KR20160024525A

KR20160024525A - Cosmetics Compositions Comprising Stachys sieboldii miq. roots Extract as Active Ingredients

Info

Publication number: KR20160024525A
Application number: KR1020140111399A
Authority: KR
Inventors: 권순우; 김진화; 오정영; 강정욱; 이근수; 표형배
Original assignee: 한불화장품주식회사
Priority date: 2014-08-26
Filing date: 2014-08-26
Publication date: 2016-03-07

Abstract

The present invention relates to a pharmaceutical composition for dermatitis, which comprises, as an active ingredient, a cosmetic composition comprising a base composition for sleep deprivation as an active ingredient and a base composition for sleep depilation. The cosmetic composition of the present invention is useful as an antioxidant, an antioxidant (inhibiting MMP-1 protein expression), a type 1 collagen biosynthesis promoting effect, a photoaging prevention effect, an acne prevention effect, a skin elasticity improving effect, a whitening effect, The effect of preventing hair loss and hair growth, improving skin cell aging, inhibiting caveolin-1 expression, improving skin elasticity improves skin moisturizing power and whitening function, smoothes skin surface by smoothly removing epidermal keratin have.

Description

(Cosmetics Compositions Comprising Stachys sieboldii miq.) As an active ingredient. roots Extract as Active Ingredients}

The present invention relates to a cosmetic composition comprising an antidepressant extract as an active ingredient. The cosmetic composition according to the present invention is excellent in active oxygen improving effect, skin elasticity improving effect, photoaging prevention effect, whitening effect, acne prevention effect, hair damage prevention effect, hair loss prevention and hair growth effect, To an anti-aging functional cosmetic composition capable of smoothing the skin surface.

Skin is a site that directly receives physical and chemical injuries from various external factors, and is an indispensable organization for us to have the ability to heal their functional and structural damage. However, our skin, for example, is damaged due to various pollutants and stresses. If such a damage does not result in cell proliferation of the skin, the skin becomes more damaged, skin elasticity is lowered , Wrinkles are generated. As the skin ages, the amount of collagen, mucopolysaccharide and hyaluronic acid in the matrix, which plays an important role in skin elasticity, decreases and the amount of insoluble collagen increases. As a result, the dermis becomes difficult to maintain the moisture of the skin, and the skin becomes rough. Furthermore, sebaceous glands and sweat glands decrease in function, gradually reducing the amount of sebum and sweat. As a result, the sebum is not properly produced, the protective film against skin hydration is lost, and the skin is finally dried, the transparency is lost, the elasticity of the skin protein is reduced, and wrinkles are formed.

On the other hand, if the moisturizing function of the skin does not function properly, it may cause skin elasticity degradation. In other words, loss of moisture of the skin is an important cause of skin irritation. The moisture balance of the skin is controlled by the epidermis. The stratum corneum of the epidermis is an accumulation of dead cells, but plays a very important role in maintaining moisture in the skin. The human body is composed mostly of water that plays an important role in the maintenance of life. The moisture content of the normal skin is 70% in the body and 20% in the stratum corneum, which supports the elasticity and flexibility of the skin. When the water content of the stratum corneum is 10% or less, it is known that the skin becomes dry and causes skin troubles. The amount of moisture in the stratum corneum produced by regular keratinization is always 10-20%. Presently, the method of water retention in the skin uses an occlusive agent. The occlusive agent is a hydrophobic substance which promotes floating of water by forming a barrier against moisture loss on the skin. The most commonly used occlusive agents are vaseline, lanolin, cocoa butter, mineral oil and silicone. To make cosmetics, sodium lauryl sulfate, dimethyl sulfoxide, croton oil ) And sodium hydroxide are added from 20 kinds to 50 kinds. When the raw materials are applied to a product, irritation such as inflammation, itching, and allergy may occur on the skin. Exposure of the skin to irritating substances damages the stratum corneum, leading to an increase in transepidermal water loss, an indicator of stratum corneum damage, which may also be a primary inflammatory response.

For this reason, conventional cosmetics have a problem that harmful side effects are generated on the skin, and thus there is a limit to obtaining a skin moisturizing effect and a protective effect.

On the other hand, Stachys sleep (Stachys sieboldii miq.) Is a perennial herbaceous plant belonging to Lamiaceae and grows at a height of 30 ~ 60 Cm. The flower is magenta and runs from the middle of May to the end of August to the upper part of the branches and stems and blooms in the layer. The roots are bulbous parts that look like bulbs and are 1 ~ 3 cm long and rhombic. The roots are red and milky. Other names are known as asterisks, asterisks, asterisks, zizam, tochoncho, supernaturalis, jiyu, earthen, tortoise, wawa. The efficacy is known to be effective for fever, seawater, pneumonia, toxin, hysteria, hemorrhage, swelling, inflammation, analgesia, gust, appendicitis, sore throat, fever diarrhea, fever and swelling. Especially recently, phenylethanoid glycoside It is known that it is effective for myasthenia gravis, cerebral infarction, prevention of senile dementia and improvement of memory.

On the other hand, skin burning or pigmentation is caused by melanin production in the skin by ultraviolet exposure. One way to prevent this is to cut off ultraviolet rays reaching the skin or apply ultraviolet ray blocking agents that absorb in advance. However, it has to be applied every time before exposure to ultraviolet rays, and it is easily washed out with water. Therefore, it is troublesome when it is used frequently, for example, and it is disadvantageous in that stability for long-term use of these agents is not established.

Vitamin C is known to have a variety of physiological functions such as acting as an enzymatic co-factor, reducing the use of vitamin E, and acting as an antioxidant. Vitamin C is known to have a whitening effect when used in large amounts. It reacts with superoxide and hydroxyl radicals when used in high doses.

