KR101142008B1 - Cosmetic composition comprising Magnolia sieboldii stem cell cultured products as active ingredient - Google Patents

Abstract

The present invention provides a cosmetic composition containing the Magnolia egg stem cell culture product as an active ingredient. Chunma Magnolia stem cell culture product of the present invention has excellent cell activity enhancing effect and collagen synthesis and MMP-1 production inhibitory effect, cosmetic composition containing the Magnolia Magnolia stem cell culture product is anti-aging, wrinkle improvement, skin whitening, skin moisturizing And it has an excellent effect on improving atopy through skin moisturizing.

Magnolia egg stem cells, culture products, aging, wrinkles, whitening, moisturizing, atopy

Description

Cosmetic composition containing Magnolia siebol stem cultured product as an active ingredient {Cosmetic composition comprising Magnolia sieboldii stem cell cultured products as active ingredient}

The present invention relates to a cosmetic composition containing the stem cell culture product as an active ingredient.

The primary function of the skin is to play an important role in protecting the human body from external stimuli such as temperature, humidity and ultraviolet rays, and performing various roles such as thermoregulation, respiration and excretion. But as you age, these functions gradually weaken and your skin ages. In particular, as the lipid composition and content of the lipid layer, which functions as a skin barrier, changes, the function decreases rapidly, and the skin moisture content decreases, causing the skin to dry, causing blemishes, freckles, pigmentation, and various skin lesions.

In addition, when the skin is exposed to strong ultraviolet rays, stress, and malnutrition, the skin accelerates aging of the skin such as wrinkle formation, loss of elasticity and keratinization.

The skin of the human body is divided into epidermis and dermis, and newly formed keratinocytes in the basal layer at the bottom of the epidermis are gradually changed from the lower layer to the upper layer, turning into keratin after about 14 days. The falling-out keratinization process occurs. The dermis, which is connected to the base layer, serves to hydrate the epidermis and maintain the elasticity of the skin. In general, skin aging such as reduced skin elasticity, skin wrinkles, etc. are caused by destruction of collagen, decrease in collagen synthesis, loss of elastin, etc., but also occur in the skin epidermis, but rather the phenomenon in the dermis.

Recently, many skin physiological studies have been conducted on the causes of wrinkles and improvement methods of skin. Actually, sunscreens, antioxidants, cell activation promoters, etc. have been used to prevent skin wrinkles in cosmetics, and also skins due to collagen synthesis Wrinkle improvement methods have been proposed.

On the other hand, based on the development of biochemistry and molecular biology, a small amount of signal substances such as various growth factors present in our body have been found, and the aging theory of the living body is being re-established based on this. It is also found that this signal decreases in vivo with age, and it is revealed that this growth factor is closely related to our aging. Most of these growth factors are produced in large quantities in stem cells that are present at the base of each tissue.

On the basis of its strong differentiation and regenerative capacity, stem cells offer great potential for regenerative medicine and have been of great interest in recent years.

Stem cells are undifferentiated cells that have the capacity to self-renewal and to differentiate into various types of tissues or organs. These stem cells are largely divided into animal-derived embryonic stem cells and adult stem cells and plant-derived plant stem cells, which are applied to cosmetics because of ethical problems and negative aspects that threaten human dignity. City, there is a problem that must be solved in consideration of the negative perception of consumers.

On the other hand, plant-derived stem cells, like human stem cells, have not only strong differentiation and regenerative capacity, but also have no ethical or safety issues such as human stem cells. There is a growing interest in utilization.

For example, caninetin, one of the plant growth factors, is present in a small amount on the leaf stem of green plants, which prevents fibroblast function from closely restoring skin elasticity, delays aging of skin cells, and improves fine lines and pigmentation. It is known to relieve skin roughness. Auxin is a plant growth hormone that is made from the growth of stems and roots. It gives skin elasticity and flexibility, and has a powerful regenerative effect and is used as a major ingredient in cosmetics. In addition, 3,9-dihydroxy-6H-benzofuro- [3,2-c] [1] benzopyran-6-one: 3,9-DHBFBP found in legumes such as soybeans and black beans has an antioxidant effect. It has been reported to have an inhibitory effect on skin aging and wrinkle formation (Beggs et al., 1985). Such plant growth factors may be present in large amounts in plant stem cells, which make up the plant's growth point, and in addition to these growth factors, various effective factors for each plant are known to be present in a large amount.

In plants, stem cells exist at shoot apical meristem (SAM) and root apical meristem (RAM). These plant stem cells are differentiated to their own personality and function to renew the tissues and organs of plants such as roots and stems. These plant stem cells are classified as pluripotent stem cells, which have the ability to form various organs through typical functions. Stem cells present in SAM and RAM achieve plant growth through differentiation. Maintaining stem cell differentiation and stem cell density is OC (organizing center) adjacent to SAM and QC (quiescent) adjacent to RAM center) Density and differentiation are regulated by communication with cells (Trends in plant science, Vol. 11 No. 5 241-246, 2006).

Unlike to animals, plants have totipotency, which was discovered by Australian botanist Gottlieb Haberlandt in 1902. Plant pluripotency refers to the ability of plant cells to differentiate into other types of cells, tissues or whole plants through dedifferentiation and redifferentiation processes. In each process, it is possible to regulate the starch flow by controlling plant hormones and growth factors. More specifically, plants can be de-differentiated to obtain callus, which is an undifferentiated cell. Callus cells can be obtained by surface disinfection, cutting and wounding of the plant, and incubation in a solid medium containing the appropriate concentration of plant hormone. Callus cells can be continuously cultured in this state, or cultured in a liquid medium to achieve suspension culture, through which a large amount of cells can be obtained. Callus cells can generate embryos through the regulation of plant hormone concentration in the medium, and form roots or shoots again. Embryos, shoots, and roots thus developed develop into complete plants through germination or growth. In other words, callus cells have long been known to have differentiation and regenerative ability. Thus, callus cells, which are undifferentiated cells, have the capacity of stem cells and thus can be called stem cells. In addition to the stem cells at the point of growth of the plant, callus cells with differentiation capacity can be defined as stem cells, which requires a new definition of plant stem cells, which means stem cells as an expanded concept for stem cells. . The expanded concept stem cells are called totipotent embryogenic stem cells and are distinguished from pluripotent stem cells, which are stem cells of traditional plants (Trends in plant science, Vol. 12 No. 6 245-252, 20070). Therefore, cultured callus cells or suspension cells can be expected to be various applications as plant stem cells. Callus cultured cells, which are undifferentiated cells as stem cells of this expanded concept, are recognized as stem cells as totipotent embryogenic stem cells, but if plant cells are ultimately recognized through the proof that they are stem cells, the utilization of plant stem cells may be further activated. This is because overcoming the problems of pluripotent stem cells, which are not present in the plant in large quantities and difficult to separate, can be fundamentally solved to supply problems by utilizing totipotent embryogenic stem cells capable of cultivating sufficient amounts as culture stem cells.

