu phenol reagent (Sigma), mix and react at room temperature for 5 minutes. 2 mL of 20% sodium carbonate was added thereto, mixed and reacted at room temperature for 1 hour. The absorbance was then measured at 680 nm using a microplate reader (UVT-06685, Thermo max, USA). At this time, gallic acid (Sigma, USA) was used as the indicator material, and a calibration curve for the concentration of the indicator was prepared, and the absorbance value of each sample was substituted to calculate the total content of phenolic compounds. The results are shown in Table 2 below.

Total phenol content (mg / g extract)
Sample 1 171.4 Sample 3 193.0 Sample 2 145.6 Sample 4 151.5 Sample 5 129.4 Sample 7 138.1 Sample 6 128.8 Sample 8 131.9 Sample 9-1 81.3 Sample 10-1 92.2 Sample 9-2 65.5 Sample 10-2 77.6 Sample 9-3 91.4 Sample 10-3 102.9 Samples 9-4 89.4 Samples 10-4 98.2 Sample 9-5 0.0 Sample 10-5 0.0

Next, the total content of flavonoids was measured as follows.

1.5 mL of each sample was mixed with 2% AlCl ₃ 6H ₂ O dissolved in an equal amount of methanol, and the mixture was reacted at room temperature for 10 minutes. The absorbance was then measured at 367 nm using a microplate reader (UVT-06685, Thermo max, USA). At this time, catechin (Sigma, USA) was used as the indicator material and the total content of flavonoids was calculated by substituting the absorbance value of each sample with a calibration curve according to the concentration of the indicator substance. The results are shown in Table 3 below.

Total flavonoid content (mg / g extract)
Sample 1 134.6 Sample 3 167.5 Sample 2 110.1 Sample 4 124.3 Sample 5 94.3 Sample 7 101.6 Sample 6 89.1 Sample 8 99.2 Sample 9-1 72.5 Sample 10-1 78.4 Sample 9-2 58.4 Sample 10-2 65.9 Sample 9-3 81.0 Sample 10-3 95.3 Samples 9-4 77.6 Samples 10-4 67.4 Sample 9-5 0.0 Sample 10-5 0.0

As shown in Tables 2 and 3, regardless of the extraction method and the extraction solvent, the content of the phenolic compound and flavonoid was higher in the extract of the composite than in the extract of the single product.

Compared with hot water (reflux) extraction and ultrasonic extraction, high pressure extraction showed higher contents of phenolic compounds and flavonoids. In particular, it was confirmed that the contents of phenolic compounds and flavonoids were highest in Samples 1 and 3 pretreated prior to high temperature extrusion. The high content of phenolic compounds and flavonoids in the extract according to the present invention means that the extract according to the present invention has a high antioxidative capacity.

Test Example 3. Antioxidant effect

The antioxidant ability of the present invention was examined in order to test the skin aging improving effect. For this, free radical scavenging test (non-enzymatic antioxidant effect test) and active oxygen scavenging test (enzymatic antioxidant effect test) were carried out.

(1) Free radical scavenging test (non-enzymatic antioxidant effect test)

To confirm the antioxidant effect, the free radical scavenging test frequently used in the art is based on the principle that the absorbance of stable 2,2-diphenyl-1-picryl-hydrazine (DPPH) shows a maximum value at 540 nm. Therefore, as the free radical DPPH is cleared by the sample and becomes a purple to transparent color, that is, the free radical scavenging rate is increased, the absorbance at 540 nm is reduced.

First, 1 mL of a solution prepared by diluting the sample prepared in the above Example with methanol was mixed with 1 mL of 0.1 mM DPPH (Sigma) solution. After incubation at 37 ° C for 15 minutes, the absorbance was measured at 540 nm using a microplate reader (UVT-06685, Thermo max, USA).

In the control group, 1 ml of DPPH and 1 ml of methanol were added and the absorbance was measured in the same manner. 1 ml of methanol and 1 ml of the sample were added to obtain the respective color correction values for the sample and the control.

The free radical scavenging ratios were calculated using the following formula 1 and shown in Table 4. In Table 4, SC ₅₀ refers to the concentration of the sample required to 50% free radical scavenging. Here, the smaller the value of SC ₅₀ , the higher the antioxidant activity.