Superoxide and hydroxyl radicals are active oxygen radicals containing oxygen which can be produced in a normal or diseased state, which acts as a causative agent in the case of sunburn or skin aging.

Vitamin C reacts with these substances to prevent skin burns or skin aging caused by ultraviolet rays.

However, vitamin C has a double bond between carbon number 2 and carbon number 3 and has hydrogen (pK = 4.2) in which acid-ionization reaction can take place in the presence of water. In addition, since it is a strong reducing agent which is easily oxidized by itself, it becomes an unstable anion type compound under high pH conditions. This instability of vitamin C is a problem when vitamin C is formulated as a solution, which is attributed to the stereochemical stress due to polar repulsive force, oxidative degradation by ascorbic acid anion, and decomposition due to water attack.

Conventional methods for stabilizing vitamin C include a method using an extremely low concentration of vitamin C, a method using a non-aqueous solvent, or a method using several derivatives of vitamin C, and the like.

Meanwhile, male-type alopecia are male hormone-dependent and have a direct relationship with the amount of male hormone. Accordingly, many studies for prevention and treatment of alopecia through inhibition of male hormone activity have been reported. On the other hand, when the function of the sebaceous gland is activated by the increase of the secretion of the male hormone, the excess sebum produced in the hair follicle is stagnated in the hair follicle due to the overgrowth of the hair follicle wall. 5-alpha-Reductase is one of the male hormones present in male hormone reactive tissues such as sebaceous glands, hair follicles, prostate, and epididymis, as a result of male hormone-induced hair loss and acne. , An enzyme involved in the metabolism of testosterone into dihydrotestosterone, and requires NADPH for its conversion. In addition, testosterone is involved in male sex adult, skeletal muscle increase, male external genitalia, scrotum growth, spermatogenesis, and dihydrotestosterone is involved in the relevant tissues such as acne, sebum increase, hair loss and enlargement of the prostate. In particular, after puberty, excessive secretion of male hormones causes acne and hair loss. To prevent excessive production of dihydrotestosterone, an active form of male hormone produced by 5-alpha-reductase, 5-alpha-reductase Studies have been actively conducted to develop anti-hair loss and anti-acne agents using an azide inhibitor.

Accordingly, the inventors of the present invention have made extensive studies on the selection of natural stone and natural extracts, which are a kind of snails and snails, and have produced extracts therefrom, and have found that they are effective in eradication of active oxygen, skin elasticity improvement effect, photoaging effect, whitening effect, The present invention has been accomplished on the basis of the fact that it has excellent hair-dressing prevention effect and hair growth effect, has a good moisturizing power, smoothes the surface of the epidermis, smoothes the surface of the skin, and excellently functions as a skin external preparation and cosmetic.

It is an object of the present invention to provide a cosmetic composition comprising an active ingredient (s) as an active ingredient.

In order to attain the above object, the present invention provides a cosmetic composition comprising an effective amount of a foundation stone extract as an active ingredient.

The present invention provides a pharmaceutical composition for dermatological use, which comprises as an active ingredient, a cornerstone sleep extract.

Hereinafter, the present invention will be described in detail.

The present invention provides a cosmetic composition comprising as an active ingredient a foundation stone extract.

In the cosmetic composition of the present invention, the composition may be a skin lotion, a skin softener, a skin toner, an astringent, a lotion, a milk lotion, a moisturizing lotion, a nutrition lotion, a massage cream, The composition is preferably used in cosmetics selected from the group consisting of a pack, a soap, a shampoo, a cleansing foam, a cleansing lotion, a cleansing cream, a body lotion, a body cleanser, an emulsion, a press powder, a loose powder and an eye shadow, It is preferred to include adjuvants which are commonly used in the scientific field and which contain the commonly used contents, wherein the adjuvants are selected from the group consisting of fatty substances, organic solvents, solubilizers, thickeners, gelling agents, softeners, antioxidants, , A stabilizer, a foaming agent, a fragrance, a surfactant, water, an ionic or nonionic emulsifier, a filler , It is more preferable to include a sequestering agent, chelating agent, preserving agent, vitamin, blocker, moisturing agent, essential oil, dye, pigment, hydrophilic or hydrophobic activator, and at least one selected from the group consisting of lipid vesicles.

In the pharmaceutical composition for dermatology of the present invention, it is preferable to further contain glycolic acid or vitamin C in addition to the cornerstone sleep extract.

The glycolic acid as an active ingredient of the present invention is commercially available as a natural product derived from sugarcane. Glycolic acid is safer for the body than retin-A, and acts to regenerate skin and prevent aging by biosynthesizing collagen.

Combination of glycolic acid, an antiaging agent, and vitamin C, an antioxidant, helps to reduce skin irritation and pain of glycolic acid, smooth skin absorption of glycolic acid, Is not only maximized, but also vitamin C is stabilized.

Synergism in the effect of using glycolic acid and vitamin C together in the foundation stone sleep extract not only overcomes the disadvantages of the organic acid having a low pH and increases the absorption and increases the effect but also stabilizes the vitamin C by the glycolic acid And complementary action that supplements the part that is not overcome in one pharmacological action.

The content of the above two active ingredients is 0.01 to 8% by weight, preferably 0.5 to 5% by weight, as a cosmetic agent, and 0.01% by weight or more as an ointment or cream agent, depending on the use form, use purpose, To 12% by weight, preferably 5 to 12% by weight, and the remainder is filled with additives and solvents necessary for each preparation.