Therefore, the researchers are trying to find a substance that prevents skin aging and improves skin wrinkles. As a result, the culture products of Magnolia ova stem cells can be effectively used as an active ingredient in cosmetic compositions for preventing skin aging and improving skin wrinkles. It was found that the invention was completed. Cosmetic composition containing stem cells culture product such as Magnolia is excellent in various skin improvement effects such as skin whitening, skin moisturizing and atopy improvement, as well as skin anti-aging and skin wrinkle improvement.

The above object is achieved by a cosmetic composition containing the Magnolia roe stem cell culture product as an active ingredient.

Preferably, the composition is characterized in that for preventing skin aging.

Also preferably, the composition is characterized in that for improving the skin wrinkles.

Also preferably, the composition is for skin whitening.

Also preferably, the composition is characterized in that for moisturizing the skin.

Also preferably, the composition is characterized in that for improving atopy.

Also preferably, the Magnolia egg stem cell culture product is an extract of a culture solution obtained by culturing stem cells of Magnolia Magnolia or an extract obtained by crushing the cultured stem cells.

Also preferably, the content of the Magnolia stem cell culture product is 0.00001 to 30% by weight relative to the total weight of the cosmetic composition.

Also preferably, the cosmetic composition is composed of solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, cleansing, oil, powder foundation, emulsion foundation, wax foundation, pack, massage cream and spray Has a formulation selected from the group.

Still another object of the present invention is to apply a cosmetic composition of the present invention to the skin of a person, to achieve a skin selected from the group consisting of anti-aging, wrinkle improvement, skin whitening, skin moisturizing and atopic improvement effect Is achieved by

Other objects and advantages of the present invention will become apparent from the following examples and claims.

The present invention relates to a cosmetic composition comprising stem cells culture product. The cosmetic of the present invention is not only very effective as a delivery formulation of stem cell culture products of the Magnolia nymph stem when applied to the skin, thereby preventing skin aging (Experiments 1, 2, 3), improving wrinkles (Experiment 7), The skin whitening (Experimental Examples 4, 5, 8), skin moisturizing (Experimental Example 6) and skin moisturizing (Experimental Example 6) is excellent in the atopic improvement effect.

The present invention relates to a cosmetic composition for preventing skin aging, improving skin wrinkles, skin whitening, skin moisturizing and atopy-containing stem cell culture product.

Cinnamon Magnolia, the active ingredient of the cosmetic composition of the present invention, is a deciduous broad-leaved arboretum (Magnolia sieboldii). Here, the shrub is about 7 meters high, the leaves are alternate, glossy with a long oval, large white flowers grow downwards in May-June, and the fruits ripen in September.

In the present invention, the 'Cheonna Magnolia stem cell culture product' includes an extract of a culture solution obtained by culturing stem cells of the Magnolia Magnolia or all extracts obtained by crushing the cultured stem cells and preferably extracting Callus cultured cells that have demonstrated differentiation ability of the magnolia are the culture products of totipotent embryogenic stem cells.

The Magnolia stem cell culture product is a stem cell culture product obtained from various sites of the Magnolia Magnolia, preferably a stem cell culture product obtained from a site selected from the group consisting of stems, leaves, roots, foliar groups and flower buds.

These Magnolia egg stem cells, germinate under the certain conditions, sterilize stems, leaves, roots, foliar group and flower buds with excellent growth, cut each wound and cut them on the MS solid medium and callus stem Induce cells. The concentrations of phytohormone auxin and cytokine in MS solid medium were properly applied to each plant site. The auxin was 2,4-D (0.10-0.50mg / l), IAA (0.50-1.00gm / l), NAA ( 0.5-1.00mg / l) cytokine maximizes callus induction by adding kinetin (0.10-0.50mg / l) and BAP (0.50-1.00mg / l) in various concentrations. In order to secure a higher content of callus stem cells, the Totipotent embryogenic stem cells thus obtained, a large amount of stem cells can be obtained through suspension culture using MS liquid medium in a gyrotory shaker, and mass production Bioreactor can also be used. Thus obtained Magnolia squeezes the stem cells at 4-8 ℃ low temperature (squeeze) to obtain a stem cell culture product.

The Magnolia stem cell culture product contains various growth factors and cytokines to promote fibroblast migration, activation effect, skin regeneration and anti-aging effect, increase collagen synthesis, inhibitory activity of tyrosinase, melanin synthesis Inhibition, moisturizing effect and atopy improvement effect can be expected.

The components included in the cosmetic composition are active ingredients, in addition to stem cell culture products, various components that maintain the activity of biological components such as preservatives, osmotic agents, pH adjusting agents, buffers, stabilizers, aluminum hydroxide, aluminum phosphate, surfactants, liposomes , Thickeners and the like, and ingredients commonly used in cosmetic compositions such as antioxidants, stabilizers, solubilizers, conventional auxiliaries such as vitamins, pigments and flavors, and carriers, and the like.