[Formula 1]

Free radical scavenging rate (%) = 100 - ((absorbance of sample / absorbance of control) x 100)

Free radical scavenging test (SC ₅₀ , / / Ml)
Sample 1 46.8 Sample 3 38.4 Sample 2 62.1 Sample 4 56.3 Sample 5 80.4 Sample 7 72.0 Sample 6 88.9 Sample 8 79.4 Sample 9-1 103.5 Sample 10-1 94.3 Sample 9-2 98.4 Sample 10-2 86.1 Sample 9-3 102.0 Sample 10-3 97.2 Samples 9-4 92.7 Samples 10-4 88.8 Sample 9-5 1,512.2 Sample 10-5 1,325.4

As shown in Table 4, regardless of the extraction method and the extraction solvent, the SC ₅₀ value of the extract of the composite was smaller than that of the single extract.

In addition, SC ₅₀ values were significantly lower in hot press extraction than in hot water (reflux) extraction and ultrasonic extraction. In particular, the lowest SC ₅₀ values were found in samples 1 and 3 pretreated prior to hot press extraction, indicating that samples 1 and 3 had the highest antioxidant activity.

(2) Active oxygen scavenging test (enzymatic antioxidant effect test)

The active oxygen scavenging test is based on the principle that active oxygen is generated by enzymatic reaction of xanthine / xanthine oxidase (Sigma). By measuring the absorbance change by oxidation of nitroblue tetrazolium (NBT) by active oxygen, the scavenging ability of active oxygen can be confirmed.

Mix the vial with a vortex mixer (Type 37600 Mixer, Mini Mix, pH 7.4) and mix with 2.4 mL of Na ₂ CO ₃ , 0.1 mL of Sigma, 0.1 mL of EDTA, 0.1 mL of BSA (bovine serum albumin, Sigma), 0.1 mL of NBT, USA) and left at 25 ° C for 10 minutes. Subsequently, 0.1 ml of xanthine oxidase was added, and the reaction was carried out at 25 DEG C for 20 minutes. The reaction was stopped by adding 6 mM CuCl ₂ and absorbance was measured at 540 nm using a microplate reader (UVT-06685, Thermo max, USA).

In the control group, the third absorbance was used instead of the sample solution, the absorbance was measured in the same manner, and the third and fourth distilled water were added instead of the xanthine oxidase solution to obtain the respective color correction values for the sample and the control group.

The active oxygen scavenging ratio was calculated using the following formula 2 and shown in Table 5 below. In Table 5, IC ₅₀ refers to the concentration of the sample required to remove 50% of the active oxygen. Here, the smaller the IC ₅₀ value, the higher the antioxidant activity.

[Formula 2]

Ratio of active oxygen scavenging (%) = 100 - ((absorbance of sample / absorbance of control) x 100)

Reactive oxygen scavenging test ₅₀ , / / Ml)
Sample 1 58.4 Sample 3 46.3 Sample 2 75.8 Sample 4 68.9 Sample 5 96.5 Sample 7 75.4 Sample 6 111.3 Sample 8 93.0 Sample 9-1 125.1 Sample 10-1 113.8 Sample 9-2 105.6 Sample 10-2 100.1 Sample 9-3 134.8 Sample 10-3 95.3 Samples 9-4 122.2 Samples 10-4 118.5 Sample 9-5 2,568.8 Sample 10-5 2,105.3

As shown in Table 5, regardless of the extraction method and the extraction solvent, the IC ₅₀ of the extract of the composite was lower than that of the extract of the single substance.

In addition, the IC ₅₀ value was significantly lower in hot press extraction than in hot water (reflux) extraction and ultrasonic extraction. In particular, the lowest IC ₅₀ values were found in samples 1 and 3 pretreated prior to hot press extraction, indicating that samples 1 and 3 had the highest antioxidant activity.

Test Example 4. Mitigation Effect of Cytotoxicity by Ultraviolet Irradiation

In order to confirm cytotoxicity mitigation effect of the complex natural extract according to the extraction method of the present invention by ultraviolet irradiation, the following experiment was conducted.