The composition of the present invention may contain various additives such as an ointment base, a moisturizing agent, a softening agent, a wetting agent, a lubricant, a preservative, etc., in addition to a cornerstone sleep extract, glycolic acid and vitamin C.

Examples of the additives include wax, stearyl alcohol, polyethylene glycol 4000, glycerin, cetanol, sodium lauryl sulfate, ethyl p-hydroxybenzoate, butyl p-hydroxybenzoate and propylene glycol. These additives are added individually or commercially May be added in the form of a component included in a possible mechanism.

The sperm wax is obtained by purifying the solid component of lead in the head of whale. It can be added to other base to give proper intensity to the ointment and added in an appropriate amount.

Stearyl alcohol acts as a lubricant and may be added in an appropriate amount. Polyethylene glycol 4000 is a representative water-soluble base material, and it is a polymer of ethylene oxide and water. It is soluble in water, good in compatibility with most medicines, and excellent in absorption of medicine.

Glycerin can be used in an appropriate amount as a softening agent and a hygroscopic agent.

Cetanol can be used in an appropriate amount as an emulsifier and a thickener.

Sodium lauryl sulfate may be used in an appropriate amount as a wetting softener to assist in mixing the composition.

Ethyl p-hydroxybenzoate and butyl p-hydroxybenzoate may be included in an effective amount as a preservative to maintain the effectiveness of the preparation. These preservatives show activity in a wide pH range and their effectiveness is enhanced by the combination of two kinds.

Propylene glycol can be used as a surfactant in an amount of 5 to 10% by weight and helps to absorb the skin of glycolic acid and vitamin C.

The above components not only make it easy to apply glycolic acid and vitamin C, which are effective ingredients of the present invention, to skin, but also have an effect on the skin such as control of moisture and lipid on skin surface, protection of skin and purification action.

The anti-oxidant effect of the present invention has excellent anti-aging effect such as active oxygen scavenging effect, MMP inhibitory effect, modulation of MMP expression by ultraviolet irradiation, skin elasticity improvement effect and mitigating skin irritation caused by ultraviolet irradiation .

In addition, primordial sleep extracts showed whitening effects such as inhibition of melanin production, which is a cause of spots, freckles and skin pigmentation.

In addition, the foundation stone extract showed excellent antibacterial activity against acne and 5 - alpha - reductase inhibitory effect which have excellent hair damage and hair loss prevention effect and anti - acne effect.

In addition, the foundation stone sleep extract improves skin moisturizing ability and whitening function by the effect of cell activity, improvement of skin cell senescence, inhibition of caveolin-1 expression, inhibition of CCN-1 expression, improvement of skin elasticity and wrinkle, Smoothes the skin surface smoothly by removing keratin.

Therefore, cosmetic compositions such as lotion, cream, emulsion, pack, powder and the like containing such a foundation stone sleep extract are effective for the active oxygen scavenging effect and the collagenase activity controlling effect, the skin wrinkle improving effect, the photoaging prevention effect, Prevention of damage, prevention of hair loss, hair growth effect, and the like.

The present invention relates to a plant extract which is characterized by extracting a natural plant from natural stone as an extraction solvent with at least one solvent selected from purified water, methanol, ethanol, glycerin, ethyl acetate, butylene glycol, propylene glycol, dichloromethane and hexane And to provide a composition for external application for skin containing as an active ingredient.

Further, the present invention is characterized in that the above extract is obtained by liquid-freezing obtained by cold-pressing and heating filtration at room temperature, furthermore, the solvent is concentrated under reduced pressure or lyophilized.

In addition, the present invention provides a composition for external application for skin, wherein the content of the foundation stone natural product is 0.001-30.0 wt% based on the freeze-dried weight of the whole composition for external application for skin. If the content of the extract is less than 0.001% by weight, the effect of improving the skin is scarcely produced. If the content of the extract is 30.0% by weight or more, the effect of increasing the content of the extract is insignificant.

The present invention also relates to a composition for external application for skin wherein the composition for external use for skin is at least one of a lotion, / O) type formulation as a main active ingredient.

In addition, the present invention is characterized in that the composition for external application for skin is a cosmetic composition or a pharmaceutical composition.

The foundation stone sleep extract can be formulated into a pharmaceutical composition such as a capsule, a liquid preparation, an ointment preparation, a patch or a sustained release formulation using a pharmaceutically acceptable carrier, and a pharmacologically acceptable base, carrier, excipient, binder Such as starch, tragacanth gum, gelatin, molasses, polyvinyl alcohol, polyvinyl ether, polyvinylpyrrolidone, hydroxypropylcellulose, methylcellulose, ethylcellulose and carboxymethylcellulose, Starch, hydrogenated vegetable oil), coloring agents, and the like, which are well known to those skilled in the art. Examples of the carriers and excipients include lactose, glucose, sucrose, mannitol, potato starch, cornstarch, calcium carbonate, calcium phosphate and cellulose.

In addition to the above, additives such as stabilizers, solubilizers, perfume fragrances such as transdermal absorption accelerators, and preservatives may be further added.

The pharmaceutical composition thus prepared can be applied to the skin once to several times a day depending on the symptoms, and the application can be controlled according to the degree of symptom improvement.

Hereinafter, the present invention will be described in more detail by way of examples. It should be noted, however, that these examples are only illustrative examples of the present invention, and the scope of the present invention is not limited to these examples.

Foundation stone sleep Preparation of extract

After shredding, the shade stone foundation was refluxed in purified water for 6 hours with an aqueous 70% (v / v) ethanol solution, cooled, and filtered through a Whatman # 5 filter paper. The filtered extract was concentrated under reduced pressure at 50 캜 or lower and lyophilized to prepare a powdery extract.