According to a preferred embodiment of the present invention, the content of the Magnolia bran stem cell culture product of the present invention is 0.00001-30% by weight, preferably 0.0001-20% by weight relative to the total weight of the cosmetic composition. In this case, when the total weight of stem cell culture products of the Magnolia sieved is less than 0.00001% by weight, the effect is difficult to appear, and when it exceeds 30% by weight, it is highly likely to cause irritation to the skin and may have a great influence on the stabilization of the formulation. In a preferred embodiment of the present invention was prepared a cosmetic composition comprising the extract of the Magnolia Magnolia 3.0% by weight, by applying it directly to the skin, skin wrinkle improvement effect and skin anti-aging effect, skin moisturizing effect, skin moisturizing atopic improvement effect and It was confirmed that the skin whitening effect is excellent.

Cosmetic composition for improving skin wrinkles of the present invention as an active ingredient includes components commonly used in cosmetic compositions in addition to the stem cell culture product of the Magnolia sieboldii, such as antioxidants, stabilizers, solubilizers, vitamins, pigments and fragrances Conventional adjuvants and carriers.

Cosmetic compositions of the invention may be prepared in any formulation conventionally prepared in the art, including, for example, solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing cleansing, It may be formulated as an oil, powder foundation, emulsion foundation, wax foundation, spray, and the like, but is not limited thereto. More specifically, it may be prepared in the form of a flexible lotion, nutrition lotion, nutrition cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray or powder.

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

When the formulation of the present invention is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used, in particular in the case of a spray, additionally chlorofluorohydrocarbon, propane Propellant such as butane or dimethyl ether.

When the formulation of the present invention is a solution or an emulsion, a solvent, a dissolving agent or an emulsifying agent is used as a carrier component, and examples thereof include water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, , 3-butyl glycol oil, glycerol aliphatic ester, polyethylene glycol or sorbitan fatty acid esters.

When the formulation of the present invention is a suspension, liquid carrier diluents such as water, ethanol or propylene glycol, suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol esters and polyoxyethylene sorbitan esters, and microcrystals are used as carrier components. Castle cellulose, aluminum metahydroxy, bentonite, agar and the like can be used.

When the formulation of the present invention is a surfactant-containing cleansing, the carrier component is an aliphatic alcohol sulfate, an aliphatic alcohol ether sulfate, a sulfosuccinic acid monoester, an isethionate, an imidazolinium derivative, a methyltaurate, a sarcosinate, a fatty acid amide ether. Sulfates, alkylamidobetaines, aliphatic alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, lanolin derivatives or ethoxylated glycerol fatty acid esters and the like can be used.

The cosmetic method of the present invention refers to all cosmetic methods for applying the cosmetic composition of the present invention to human skin. That is, all methods known in the art for applying the cosmetic composition to the skin belong to the cosmetic method of the present invention.

The cosmetic composition of the present invention may be used alone or in combination, or may be used by overlapping with other cosmetic compositions other than the present invention. In addition, the cosmetic composition with excellent skin protection effect according to the present invention can be used according to a conventional method of use, the number of times of use can be varied according to the user's skin condition or taste.

When the cosmetic composition of the present invention is a soap, surfactant-containing cleansing or surfactant-free cleansing formulation, it may be wiped off, peeled off or washed with water after application to the skin. As a specific example, the soap is liquid soap, powdered soap, solid soap or oil soap, the surfactant-containing cleansing formulation is a cleansing foam, cleansing water, cleansing towel or cleansing pack, the surfactant-free cleansing formulation is a cleansing cream , Cleansing lotion, cleansing water or cleansing gel, but is not limited thereto.

When the cosmetic composition comprising the active ingredient of the present invention is applied to the skin of a person, a skin aging prevention effect, skin wrinkle improvement effect, skin moisturizing effect, atopy improvement and skin whitening effect can be obtained.

Hereinafter, the present invention will be described in more detail through experimental examples. These experimental examples are only for explaining the present invention more specifically, it will be apparent to those skilled in the art that the scope of the present invention is not limited by these experimental examples in accordance with the gist of the present invention. .

Preparation Example 1: cultivation of Magnolia egg stem cells and production of culture products

After germinating Magnolia sieboldii seeds under certain conditions to surface sterilize stems, leaves, roots, foliar and flower buds with excellent growth, cut each and cut off, and then auxin is 2,4-D (0.10-0.50). mg / l), IAA (0.50-1.00gm / l), NAA (0.5-1.00mg / l) and cytokine added kinetin (0.10-0.50mg / l) and BAP (0.50-1.00mg / l) Callus stem cells were induced on the MS solid medium (Sigma) for about 2 weeks. These were separated separately and placed on fresh solid medium and subcultured at intervals of 4-5 to maintain callus.

Then, in order to secure more content of callus stem cells, Totipotent embryogenic stem cells, LB (Luria Bertani) Broth was incubated for 28 days in a 25 ° C suspension culture (gyrotory shaker). The obtained Magnolia squeezed the stem cells at low temperature to obtain a stem cell culture product, and then carried out a cytotoxicity test. Afterwards, the spermatozoa stem cell culture product was dissolved in 50% 1,3-butylene glycol aqueous solution, and diluted to a final concentration of 3% (v / v), which is a safe concentration, to the final concentration. Used for.

Comparative Production Example 1: Preparation of Magnolia egg Extract

For the comparative experiment on the stem cell culture of Magnolia Magnolia, the Magnolia Magnolia extract was prepared as follows.

Magnolia egg outpost was washed with clean water, dried and finely powdered. 70% alcohol solution was added to the powdered Magnolia egg powder in an amount of 10 times the weight of the powder, and then immersed for 5 days, filtered through Whatman filter paper, and then aged at 4 ° C. for 7 days at low temperature. After filtration and concentration using a vacuum concentrator, the concentrated powder was dissolved in 50% 1,3-butylene glycol aqueous solution so that the final concentration was 3% (v / v) and used in the experiment of the present invention.

Experimental Example 1 Cell Activity Enhancement Effect of Stem Cell Cultures

The following experiment was carried out to compare the cell activity enhancing effect of the Magnolia egg stem cell culture product prepared in Preparation Example 1 and the Magnolia Magnolia extract prepared in Comparative Preparation Example 1.