Fibroblasts were dispensed at 1 × 10 ⁵ cells in 24-well plates, and then adhered for 24 hours. Each well was washed once with PBS, and 500 μL of PBS was added to each well. Then, ultraviolet rays of 10 mJ / cm ² were irradiated to the fibroblasts using an ultraviolet B (UVB) lamp (Model: F15T8, UVB 15W, Sankyo Dennki, Japan). Then PBS was removed and 1 mL of cell culture medium (DMEM supplemented with 10% FBS) was added.

Here, the samples produced in the above examples were treated at a concentration of 0.01%, and then cultured for 24 hours. Subsequently, the medium was removed, and 500 μL of the cell culture medium and 60 μL of the MTT solution (2.5 mg / mL) were added to each well, followed by culturing in a 37 ° C. CO ₂ incubator for 2 hours. The medium was removed and 500 μL of isopropanol-HCl (0.04 N) was added. The cells were lysed by shaking for 5 minutes, and 100 μL of the supernatant was transferred to a 96-well test plate. The absorbance at 565 nm was then measured using a microplate reader (UVT-06685, Thermo max, USA).

The cell viability (%) was calculated by the following formula 3, and the cytotoxicity reduction rate by ultraviolet was calculated by the following formula 4. Table 6 shows cytotoxicity reduction rates by ultraviolet light.

[Formula 3]

Cell survival rate (%) = [(St-Bo) / (Bt-Bo)] x 100

Bo: Absorbance at 565 nm of the well, which was color-developed only in the cell culture medium

Bt: absorbance at 565 nm of the well, in which the well not treated with the sample was subjected to the color reaction,

St: Absorbance at 565 nm of the well, in which the well treated with the sample was subjected to color reaction

[Formula 4]

(%) = [1- (St-Bo) / (Bt-Bo)] x 100

· Bo: Cell viability of wells not irradiated with ultraviolet light and not treated with samples

Bt: Cell viability of wells irradiated with ultraviolet light and not treated with the sample

St: Cell viability of the well irradiated with ultraviolet light and treated with the sample

Cytotoxic Relaxation Rate (%)
Sample 1 40.2 Sample 2 34.8 Sample 3 51.3 Sample 4 43.6 Sample 5 12.3 Sample 6 11.7 Sample 7 18.4 Sample 8 18.1

As shown in Table 6, the extract according to the present invention effectively protects against cytotoxicity caused by ultraviolet rays and exhibits a high cell survival rate. Particularly, the extract obtained by hot press extraction is superior to the extract obtained by hot water (reflux) extraction and ultrasonic extraction.

Test Example 5. Effect of Inhibiting MMP-1 Expression after UV Irradiation

As mentioned above, the enzyme MMP (Matrix metalloproteinase), which degrades collagen, can be promoted by ultraviolet irradiation. In this case, collagen degradation is activated, resulting in reduced skin elasticity and wrinkles .

In this test example, the effect of suppressing the expression of MMP-1 by the sample was confirmed by measuring the concentration of MMP-1 after UV irradiation and sample addition associated with skin elasticity reduction, wrinkle formation, and aging. Enzyme immunoassay (ELISA) was performed as follows.

UVA was irradiated to human dermal fibroblasts at an energy of 5 J / cm ² using a UV chamber. In order to determine ultraviolet irradiation dose and incubation time, preliminary experiments confirmed the conditions under which the amount of MMP expression in the fibroblasts was maximized. The negative control was wrapped in silver foil and kept in the UVA environment for the same time. UVA emission was measured using a UV radiometer. During UVA irradiation, the cells were kept in a previously dispensed state. UVA was irradiated and the medium was replaced with a medium containing the sample. After culturing for 24 hours, the medium was recovered and coated on a 96-well plate.

As the primary antibody, MMP-1 (Ab-5) monoclonal antibody and MMP-2 (Ab-3) monoclonal antibody were treated and reacted at 37 ° C for 60 minutes. The secondary antibody, anti-mouse IgG (alkaline phosphatase-conjugated), was reacted for 60 minutes and reacted with alkaline phosphatase substrate solution (1 mg / ml ρ-nitrophenyl phosphate in diethanolamine buffer) at room temperature for 30 minutes. The absorbance was then measured at 405 nm using a microplate reader (UVT-06685, Thermo max, USA).

At this time, as a negative control group, a sample without addition of the sample was used, and retinol, which is known to have excellent inhibitory effect on MMP-1 expression, was used as a positive control group. The results are shown in Table 7 below and are shown graphically in FIG.