Cytotoxicity of the sample

The cytotoxicity of the obtained crude extracts of groundnut silk to skin cells was evaluated.

Fibroblasts diluted in cell culture medium (DMEM supplemented with 10% FBS) were added to 96-well test plates at 1 × 10 ⁵ cells / mL for 24 hours. The sample obtained in Example 1 diluted to an appropriate concentration in each well was treated and cultured for 24 hours. After 24 hours, the medium was removed, and a cell culture medium 200 containing a solution of MTT (3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyl tetrazolium bromide) (2.5 mg / And incubated in a 37 ° C CO ₂ incubator for 2 hours. The medium was removed and 100 μl of DMSO (dimethyl sulfoxide) was added. After shaking for 5 minutes to dissolve the cells, absorbance at 565 nm was measured in a microplate reader. The cell viability (%) was determined by Equation (1) and the concentration of the sample that did not affect cell survival was determined.

Bo: 565 nm absorbance value of a well that underwent chromogenic reaction in cell culture medium

Bt: 565 nm absorbance value of the well that underwent chromogenic reaction without treatment of the sample

St: a 565 nm absorbance value of the well treated with the color-developed sample

As can be seen in Table 1, it was confirmed that the 100% survival concentration of the skin fibroblasts of each sample was less than 2% of the cornerstone sleep extract of Example 1, which is a safe sample showing no cytotoxicity.

Name of sample 100% cell viability concentration Example 1 2%

NBT Experiment on antioxidant effect test

The antioxidative activity of NKT (Nitro Blue Tetrazolium) was measured by using the green tea extract, which is well known as an antioxidant, as a comparative sample in laboratory conditions in order to confirm the antioxidative effect of the extract of the extract of the groundmass extract obtained in Example 1.

In order to measure the antioxidative effect, active oxygen produced by xanthine and xanthine oxidase is measured by the NBT method, and the effect of the test substance on the removal of active oxygen, that is, the effect of scavenging reactive oxygen is evaluated. And active oxygen is generated by xanthine and xanthine oxidase. The active oxygen is reacted with Nitro Blue Tetrazolium (NBT), and the blue color produced thereby is measured at a wavelength of 560 nm to measure the active oxygen scavenging rate.

(0.1 ml), 3 mM EDTA solution (0.1 ml), BSA solution (0.1 ml) and 0.72 mM NBT solution (0.1 ml) were added to the solution, and 0.1 ml And left at 25 캜 for 10 minutes. A solution of xanthine oxidase (0.1 ml) was added, stirred rapidly, and incubated at 25 ° C for 20 minutes. Then, 6 mM CuCl 2 solution (0.1 ml) was added to stop the reaction, and the absorbance St at 560 nm was measured. The blank test is carried out in the same manner as above using distilled water instead of the sample solution, and the absorbance Bt is measured. Also, the blank of the sample solution is measured by the same procedure using distilled water instead of the enzyme, and the absorbance Bo is measured.

The inhibition rate was calculated according to Equation (2), and the results are shown in Table 2.

St: absorbance at 560 nm after enzyme reaction of the sample solution

Bt: Absorbance at 560 nm after enzyme reaction in blank test solution

So: absorbance of 560 nm before reaction in the absence of enzyme in the sample solution

Bo: absorbance at 560 nm before the reaction in the absence of enzyme in the blank test solution

As can be seen in Table 2, the antioxidant activity of cornstal moth extract showed excellent antioxidative activity similar to that of green tea extract having excellent antioxidant activity.

Name of sample content Antioxidative effect (%) Example 1 0.2 wt% 92.35 Example 1 0.1 wt% 86.91 Example 1 0.05 wt% 82.43 Example 1 0.005 wt% 22.31 Green tea extract 0.2 wt% 96.58

Free radical Measurement of scavenging activity

To determine the free radical scavenging activity of the cornerstone sleep extract obtained in Example 1, free radical scavenging activity was measured using a DPPH method using an extract having an excellent antioxidative activity such as green tea extract under laboratory conditions as a comparative sample.

The DPPH method measures free radical scavenging activity by reducing power using a free radical called DPPH (2,2-Di (4-tert-octylphenyl) -1-picrylhydrazyl free radical). The degree of reduction of the absorbance by reduction of DPPH by the test substance is compared with the absorbance of the blank test solution and the free radical scavenging ratio is measured at a wavelength of 560 nm.

For determination of DPPH free radical scavenging activity, base layer sleep extracts of 0.2%, 0.1%, 0.05% and 0.005% concentration were prepared. The extracts of the above concentrations were placed in a 96-well plate, and DPPH prepared from 100 uM methanol solution was added thereto to make the total volume of the solution 200 μl. After incubation at 37 ° C for 30 minutes, absorbance was measured at 560 nm. The free radical scavenging activity (%) was calculated by the following equation (3).

A: absorbance of the control well without treatment of the cornerstone sleep extract of the present invention

B: Absorbance of the experimental group well treated with the base laying extract of the present invention

As can be seen in Table 3, it was found that the cornerstone sleep extract of Example 1 exhibits excellent free radical scavenging activity and exhibits an excellent antioxidative effect as that of green tea extract having excellent antioxidant activity.

Name of sample content Antioxidative effect (%) Example 1 0.2 wt% 92.59 Example 1 0.1 wt% 91.76 Example 1 0.05 wt% 90.33 Example 1 0.005 wt% 30.79 Green tea extract 0.2 wt% 92.84

After UV irradiation Foundation stone sleep By extract MMP -1 expression inhibition evaluation

In this Experimental Example, enzyme immunoassay (ELISA) was carried out to measure the concentration of MMP-1 after UV irradiation and sample addition of the cornerstone sleep extract obtained in Example 1.