First, the concentrations of stem cells culture product of the Magnolia sieboldii were used in the experiment by 100 ㎍ / ㎖ each. The experiment was first inoculated human normal fibroblasts to 1 × 10 ⁴ cells in each well of a 96-well microplate, and incubated for 48 hours at 37 ℃ in DMEM medium.

Subsequently, the experimental group replaced with DMEM medium containing 2.5% of serum at a final concentration of 100 mg / ml of the Magnolia egg stem cell culture product of Preparation Example 1 was further cultured for 24 hours. In addition, as a comparative experiment, the comparison group, the Magnolia Magnolia, which was replaced with DMEM medium containing 2.5% of the Magnolia Magnolia extract of Comparative Preparation Example 1 at 100 ㎍ / ml, instead of the stem cell culture product of Preparation Example 1 The control group replaced with DMEM medium containing 2.5% of the serum without the stem cell culture was further cultured for 24 hours.

After incubation, 10 μl of 5 mg / ml MTT solution (Sigma) was added to observe the cell viability, and 20 μl of dimethyl sulfoxide solution was added to the well to dissolve, and the absorbance was measured by a microplate reader. The cell activity rate was determined by the assay. Cell activity rate was calculated according to the following equation.

In the above equation, D1 absorbance is the absorbance of the experimental group, D0 absorbance is the absorbance of the control group, and the absorbance is the value measured at 570 nm.

Cell Activity Effect of Stem Cell Culture of Stem Cells (Experimental Concentration 100㎍ / ㎖) Test substance Cell activity rate (%) Preparation Example 1 (stem) 88.2 Preparation Example 1 (leaf) 86.3 Preparation Example 1 (root) 87.3 Preparation Example 1 (leaf garden) 87.5 Preparation Example 1 (Flower Snow) 88.0 Comparative Preparation Example 1 68.4

According to Table 1, it was found that the cell activity rate is about 1.28 times better than that of the Magnolia egg extract of Comparative Example.

Experimental Example 2 Enhancement of Collagen Synthesis of Magnolia Egg Extract

Human normal fibroblasts were seeded to 1 × 10 ⁶ cells in each well of a 48 well microplate and incubated for 24 hours at 37 ° C. in DMEM medium. Subsequently, the experimental group replaced with DMEM medium without serum at the final concentration of the Magnolia egg stem cell culture product to 100 μg / ml, and the Magnolia Magnolia egg extract of Comparative Preparation Example 1 was replaced with 100 μg / ml instead of the stem cell culture product. Comparative group replaced with serum-free DMEM medium, control group replaced with serum-free DMEM medium containing no Magnolia egg extract was further cultured for 24 hours. After incubation, the supernatant of each well was collected and the amount of newly synthesized collagen was measured using a procollagen type I C-peptide (PICP) kit (Takara, Japan). The amount of PICP was converted to ng / ml, and the collagen biosynthesis increase rate was calculated according to Equation 2 below and the results are shown in Table 2.

Collagen Biosynthesis Enhancement Effect of Magnolia vulgaris Extract Test substance Collagen Biosynthesis
% Increase Preparation Example 1 (stem) 45.2 Preparation Example 1 (leaf) 44.2 Preparation Example 1 (root) 44.8 Preparation Example 1 (leaf garden) 43.8 Preparation Example 1 (Flower Snow) 44.6 Comparative Preparation Example 1 34.3

According to Table 2, the increase rate of collagen biosynthesis of the Magnolia Magnolia extract was about 1.3 times higher than that of the Magnolia Magnolia stem extract of Comparative Example 1.

Experimental Example 3. Inhibitory Effect on the Production of MMP-1 (MatrixMetalloProteinase-1) of Stem Cell Cultures

Human normal fibroblasts were seeded into 1 × 10 ⁶ cells in each well of a 48-well microplate and incubated for 24 hours at 37 ° C. in DMEM medium. Subsequently, the experimental group replaced with DMEM medium without serum by making the final concentration of the Magnolia egg stem cell culture product according to Preparation Example 100 to 100 μg / ml, and the Magnolia egg extract of Comparative Preparation Example 1 instead of the stem cell culture product Was replaced with DMEM medium without serum at 100 μg / ml, and the control group replaced with DMEM medium without serum containing no Magnolia egg extract was further cultured for 48 hours. After incubation, the supernatant of each well was collected and the amount of newly synthesized MMP-1 was measured using an MMP-1 assay kit (Amersham, USA), and then the amount of MMP-1 was converted to mg / ml. MMP-1 production inhibition was calculated according to the following equation (3) and the results are shown in Table 3.

MMP-1 Production Inhibitory Effect of Magnolia vulgaris Extract Test substance MMP-1 generation
% Inhibition Preparation Example 1 (stem) 86.9 Preparation Example 1 (leaf) 85.8 Preparation Example 1 (root) 87.3 Preparation Example 1 (leaf garden) 86.1 Preparation Example 1 (Flower Snow) 89.0 Comparative Preparation Example 1 74.3

According to Table 3, the MMP-1 production inhibition rate of the Magnolia Magnolia extract was about 1.17 times higher than that of the Magnolia Magnolia stem extract of Comparative Example 1.

Experimental Example 1-3 it was confirmed that the stem cells culture product of the Magnolia nymph of the present invention is excellent in cell activity enhancement effect, collagen synthesis enhancement effect and MMP-1 production inhibitory effect.