MMP-1 expression inhibition rate (%)
Sample 1 25.4 Sample 2 18.8 Sample 3 36.6 Sample 4 29.9 Sample 5 12.1 Sample 6 9.7 Sample 7 15.4 Sample 8 11.3 Positive Control - Retinol 22.5

As shown in Table 7, the hot press extraction showed a better effect than the inhibitory rate of 22.5% of MMP-1 expression of retinol used as a positive control. This means that the extract according to the present invention can inhibit the expression of MMP enzyme to effectively block collagen degradation, thereby inhibiting skin elasticity and wrinkle formation, thereby preventing skin aging.

Test Example 6. Anti-aging effect

To test the anti-aging effect of the composition according to the present invention, nutritional creams were prepared according to the ingredients and contents of Table 8 below.

number ingredient Formulation Example 1 Formulation Comparative Example 1 One Pro-type glycerin monostearate 2.0 2.0 2 Stearic acid 1.5 1.5 3 Cetearyl alcohol 2.2 2.2 4 Wax 1.0 1.0 5 Squalane 3.0 3.0 6 Hardened vegetable oil 1.0 1.0 7 Sorbitan stearate 0.6 0.6 8 Mineral oil 5.0 5.0 9 Polysorbate 60 1.5 1.5 10 Dimethicone 1.0 1.0 11 Trioctanoin 5.0 5.0 12 Betaine 3.0 3.0 13 Triethanolamine 1.0 1.0 14 glycerin 5.0 5.0 15 Sodium hyaluronate 3.0 3.0 16 Sample 1 0.1 - 17 Distilled water Balance Balance 18 Preservative, fragrance, pigment a very small amount a very small amount

First, the following experiment was conducted to measure the wrinkle improvement effect. The nutritional creams of Formulation Example 1 and Formulation Comparative Example 1, prepared according to Table 8 above, were applied to the crow feets region for 12 weeks for 10 women aged 30 or older who are undergoing skin aging for 12 weeks. The depth (μm) of the wrinkles was measured using a skin visiometer (SV-600, C + K, Germany) using image analysis of the eye wrinkle replica at 4 weeks, 8 weeks and 12 weeks. The average of the 10 measurements is shown in Table 9 below.

Wrinkle depth (μm)
Formulation Example 1 Formulation Comparative Example 1 Nourishing cream before application 348 ± 15 324 ± 10 Nutrition cream 12 weeks after application 290 ± 16 318 ± 15 Wrinkle depth reduction rate (%) 16.6 1.9

Next, in order to measure the effect of improving skin barrier damage, the following experiment was conducted. Acetone was used to damage the skin barrier. When the amount of transepidermal water loss (TEWL) reached 4.0 g / m ² / h by dispensing acetone into the aliquots of 8- to 12-week old reared mice, the nutritional creams of Formulation Example 1 and Comparative Example 1 were prepared Respectively.

TEWL was measured with an evaporator Tewameter 210 from C + K (Cologne, Germany). The extent to which the TEWL was reduced through the difference between the TEWL measurement value and the initial TEWL measurement value after 6 hours of application was evaluated to confirm the degree of restoration of the skin barrier function. The skin barrier recovery rate was calculated according to the following formula 3, and the results are shown in Table 10 below.

[Formula 5]

B (Barrier Recovery) = (1- (B _{t = 6} - B _{t = 0} ) / (B _{t = d} - B _{t = 0} )) x 100

B _{t = 6} : TEWL measurement value after 6 hours from skin barrier damage

· B _{t = 0} : TEWL measured before skin barrier damage

B _{t = d} : TEWL measured immediately after skin barrier damage

Skin barrier recovery rate (%)
Formulation Example 1 Formulation Comparative Example 1 Nourishing cream application 6 hours after 47 12

As shown in Tables 9 and 10, the nutritional cream of Formulation 1 containing the extract according to the present invention can significantly reduce the depth of wrinkles compared to the nutritional cream of Formulation Comparative Example 1 which does not belong to the present invention In addition, the damage recovery rate of the skin barrier is excellent and the loss of transdermal water can be minimized. This means that the composition according to the present invention can efficiently inhibit and prevent skin aging.