UVA was irradiated to human dermal fibroblasts at an energy of 6.3 J / cm < 2 > using a UV chamber. Ultraviolet irradiation dose and incubation time were established by preliminary experiment to maximize MMP expression level in fibroblasts. Negative controls were wrapped in silver foil to maintain the same time in the UVA environment. UVA emission was measured using a UV radiometer. During UVA irradiation, the cells were kept in the previously dispensed medium, irradiated with UVA, exchanged with the medium containing the sample, cultured for 24 hours, and the medium was recovered and coated on 96 wells. The primary antibody (MMP-1 (Ab-5) monoclonal antibody) was treated and reacted at 37 ° C for 60 minutes. The secondary antibody, alkaline phosphatase-conjugated anti-mouse IgG (whole mouse, alkaline phosphatase conjugated) was reacted for about 60 minutes and then incubated with an alkaline phosphatase substrate solution (containing 1 mg / ml ρ-nitrophenylphosphate in diethanolamine buffer solution ) Was reacted at room temperature for 30 minutes and the absorbance was measured at 405 nm with a microplate reader. As a control group, a sample to which no sample was added was used.

The MMP-1 expression induced by ultraviolet irradiation showed an inhibition rate of more than 48% at 0.02 wt%, compared to the control without the sample, which was significantly superior to the inhibitory rate of retinol used as the control group .

Test group MMP-1 expression inhibition rate (%) Control group - Example 1 0.02 wt% 48 Example 1 0.01 wt% 35 Example 1 0.005 wt% 22 Retinol 41

Promoting effect of type 1 procollagen biosynthesis

Procollagen assay was performed by the following method in order to examine the effect of accelerating the biosynthesis of type 1 procollagen, which is a skin substrate component, after each of the extracts of the extracts of crude extract obtained from Example 1 were added to the skin cell culture broth Respectively.

Human dermal fibroblasts isolated from the epidermal tissue of newborns were purchased from Modern Tissue Technology (MTT, Korea) and supplemented with 10% fetal bovine serum (FBS) in DMEM / F-12 (3: And cultured at a concentration of 1 × 10 ⁴ cells / mL. When 70 ~ 80% of the cells were grown, the cells were subcultured at a ratio of 1: 3, and the cells cultured in the third to fourth passages were used for the experiment. For the measurement of the amount of procollagen, the fibroblasts were cultured in a 48-well plate at a concentration of 90% or more, the respective samples were added at an appropriate concentration, and after 24 hours, the amount of procollagen liberated in the medium was measured with procollagen Type 1 C-peptide EIA kit (MK101, Takara, Japan).

As shown in Table 5, the cornerstone sleep extract of Example 1 exhibited about 30% promoting effect of procollagen biosynthesis at a concentration of 0.001% by weight, which is lower than that of TGF-beta, a cell signaling substance used as a positive control, Activity.

Name of sample content MMP-1 expression inhibition rate Example 1 0.001 wt% 29.2% TGF-beta 0.001 wt% 33.7%

By ultraviolet irradiation Skin cell Optical damage Defense effectiveness verification experiment

This Example was carried out in order to evaluate the mitigation effect of cytotoxicity by ultraviolet irradiation of the cornerstone sleep extract obtained in Example 1. Fibroblasts were inoculated into 96-well test plates at 1 × 10 ⁵ cells / mL for 24 hours. After 24 hours, samples of Tungbok sleeping extract were treated and incubated for 24 hours under the same culture conditions. Then the medium was removed and each well was washed once with PBS and 100 쨉 l PBS was added to each well. The cells were irradiated with 10 mJ / cm 2 of ultraviolet light using ultraviolet B (UVB) lamp (Model: F15T8, UVB 15W, Sankyo Dennki, Japan) Was added. The extracts were then treated for 24 hrs. After 24 hours, the medium was removed, and 200 μl of MTT solution (2.5 mg / ml) containing cell culture medium per well was added and cultured in a 37 ° C CO ₂ incubator for 2 hours. The medium was removed and 100 μl of DMSO (dimethyl sulfoxide) was added. Cells were lysed by shaking for 2 minutes and absorbance at 565 nm was measured in a microplate reader. The cell viability (%) was measured by the equation (4) and the cytotoxic relaxation rate by ultraviolet was calculated by the formula (5).

Bo: Absorbance at 565 nm of wells that had undergone color development of the cell culture medium

Bt: Absorbance at 565 nm of a well that underwent chromogenic reaction in a well not treated with a sample

St: absorbance at 565 nm of a well subjected to color development reaction in a well treated with a sample

Bo: Cell survival rate of wells not irradiated with ultraviolet light and not treated with sample

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

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

As a result of the experiment, it can be shown that the cornerstone sleep extract of Example 1 mitigates cytotoxicity by ultraviolet rays by 19.3% at a concentration of 0.125%, thereby preventing cell damage by ultraviolet rays (Table 6).

Name of sample Treatment concentration (%) Cytotoxic Relaxation Rate (%) Example 1 0.125 19.3

Tyrosinase Experiment of inhibition effect measurement

The present example is to determine the whitening effect by checking the degree of suppression of the enzyme function of tyrosinase to confirm the whitening effect of the sample obtained in Example 1. [

Tyrosinase is an enzyme that helps the production of melanin by stimulating the oxidation process of tyrosine in vivo. This example is a method (Pomerantz SH: J. Biochem., 24: 161-168, 1996) in which the degree of inhibition of the function of the enzyme to inhibit the formation of a black polymer called melanin by oxidation of tyrosine is measured To determine the whitening effect.