Experimental Example 4. Inhibitory effect of tyrosinase activity

Tyrosinase activity inhibition experiment was conducted to determine the degree of inhibition of tyrosinase activity of stem cells culture product according to Preparation Example 1. As tyrosinase activity inhibiting substrate, 1.5 mmol / L tyrosine (Sigma, USA) dissolved in sodium phosphate buffer (pH 6.8) was used. Chunma Magnolia stem cell culture product was prepared by diluting each concentration from 0 to 3.0% using 0.05 M sodium phosphate buffer (pH 6.8), and adding 40 μl of tyrosine to 240 μl of the sample solution. 20 μl of tyrosinase (Sigma, 1,500 U / mL) was added and reacted at 37 ° C. for 15 minutes, and then absorbance at 490 nm (absorbance of the experimental group, but the control absorbance of 0% of the Magnolia siebolis). As a comparison group, the extract of the Magnolia Magnolia according to Comparative Preparation Example 1 was used, and arbutin, a whitening agent, was used as a positive control group. The stem cell culture products of the Magnolia sieved stems, leaves, roots, leaf gardens, and flower buds were used. However, all stems showed similar patterns, and only stem cultures were described by concentration. The cultures of the flower buds described only the results of 0.5% without any remaining concentrations. Inhibition of tyrosinase activity was calculated according to Equation 4 below, and the results are shown in Table 4 below.

Experimental Sample (%) % Inhibition of tyrosinase activity Magnolia egg extract amount 0.50 35.2 Magnolia egg stem cell culture 0.50 (stem) 47.2 Magnolia egg stem stem culture product 1.00 (stem) 49.8 Magnolia egg stem stem culture product 1.50 (stem) 56.9 Magnolia egg stem cell culture product 2.00 (stem) 61.6 Magnolia egg stem stem culture product 2.50 (stem) 70.4 Magnolia egg stem stem culture product 3.00 (stem) 92.1 Magnolia egg stem culture 0.50 (leaves) 47.9 Magnolia egg stem cell culture 0.50 (root) 48.1 Magnolia egg stem stem culture product 0.50 47.1 Magnolia Egg Stem Cell Culture 0.50 48.3 Arbutin 2.50 71.2

As can be seen from the results of Table 4, when the barley stem cell culture product of the present invention treated 0.50% to 3.00%, it showed an excellent effect on the inhibition of tyrosinase activity, and the activity inhibition degree was also It was found to increase depending on the culture concentration. In addition, even when compared with the same concentration of arbutin, the effect of arbutin appeared to be similar or higher, indicating that the magnolia is excellent in whitening effect of stem cell culture products.

Experimental Example 5. Melanin synthesis inhibitory effect

Melanoma-derived melanoma cells B-16 F1 (Melanocyte, B16-F1 Korea Cell Line Bank) were seeded in 12-well plates (Falcon, USA) to 1 × 10 ⁴ cell / well and cultured for one day to stabilize the cells. Then, the Magnolia egg stem cell culture product according to Preparation Example 1 was treated for each concentration of 0 to 2.0% per well and cultured for 3-4 days. The cultured plate was centrifuged, the culture solution was removed, cells were lysed with 500 μl of 1N NaOH solution dissolved in dimethyl sulfoxide (DMSO), and then heated in a 90 ° C. water bath for 10 minutes. After centrifugation again, the supernatant was absorbed at a wavelength of 570 nm (absorbance of the experimental group, but in the case of 0% of the Magnolia egg extract, the control absorbance).

The addition group to which the Magnolia egg extract was added was used as a comparison group, and arbutin, a whitening agent, was used as a positive control. Melanin generation inhibition (%) was calculated according to Equation 5 below, and the results are shown in Table 5. The stem cell culture products of the Magnolia sieved stems, leaves, roots, leaf gardens, and flower buds were used. However, all stems showed similar patterns, and only stem cultures were described by concentration. The cultures of the flower buds described only the results of 0.5% without any remaining concentrations.

Experimental Sample (%) % Melanin production inhibition Magnolia Egg Extract 0 0 Magnolia Egg Extract 0.5 31.2 Magnolia Egg Extract 1.0 49.7 Magnolia Egg Extract 1.5 55.9 Magnolia Egg Extract 2.0 79.3 Magnolia egg stem cell culture 0.5 (stem) 36.6 Magnolia egg stem cell culture product 1.0 (stem) 56.8 Magnolia egg stem culture product 1.5 (stem) 68.4 Magnolia egg stem culture 2.0 (stem) 86.8 Magnolia egg stem culture 0.5 (leaf) 36.4 Magnolia egg stem culture 0.5 (root) 36.8 Magnolia egg stem cell culture product 0.5 36.2 Magnolia egg stem cell culture 0.5 (flower bud) 37.0 Arbutin 0.5 50.4

As can be seen from the results of Table 5, the treatment of the Magnolia egg stem cell culture product of the present invention showed an excellent effect on the inhibition of melanin production when 0.50% ~ 2.00%, and also the activity inhibition degree of the Magnolia egg stem cell culture product It was found to increase depending on the concentration. And compared with the same concentration of arbutin, arbutin shows the same effect as can be seen that the whitening effect of the stem cell culture product of the Magnolia bran.

Thus, the Magnolia of the present invention is to prepare a cosmetic containing a stem cell culture product as an active ingredient to present a prescription example, but the cosmetic composition of the present invention is not limited to this prescription example. In addition, 3% (w / v) of the culture product obtained from the stem of the Magnolia egg stem cell culture product of Preparation Example 1 was used as the Magnolia egg stem cell culture product used in the following prescription example. In addition to the stem, cultures of leaves, roots, foliar groups, and flower buds may be applied at the same concentration.

Formulation Example 1 Softening Cosmetic (Skin Lotion)

As shown in Table 6, the flexible cosmetic was prepared according to a conventional method.

ingredient Content (% by weight) Magnolia egg stem cell culture 3.0 glycerin 5.0 1,3-butylene glycol 3.0 PEG 1500 1.0 Allantoin 0.1 DL-Panthenol 0.3 Benzophenone-9 0.04 EDTA-2Na 0.02 Sodium hyaruronate 5.0 ethanol 10.0 Octyldodec-16 0.2 Polysorbate 20 0.2 incense 0.02 Purified water to 100

Formulation Example 2. Nutrients (Milk Lotion)

Nutrients were prepared according to a conventional method as shown in Table 7.