Hereinafter, other formulation examples of the present invention will be exemplified. However, the formulation of the cosmetic composition according to the present invention should not be construed as being limited thereto, and ordinary variations of the person skilled in the art are possible within the scope of the present invention.

Formulation Example 2. Lotion number ingredient content(%) One Purified water Balance 2 Butylene glycol 5.00 3 Sample 1 2.00 4 glycerin 4.00 5 Disodium EDTA 0.02 6 Carbomer 0.10 7 Betaine 0.50 8 Hyaluronic acid 0.01 9 ethanol 4.00 10 PEG-60 Hydrogenated Castor Oil 0.30 11 Methyl paraben 0.10 12 Phenyl trimecicone 0.05 13 Flavoring agent a very small amount 14 coloring agent a very small amount

Formulation Example 3. Lotion number ingredient content(%) One Purified water Balance 2 Methyl paraben 0.20 3 Sample 1 5.00 4 glycerin 5.00 5 Butylene glycol 7.00 6 Carbomer 0.10 7 Xanthan gum 0.05 8 Cetylaryl alcohol 3.00 9 Glyceryl stearate 0.50 10 Propylparaben 0.10 11 Mineral oil 3.00 12 Vegetable squalane 3.00 13 Cetyl ethyl hexanoate 3.00 14 Arginine 0.10 15 Flavoring agent a very small amount

Formulation Example 4. Cream number ingredient content(%) One Purified water Balance 2 Methyl paraben 0.20 3 Sample 1 5.00 4 glycerin 5.00 5 Butylene glycol 7.00 6 Carbomer 0.20 7 Xanthan gum 0.08 8 Cetearyl alcohol 3.00 9 Glyceryl stearate 0.50 10 Behenyl alcohol 2.00 11 Aceh Litchi 0.20 12 Propylparaben 0.10 13 Mineral oil 3.00 14 Vegetable squalane 3.00 15 Cetyl ethyl hexanoate 3.00 16 Arginine 0.20 17 Flavoring agent a very small amount

Formulation Example 5. Essence number ingredient content(%) One Purified water Balance 2 Methyl paraben 0.20 3 Sample 1 5.00 4 Hyaluronic acid 0.05 5 Butylene glycol 2.00 6 Carbomer 0.08 7 Xanthan gum 0.04 8 Cetearyl alcohol 0.50 9 Glyceryl stearate 0.50 10 Propylparaben 0.10 11 Mineral oil 1.00 12 Vegetable squalane 2.00 13 Cetyl ethyl hexanoate 1.00 14 Arginine 0.08 15 ethanol 4.00 16 PEG-60 Hydrogenated Castor Oil 0.30 17 Flavoring agent a very small amount

Formulation Example 6. Pack number ingredient content(%) One Purified water Balance 2 glycerin 10.00 3 Sample 1 5.00 4 Butylene glycol 5.00 5 kaoline 4.00 6 Caprylic / capric triglyceride 4.00 7 Cyclomethicone 1.00 8 Magnesium aluminum silicate 1.00 9 PEG-100 stearate, glyceryl stearate 2.00 10 Xanthan gum 0.10 11 ethanol 3.00 12 Methyl paraben 0.20 13 Clofenine 0.10 14 Titanium dioxide 0.50 15 Flavoring agent a very small amount

Claims (7)

The present invention relates to an anti-aging cosmetic composition comprising extracts of K. chinensis,
Wherein said antiaging agent exhibits wrinkle prevention or improvement, skin moisturizing power enhancement, skin elasticity improvement, skin barrier improvement, and antioxidation. The method according to claim 1,
The cosmetic composition for anti-aging according to any one of claims 1 to 3, wherein the starch, ginseng, juli, rhubarb and stonite are contained in a weight ratio of 1: 1: 2: 1: 1. The method according to claim 1,
Wherein said extract is extracted by hot water extraction, ultrasonic extraction or high-temperature pressure extraction. The method according to claim 1,
Wherein the extract is extracted using at least one solvent selected from the group consisting of water, a lower alcohol, a polyhydric alcohol and a hydrocarbon-based solvent as an extraction solvent. The method of claim 3,
Wherein the rosin is pre-roasted before extracting the marigold, Chaetocerosus, Quercus, Rhizoma and Rhodochrous meteorite. delete delete

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