The inhibitory activity of each sample against tyrosinase was determined by adding 15 μl of a sample to a 96-well plate, adding 150 μl of 50 mM phosphate buffer (pH 6.5) and 25 μl of 1.5 mM L-tyrosine solution, followed by addition of mushroom tyrosinase (1,500 units / After incubation at 37 ° C for 20 minutes, the inhibition rate against tyrosinase was measured by measuring the absorbance at 490 nm using a microplate reader (ELx800, USA). The inhibition rate (%) for tyrosinase was calculated according to Equation (6), and the IC ₅₀ value is the concentration of substance that inhibits tyrosinase enzyme activity by 50%.

A: absorbance before reaction of the well containing the sample

B: absorbance after reaction of the well containing the sample

C: Absorbance before reaction of well without sample

D: absorbance after reaction of well without sample

As shown in Table 7, the IC ₅₀ value of the sample of Example 1 was 0.38% as shown in Table 7. As shown in Table 7, the tyrosinase inhibitory effect was similar to that of arbutin, a known whitening agent.

Name of sample Tyrosinase inhibitory effect (IC ₅₀ ) Example 1 0.38% Arbutin 0.45%

B16F1 Melanocyte Melanin synthesis inhibition experiment

The present example is to determine the whitening effect of B16F1 melanocyte in view of the degree of inhibition of melanin formation in order to confirm the whitening effect of the foundation stone sleep extract obtained in Example 1.

The B16F1 melanocyte used in this example is a cell strain derived from a mouse, and is a cell that secretes a melanin pigment called melanin. During the artificial culture of these cells, samples were treated to compare the degree of reduction of melanin pigment. The B16F1 melanocyte used in this example was purchased from ATCC (American Type Culture Collection, Accession No. 6323). Melanin synthesis inhibitory effect of B16F1 melanocyte was measured as follows. B16F1 melanocytes were dispensed into 6-well plates at a concentration of 2 x 10 ⁶ cells / mL per well, and the cells were treated at a concentration that did not induce toxicity after 72 hours of incubation. After incubation for 72 hours, the cells were detached with trypsin-EDTA, and the number of cells was measured, followed by centrifugation to recover the cells. Quantification of intracellular melanin was carried out with a slight modification of the method of Lotan: Cancer Res., 40, 3345-3350 (1980). The cell pellet was washed once with PBS (phosphate buffered saline), and 1 ml of homogenization buffer (50 mM sodium phosphate, pH 6.8, 1% Triton X-100, 2 mM PMSF (phenylmethanesulphonylfluoride)) was added and vortexed for 5 minutes to disrupt the cells Respectively. After centrifugation (3,000 rpm, 10 min), 1N NaOH (containing 10% DMSO (dimethyl sulfoxide)) was added to the cell filtrate to dissolve the extracted melanin, and the absorbance of melanin was measured at 405 nm using a microplate reader. Was measured to determine the inhibition rate (%) of melanin formation in the sample. Melanin formation inhibition rate (%) of B16F1 melanocyte was calculated according to Equation (7).

A: Amount of melanin in wells to which no sample was added

B: Amount of melanin in the well to which the sample was added

The inhibitory effect of B16F1 melanocyte on melanin synthesis was tested. The IC ₅₀ of the extracts of cornstarch was 0.19%, which was similar to that of arbutin, a conventional whitening agent (Table 8).

Name of sample Treatment concentration (%) Melanin synthesis inhibitory effect (IC ₅₀ ) Example 1 0.1 0.19% Arbutin 0.1 0.22%

Antimicrobial activity against acne bacteria

In Example 10, a Paper Disc Test was conducted to confirm the antibacterial activity against the acne bacterium of the foundation stone sleep extract obtained in Example 1. First, the cells were pre-cultured for 48 hours in a BHI liquid medium (Brain-Heart Infusion Broth; 3.7%) to activate the epidermis Propionibacterium acnes, the cause of acne. The prepared culture broth was applied to 0.1 ml of BHI solid medium (Brain-Heart Infusion Broth; 3.7%; agar 1.5%) and dried. Each of the samples obtained in Example 1 was diluted to 12% (w / v) in a 95% aqueous ethanol solution, and 50 μl of the diluted sample was dispensed on a paper disk having a diameter of 8 mm, and then placed on the solid medium prepared above. For 3 days. The antimicrobial activity was evaluated by observing the growth inhibition zone around the paper disk and measuring the size of the inhibition zone.

As a result of measuring the antimicrobial activity against acne bacterium, it was found that the size of the bacterium growth inhibition ring of the antimicrobial extract was 12 mm.

Foundation stone sleep The 5 alpha - Reductase Activity inhibition evaluation experiment

5 alpha - reductase activity The 5 - alpha - reductase used in the inhibition assay was produced by the enzyme produced by the forebrain - derived fibroblasts.

Fibroblasts were inoculated into microplate holes every 10,000 cells per well and cultured. Radiolabelled testosterone was added to each hole at 0.1 mμCi with 3H (tritium), followed by culturing to determine whether fibroblasts were used. As a control group, we did not enter the foundation stone sleep extract. After 24 hours of incubation, the supernatant is obtained and steroids are obtained with 1 ml of 1: 1 ethyl acetate-cyclohexane extraction solvent. The obtained steroid was placed on a thin layer chromatography plate and developed with a chloroform / methanol mixture 98/2 (v / v). The radioactivity of the points corresponding to testosterone and dihydrotestosterone was measured using a densitometer to calculate the conversion rate, and the result was compared with the control group (the conversion rate when the extract was not added) Alpha-reductase inhibitors were evaluated.