ingredient Content (% by weight) Magnolia egg stem cell culture 3.0 Glyceryl Stearate SE 1.5 Cetearyl alcohol 1.5 lanolin 1.5 Polysorbate 60 1.3 Sorbitan stearate 0.5 Cured Plants 1.0 Mineral logistics 5.0 Squalane 3.0 Trioctanoine 2.0 Dimethicone 0.8 Carboxy Vinyl Polymer 0.12 glycerin 5.0 1,3-butylene glycol 3.0 Sodium hyaluronate 2.0 Triethanolamine 0.12 incense 0.01 Purified water to 100

Formulation Example 3. Nutrition Cream

Nutritional cream was prepared according to a conventional method as shown in Table 8.

ingredient Content (% by weight) Magnolia egg stem cell culture 3.0 Lipophilic monostearate 2.0 Cetearyl alcohol 2.2 Stearic acid 1.5 Polysorbate 60 1.5 Sorbitan stearate 0.6 Cured Plants 1.0 Mineral logistics 5.0 Squalane 3.0 Trioctanoine 2.0 Dimethicone 1.2 Sodium magnesium silicate 0.1 glycerin 5.0 1,3-butylene glycol 3.0 Sodium hyaluronate 2.0 Triethanolamine 1.0 incense 0.02 Purified water to 100

Formulation Example 4 Massage Cream

Massage cream was prepared according to a conventional method as shown in Table 9.

ingredient Content (% by weight) Magnolia egg stem cell culture 3.0 Lipophilic monostearate 1.5 Cetearyl alcohol 1.5 Stearic acid 1.0 Polysorbate 60 1.5 Sorbitan stearate 0.6 Isostearyl isostearate 5.0 Mineral logistics 35.0 Squalane 5.0 Dimethicone 0.5 Hydroxyethyl cellulose 0.12 glycerin 6.0 Triethanolamine 0.7 incense 0.01 Purified water to 100

Formulation Example 5. Essence

Essence was prepared according to a conventional method as shown in Table 10 below.

ingredient Content (% by weight) Magnolia egg stem cell culture 3.0 glycerin 10.0 Betaine 5.0 PEG 1500 2.0 Allantoin 0.1 DL-Panthenol 0.3 EDTA-2Na 0.02 Benzophenone-9 0.04 Hydroxyethylcellulose 0.1 Sodium hyaruronate 8.0 Carboxy Vinyl Polymer 0.2 Triethanolamine 0.18 Octyldodecanol 0.3 Octyldodes-16 0.4 ethanol 6.0 incense 0.02 Purified water to 100

Formulation Example 6 Pack

To prepare a pack according to the conventional method as shown in Table 11.

ingredient Content (% by weight) Magnolia egg stem cell culture 3.0 Poly Secret Alcohol 15.0 Cellulose sword 0.15 glycerin 3.0 PEG 1500 2.0 DL-Panthenol 0.4 Cyclodextrin 0.15 Allantoin 0.1 Glycyrisin Monoammonium 0.3 Nicotinamide 0.5 ethanol 6.0 PEG 40 Cured Castor Oil 0.3 incense 0.02 Purified water to 100

Experimental Example 6. Moisturizing effect of the cosmetic

Cosmetics containing a stem cell culture product (Example 1), which is cultured from stems of the Magnolia egg prepared in Preparation Example 1 in order to measure the moisturizing effect on the skin in cosmetics containing stem cell culture product Cosmetics (Comparative Example 1) that do not contain the stem cells culture product (comparative Example 1), cosmetics of the formulation containing the extract of the Magnolia Magnolia prepared in Comparative Preparation Example 1 in place of the stem cell culture product (Comparative Example 2) It was prepared in a composition of 12.

ingredient Example 1 Comparative Example 1 Comparative Example 2 Magnolia egg stem culture 3.0 - - Magnolia Egg Extract - - 3.0  vaseline 7.0 7.0 7.0  Liquid paraffin 10.0 10.0 10.0  Beeswax 2.0 2.0 2.0  Polysorbate 60 2.0 2.0 2.0  Sorbitan sesquioleate 2.5 2.5 2.5  Squalane 3.0 3.0 3.0  Propylene glycol 6.0 6.0 6.0  glycerin 4.0 4.0 4.0  Triethanolamine 0.5 0.5 0.5  Carboxy Vinyl Polymer 0.5 0.5 0.5  Tocopheryl acetate 0.1 0.1 0.1  incense 0.01 0.01 0.01  Purified water to 100 to 100 to 100

The units in the table are by weight (% by weight).

The skin moisturizing effect on the cosmetics of Example 1 and Comparative Examples 1 and 2 was measured as follows.

After dividing 30 healthy women into three groups (A, B, C) at 25 ° C., 45% relative humidity and no air flow, the cosmetics of Example 1 were applied to group A for group B. After applying the cosmetic of Example 2, the cosmetics of Comparative Example 1 for the group C, according to Table 12, after applying the appropriate amount to the upper arm of the arm three times a day for 6 weeks, TEWAMETER TM210 The transdermal moisture loss, that is, the TEWL (Transepidermal Water Loss) value was measured as the TEWL value change ΔTEWL over time, and the CORNEOMETER CM 820 PC (C + K electronic GmbH. ) To measure the moisturizing effect by quantifying the moisture content of the skin from 0 to 150 with the change of electrical conductivity according to the epidermal moisture content. The experimental results are shown in Table 13 below.

Skin moisturizing effect Test Product △ TEWL Corneometer value Example 1 1.2 120 Comparative Example 1 6.1 86 Comparative Example 2 2.3 111

As can be seen from the results of Table 13, the cosmetic containing the stem cell culture product of Magnolia ovary (Example 1) is a cosmetic containing only the extract of Magnolia ovary stem cell culture (Comparative Example 1) and extract of Magnolia ovary Compared with (Comparative Example 2), the amount of transdermal moisture was greatly improved, and it was found that the stem cell culture product of the Magnolia nectar exhibits a moisturizing effect.