A = conversion of testosterone to dihydrotestosterone (when no extract is added)

B = conversion of testosterone to dihydrotestosterone (upon addition of extract)

The results of the experiment show that the extracts of Chaemannia ash have an inhibitory effect on 5 alpha-reductase (Table 9).

Example 1 Weight (%) One 0.1 0.05 0.025 0.01 0.005 0.0025 Inhibition rate (%) 93.4 86.8 79.4 76.4 58.8 47.2 1.9

Foundation stone sleep Extract Caveolin -1 expression inhibitory effect

For the measurement of caveolin-1 expression in cells, FITC-labeled caveolin-1 antibody was used and FITC-conjugated caveolin-1 antibody (Santa Cruz, USA) was used. The cultured cells were harvested according to the experimental conditions and rinsed twice with cold PBS containing 0.5% BSA. Cells were collected by adding 50 ㎕ of FITC-conjugated caveolin-1 antibody diluted 1: 100 and incubated at 4 ℃ for 30 min. Rinse twice with 500 μl to remove residual antibody. Then, 500 μl of PBS supplemented with 0.5% BSA was added, and FACS analysis was performed. The instrument was FACSCalibur (BD, USA) and WinMDI was used for analysis (Table 10). As a result of this experiment, it was confirmed that the amount of caveolin-1 expressed in the cells after the treatment of the base coat extract was reduced compared with that of the control group.

FACS analysis of caveolin-1 expression level after treatment Sample (extract) Concentration (%) Caveolin-1 Expression Inhibitory Effect (%) Example 1

0.05 38.35 0.025 21.65 0.0125 14.83 0.00625 8.34 0.00313 6.54

Skin elasticity improvement test

A cosmetic composition containing the foundation stone extract obtained in Example 1 was prepared and evaluated for its effect of improving skin elasticity in humans compared with Comparative Example 1.

The cosmetics used in the comparative experiment are in the form of a cream, and the composition thereof is shown in Table 11. First, the b) image recorded in Table 11 was heated and stored at 70 ° C. To this was added a phase, which was pre-emulsified, uniformly emulsified with a homomixer, and then slowly cooled to prepare a cream (Example 13, Comparative Example 1). The cream prepared in Example 13 was applied to the right side of the face and the cream prepared in Comparative Example 1 was applied to the left side of the face for 2 consecutive months for 15 consecutive patients (20 to 35 year-old female) . The skin elasticity was measured by using a skin elasticity meter (SEM 575, C + K Electronic Co., Germany) before and after using the product for 2 months. The experimental results are shown in Table 12 as ΔR7 values of Cutometer SEM 575, where R7 values indicate the viscoelasticity properties of the skin.

As shown in Table 12, it can be seen that the cream containing the cornerstone extract of Example 1 has excellent skin elasticity improving effect.

division Example 13 Comparative Example 1 end Stearyl alcohol 8 8 Stearic acid 2 2 Stearic acid cholesterol 2 2 Squalane 4 4 2-octyldodecyl alcohol 6 6 Polyoxyethylene (25 mole addition) alcohol ester 3 3 Glyceryl monostearate Aster 2 2 I Foundation stone sleep extract One - Propylene glycol 5 5 Purified water Balance Balance

Note) Unit: wt%

Experimental product Skin elasticity effect (△ R7) Example 13 0.38 Comparative Example 1 0.17

n = 20, p < 0.05

Whitening efficacy Clinical evaluation

The present example was evaluated by comparing the skin whitening effect with that of the Comparative Example 1 to a human using a cream cosmetic formulation containing the foundation stone sleep extract obtained in Example 13. 20 women (20 to 35 years old) were applied the cream prepared in Example 13 on the right side of the face and the cream prepared in Comparative Example 1 on the left side of the face for 2 consecutive months for 2 consecutive days . After the completion of the test, the color of the applied area of the left and right sides of the face was analyzed with an image analyzer and color change (L) of the color was measured using a Minolta CR300. Objective visual observation by a plurality of experts Subjective visual observation was performed to measure the effect according to the following grades. The results are shown in Table 13 below. At this time, the degree of whitening efficacy was classified into the following seven grades and evaluated.

Whitening efficacy Visual evaluation criteria:

-3: Very bad

-2: worse

-1: slightly worse

0: No change

1: slight improvement

2: Improvement

3: Very much improved

As shown in Table 13, it can be seen that the whitening effect is excellent in the facial skin of the experimenter who applied the cream containing the foundation stone sleep extract.

Change in skin color brightness (L) Objective evaluation of expert Subjective evaluation of the subject Example 13 Comparative Example 1 Example 13 Comparative Example 1 Example 13 Comparative Example 1 medium 4.93 3.27 2.5 1.5 2.8 1.3

Other examples are shown below. In other words, lotion, milky lotion and essence liquid containing the cornerstone sleep extract obtained in Example 1 were prepared in Examples 15 to 17. These lotion, essence and cosmetic liquids containing the foundation stone extract of the present invention can be used as an active oxygen scavenging effect, a skin wrinkle improving effect, a skin elasticity improving effect, a photoaging prevention effect, a whitening effect, an acne prevention effect, a hair damaging prevention effect, It showed excellent effect on skin improvement.

Foundation stone sleep Manufacture of lotion containing extract

0.05 g of polypyrrolidone, 0.1 g of oleyl alcohol, 0.2 g of polyoxyethylene monooleate, 0.2 g of fragrance, 0.1 g of p-hydroxybenzoic acid methyl ester, a small amount of antioxidant and a small amount of pigment are mixed and dissolved in 8 g of 95% ethanol . 0.05 g of the cornerstone sleep extract obtained in Example 1 and 5 g of glycerin were dissolved in 85.33 g of purified water. The mixture was added to the mixture and stirred to obtain a skin lotion having skin-improving effect.