In addition, in the Corneometer value, which measured the moisture content of the skin, Example 1 (cosmetics containing the Magnolia egg stem cell culture product) was compared to Comparative Example 1 (cosmetics containing the Magnolia egg stem cell culture product) and Comparative Example 2 ( It can be seen that a higher increase compared to cosmetics containing the extract of Magnolia sieboldii. In other words, the skin moisture content of the cosmetic containing the stem cell culture product of the Magnolia Magnolia was measured high, which proves that the moisturizing effect of the cosmetic containing the stem cell culture of the Magnolia Magnolia is very effective.

Experimental Example 7. Skin wrinkle improvement effect of cosmetics containing stem cell culture product

With the same composition as shown in Table 12, the cosmetic containing the stem cell culture product of the Magnolia bran of the present invention (Example 1), the cosmetic containing no stem cell culture product of the Magnolia bran (Comparative Example 1) The effect of improving skin wrinkles on cosmetics containing the extract (Comparative Example 2) was measured. The measurement was carried out by mechanical evaluation method and visual evaluation method by professional personnel as follows.

7-1. Mechanical evaluation

Thirty women aged 30 or older were divided into three groups, and the cosmetics of Example 1 were administered to Group A, the cosmetics of Comparative Example 1 to Group B, and the cosmetics of Comparative Example 2 to Group C were formulated according to Table 12. (3 times / day, 0.2 g / time) was applied to the area of the upper left side of the upper arm (2 times / day, 0.2 g / time) for 3 months, and a replica of the skin wrinkles was floated using a transparent silicone solution (SKINVISIOMETER SV400, C + K Electronics GmbH Germany) was used to measure the improvement of skin wrinkles. Three-dimensional analysis of the image of the replica with a CCD camera, the sum of the roughness of each wrinkle (R _m : m is an integer of 1 or more) divided by the number of wrinkles is the average wrinkle roughness (R _z ) of the following formula (6) Wrinkle improvement effect was analyzed. The experimental results are shown in Table 14 below.

Skin wrinkle improvement (equipment evaluation) Example 1 △ R _z Comparative Example 1 △ R _z Comparative Example 2 △ Rz Subject 1 -29 Subject 21 -11 Subject 31 -23 Subject 2 -31 Subject 22 -5 Subject 32 -19 Subject 3 -26 Subject 23 3 Subject 33 -8 Subject 4 -13 Subject 24 -7 Subject 34 -21 Subject 5 -37 Subject 25 -15 Subject 35 -15 Subject 6 -28 Subject 26 7 Subject 36 -12 Subject 7 -35 Subject 27 -8 Subject 37 -20 Subject 8 -22 Subject 28 -2 Subject 38 -29 Subject 9 -25 Subject 29 -13 Subject 39 -17 Subject 10 -27 Subject 30 -2 Subject 40 -23 Average -27.3 Average -5.3 Average -18.7

As can be seen in Table 14, in the case of cosmetics containing no stem cells culture product (Comparative Example 1) did not show the effect of improving wrinkles, cosmetics containing stem cells culture product (Example 1) In the case of the wrinkle improvement was found to be excellent. In addition, in the case of the cosmetic containing the stem cell culture product (Example 1) and the cosmetic containing the extract of the Magnolia Magnolia (Comparative Example 2), the height (roughness) of the skin wrinkles after 3 months (T3) was 27.3, respectively. It was found that the lowering of about 18.7 μm and about 18.7 μm (p <0.01) resulted in better wrinkle improvement compared to Comparative Example 2. Through this, it can be seen that the stem cell culture product of the Magnolia sieved wrinkles.

7-2. Visual evaluation of experts

Thirty women aged 30 and older were divided into three groups, and the cosmetics of Example 1 were administered to Group A, the cosmetics of Comparative Example 1 to Group B, and the cosmetics of Comparative Example 2 to Group C according to Table 12. The degree of improvement of skin wrinkles (ΔW) was measured at one month intervals (T0, T1, T2, T3) while applying for 3 months around the eyes. The measurement was based on two expert double-blind test objectives (Table 15) and subjects' subjective assessments (Table 16), followed by a global photodamage score (0: none, 1: none / mild, 2). : mild, 3: mild / moderate, 4: moderate, 5: moderate / severe, 6: severe, 7: very severe, Ref.: Arch Dermatol vol. 137 (8): 1043-1051, 2001) It was.

Skin wrinkle improvement effect (expert evaluation by expert) Skin wrinkle improvement Example 1 Comparative Example 1 Comparative Example 2 T0 T1 T2 T3 T0 T1 T2 T3 T0 T1 T2 T3 Subject 1 4 4 3 2 Subject 11 3 3 3 2 Subject 21 5 4 4 3 Subject 2 3 2 One One Subject 12 4 4 4 3 Subject 22 3 3 3 2 Subject 3 3 One One One Subject 13 5 4 3 3 Subject 23 3 3 3 3 Subject 4 3 2 2 2 Subject 14 3 3 3 3 Subject 24 4 4 3 2 Subject 5 2 0 0 0 Subject 15 2 2 One One Subject 25 6 6 4 3 Subject 6 4 2 2 One Subject 16 4 4 3 3 Subject 26 4 4 2 2 Subject 7 5 3 One One Subject 17 4 4 3 3 Subject 27 3 2 2 2 Subject 8 5 5 4 4 Subject 18 3 3 3 3 Subject 28 3 3 One One Subject 9 3 2 2 One Subject 19 4 3 3 3 Subject 29 2 One One 0 Subject 10 4 3 One One Subject 20 3 2 One 2 Subject 30 4 4 4 2 △ W ( T3 -T0) = 2.2 ΔW ( T3 -T0) = 0.9 ΔW ( T3 -T0) = 1.7

Skin wrinkle improvement effect (subjective evaluation of subject) Skin wrinkle improvement Example 1 Comparative Example 1 Comparative Example 2 T0 T1 T2 T3 T0 T1 T2 T3 T0 T1 T2 T3 Subject 1 4 4 3 One Subject 11 3 3 3 2 Subject 21 5 5 3 3 Subject 2 3 2 One 0 Subject 12 4 4 4 2 Subject 22 3 2 2 2 Subject 3 3 One One One Subject 13 5 4 4 2 Subject 23 3 3 3 2 Subject 4 3 2 2 One Subject 14 3 3 3 2 Subject 24 4 3 2 2 Subject 5 2 0 0 0 Subject 15 2 2 One 0 Subject 25 6 6 4 4 Subject 6 4 2 2 One Subject 16 4 4 4 4 Subject 26 4 4 3 3 Subject 7 5 3 One One Subject 17 4 4 3 3 Subject 27 3 2 One One Subject 8 5 5 4 3 Subject 18 3 3 3 3 Subject 28 3 3 One 0 Subject 9 3 2 2 One Subject 19 4 3 3 3 Subject 29 2 2 One 0 Subject 10 4 3 One One Subject 20 3 3 3 2 Subject 30 4 4 3 2 △ W ( T3 -T0) = 2.6 △ W ( T3 -T0) = 1.2 △ W ( T3 -T0) = 1.8

As can be seen from the results of Tables 15 and 16, the cosmetic containing the stem cell culture product of Magnolia sieboldii (Example 1) showed a skin wrinkle improvement effect in both an objective and subjective evaluation.

In the case of Comparative Example 1, both the expert and the subject showed a slight improvement of wrinkles, but it was not a significant improvement in the range, and it seems to be a result of long-term use of the conventional cosmetics. In addition, it can be seen that the cosmetics containing the stem cell culture products (Example 1) have a more effective skin wrinkle improvement effect than the cosmetics containing the extract of the Magnolia sieboldii (Comparative Example 2).

Experimental Example 8. Skin whitening effect of cosmetics containing stem cell culture product

With the same composition as shown in Table 12, the cosmetic containing the stem cell culture product of the Magnolia sieboldii stem of the present invention (Example 1) and the cosmetic composition (Comparative Example 1) and the cheonna Magnolia extract containing no stem cell culture products Skin whitening effect for the cosmetic containing (Comparative Example 2) was measured.

Three groups (A, thirty-five) of thirty healthy adult men and women with blemishes, freckles and pigmentation in order to test pigmentation improvement as a skin whitening effect on cosmetics including stem cell culture products of the present invention After dividing by B, C), the cosmetics of Example 1 were included in the A group, the cosmetics of Comparative Example 1 were included in the B group, and the stem cell culture product of the Magnolia bran was included in the C group. After applying the appropriate amount of the cosmetic preparation of Comparative Example 2 containing the extract of Magnolia Magnolia twice a day for 12 weeks, the change in skin color was measured using chromata (Minolta CR300) to measure the change in color brightness (ΔL), Objective visual observation by a plurality of skilled workers and subjective visual observation by a subject were performed, and the effects were measured according to the following classification. The results are shown in Table 17 below. At this time, the degree of pigmentation improvement was evaluated by classifying into the following seven grades.

    <Color Deposition Improvement Evaluation Criteria>

       -3: very worse

-2: worse

-1: slightly worse

0: no change

1: slightly improved

2: improve

3: highly improved

Assessment of pigmentation improvement Skin-colored
Change in brightness (ΔL) Skilled
Objective evaluation Subject's
Subjective evaluation A B C A B C A B C Subject 1 6.0 0.6 5.4 Subject 11 3 0 One Subject 21 3 0 2 Subject 2 6.2 0.0 4.9 Subject 12 3 0 One Subject 22 3 0 One Subject 3 7.1 0.1 4.2 Subject 13 2 0 2 Subject 23 2 0 One Subject 4 5.9 1.2 5.7 Subject 14 2 One One Subject 24 2 0 One Subject 5 6.6 0.2 3.2 Subject 15 3 0 3 Subject 25 2 One 0 Subject 6 7.2 0.1 4.1 Subject 16 2 0 2 Subject 26 3 0 2 Subject 7 4.9 0.5 3.7 Subject 17 2 0 One Subject 27 2 0 One Subject 8 7.6 0.3 3.8 Subject 18 2 0 One Subject 28 3 0 One Subject 9 7.4 0.0 4.6 Subject 19 3 One 0 Subject 29 One 0 One Subject 10 6.5 0.9 5.1 Subject 20 One 0 One Subject 30 3 0 One medium 6.54 0.39 4.47 medium 2.3 0.2 1.3 medium 2.4 0.1 1.1

As can be seen in Table 17, the cosmetic containing the stem cell culture product of the Magnolia Magnolia (Example 1) contains a cosmetic (Comparative Example 1) and the Magnolia Magnolia extract that do not contain any of the Magnolia egg stem cell culture products. It was found that the pigmentation improvement effect was superior to the cosmetics (Comparative Example 2).

In addition, in the objective evaluation by the expert and the subjective evaluation of the subject, it can be seen that the cosmetic containing the stem cell culture product of the Magnolia sieboldii (Example 1) of the present invention effectively improves skin pigmentation.

Claims (10)

Granola is a cosmetic composition containing a stem cell culture product as an active ingredient, wherein the stem cells are derived from callus. According to claim 1, wherein the composition is a cosmetic composition, characterized in that for preventing skin aging. The cosmetic composition according to claim 1, wherein the composition is for improving skin wrinkles. According to claim 1, wherein the composition is a cosmetic composition, characterized in that for skin whitening. According to claim 1, wherein the composition is a cosmetic composition, characterized in that for moisturizing the skin. delete The culture solution according to any one of claims 1 to 5, wherein the product of the Magnolia egg stem cell culture is obtained by culturing stem cells of a site selected from the group consisting of stems, leaves, roots, leaf stems, and flower buds of the Magnolia egg. Cosmetic composition, characterized in that the extract obtained by crushing the extract or cultured stem cells. The cosmetic composition according to claim 7, wherein the content of the Magnolia sieved stem cell culture product is 0.00001 to 30% by weight based on the total weight of the cosmetic composition. The cosmetic composition according to any one of claims 1 to 5, wherein the cosmetic composition is a solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, cleansing, oil, powder foundation, emulsion emulsion, wax foundation Cosmetic composition, characterized in that it has a formulation selected from the group consisting of a pack, massage cream and spray. delete