Foundation stone sleep Manufacture of emulsion containing extract

0.2 g of cetyl alcohol, 10 g of squalane, 2 g of vaseline, 0.2 g of p-hydroxybenzoic acid ethyl ester, 1 g of glycerin monoestearylate, 1 g of polyoxyethylene (20 mols) monooleate and 0.1 g of perfume were mixed and dissolved at 70 캜 0.5 g of the foundation stone sleep extract obtained in Example 1, 5 g of dipropylene glycol, 2 g of polyethylene glycol-1500, 0.2 g of triethanolamine and 76.2 g of purified water were heated to 75 캜 and dissolved. The two were mixed and emulsified and then cooled to obtain an oil-in-water (O / W) type milky lotion.

Foundation stone sleep Extract-containing Serum Produce

To 5 g of 95% ethyl alcohol, 1.2 g of polyoxyethylene sorbitan monooleate, 0.3 g of chitoolose, 0.2 g of sodium hyaluronate, 0.2 g of vitamin E-acetate, 0.2 g of sodium permanganate, 0.1 g of p-hydroxybenzoic acid ethyl ester 1 g of the foundation stone sleep extract obtained in Example 1 and an appropriate amount of pigment were mixed to obtain a serum having skin-improving effect.

Foundation stone sleep In the extract Glycolic acid And vitamin C

45 g of glycolic acid (glycolic acid 99%, manufactured by Aldrich) and 30 g of vitamin C (L-ascorbic acid, Sigma) were well mixed into 450 g of the foundation stone sleep extract.

Pigmentation, removal of fine lines, acne scars and oxidative testing of vitamin C

The composition of Example 18 was applied to the pigmentation site for four months in the morning and evening twice a day, with three groups of 50 healthy male and female volunteers with severe pigmentation on the eyes and cheeks. The extent to which pigmentation was removed weekly and the extent to which the skin was stimulated was observed and recorded.

The composition of Example 18 was applied to the affected area twice a day with three groups of 50 male and female patients with brow skin acne on the forehead, Were visually observed and shown in the following table.

The degree of oxidation of vitamin C in air by oxygen was visually observed at room temperature. The criteria for the oxidation of vitamin C were evaluated in terms of degree of discoloration of white to yellow, and are shown in Table 14 below.

Test substance Deposited pigment removal (persons) Treatment of acne and reduction of pores (persons) Degree of vitamin C oxidation splendor available invalidity splendor available invalidity Yellowing time Example 18 35 12 3 43 7 0 More than 90 days

As a result of application of the composition of Example 18 of the present invention to a patient, reduction of the pores of the face, treatment of acne, and whitening effect on sun-exposed skin were shown.

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the invention is not limited to the disclosed exemplary embodiments, but, on the contrary, This is possible.

Claims

Claims [1] A cosmetic composition comprising as an active ingredient an extract of Tranquil Primrose.

The cosmetic composition according to claim 1, wherein the cosmetic composition exhibits an effect of improving active oxygen.

The cosmetic composition according to claim 1, wherein the cosmetic composition exhibits an inhibitory effect on MMP-1 protein expression.

The cosmetic composition according to claim 1, wherein the cosmetic composition exhibits a type 1 procollagen biosynthesis promoting effect.

The cosmetic composition according to claim 1, wherein the cosmetic composition exhibits a photoaging effect.

The cosmetic composition according to claim 1, wherein the cosmetic composition exhibits an anti-acne effect.

The cosmetic composition according to claim 1, wherein the cosmetic composition exhibits an effect of improving skin elasticity.

The cosmetic composition according to claim 1, wherein the cosmetic composition exhibits a whitening effect.

The cosmetic composition according to claim 1, wherein the cosmetic composition exhibits an effect of preventing hair damage.

The cosmetic composition according to claim 1, wherein the cosmetic composition exhibits hair loss prevention and hair growth effect.

The cosmetic composition according to claim 1, wherein the cosmetic composition exhibits an effect of improving skin cell senescence.

The cosmetic composition according to claim 1, wherein the cosmetic composition exhibits an effect of inhibiting caveolin-1 expression.

The composition according to claim 1, wherein the composition is selected from the group consisting of a skin lotion, a skin softener, a skin toner, an astringent, a lotion, a milk lotion, a moisturizing lotion, a nutrition lotion, a massage cream, a nutritional cream, Wherein the cosmetic composition is used in cosmetics selected from the group consisting of soap, shampoo, cleansing foam, cleansing lotion, cleansing cream, body lotion, body cleanser, emulsion, press powder, loose powder and eye shadow.

2. The cosmetic composition according to claim 1, wherein the composition further comprises adjuvants which are commonly used in cosmetics or dermatological science and contain a commonly used content.

15. The method of claim 14, wherein the adjuvant is selected from the group consisting of a fatty substance, an organic solvent, a solubilizer, a thickening agent, a gelling agent, a softening agent, an antioxidant, a suspending agent, a stabilizer, a foaming agent, a perfume, a surfactant, At least one selected from the group consisting of nonionic emulsifiers, fillers, sequestering agents, chelating agents, preservatives, vitamins, blocking agents, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic active agents and lipid vesicles &Lt; / RTI >

A pharmaceutical composition for dermatological use, which comprises a foundation stone sleep extract as an active ingredient.

The pharmaceutical composition for dermatological use according to claim 16, which further comprises glycolic acid or vitamin C in addition to the foundation stone sleep extract.