KR20200026646A - Cosmetic Composition for Improving Skin Condition Comprising Plant cell complex cultures to improve skin radiance and vitality - Google Patents

Abstract

The present invention relates to a cosmetic composition for improving a skin condition, comprising plant cell culture extracts/filtrates as a complex and, more specifically, to a cosmetic composition which comprises, as an effective component, one or more selected from the group consisting of Leontopodium Alpinum Callus culture or an extract thereof, Eryngium Maritimum Callus culture or an extract thereof, Crithmum Maritimum Callus culture or an extract thereof, Malus Domestica fruit cell culture or an extract thereof, Vitis Vinifera (Grape) fruit cell culture or an extract thereof, an Adenium Obesum leaf cell extract, a Lavender leaf cell extract, a Vitex Negundo leaf cell extract, Nelumbo Nucifera leaf cell culture powder, a Commiphora Myrrha leaf cell extract, and a Jasmine leaf cell extract, and thus exhibits effectiveness in a skin brightening function, a skin invigorating function, prevention or reduction of skin wrinkles, enhancement or improvement of skin elasticity, skin tightening, protection of skin barrier functions, skin protection from UV light, alleviation of atopic dermatitis, and anti-inflammatory, antioxidant, and anti-aging functions.

Description

Cosmetic composition for Improving Skin Condition Comprising Plant cell complex cultures to improve skin radiance and vitality

The present invention relates to a cosmetic composition for skin improvement comprising a plant cell complex culture or extract thereof, and more particularly, Edelweiss callus culture or its extract, Seaside Eringo callus culture or its extract, Roxamfire callus Cultures or extracts, apple cell cultures or extracts thereof, grape cell cultures or extracts thereof, desert rosette leaf cell cultures or extracts, lavender leaf cell cultures or extracts, Chinese chestnut leaf cell cultures or extracts, By containing lotus leaf cell culture or extract, myrrh leaf cell culture or extract, Arabian jasmine leaf cell culture or extract as an active ingredient, to give skin vitality, increase skin vitality, prevent or improve skin wrinkles, improve skin elasticity, improve , Skin convergence, skin barrier function protection, skin protection from ultraviolet light, antioxidant, anti-aging It relates to a cosmetic composition with the function.

Recent research has identified a biological clock molecular network that governs circadian rhythms. Since the 1970s, the biological clock that regulates the expression of genes related to physiological metabolism and body rhythms of organisms according to the day and night cycles of the earth's rotation is becoming more detailed since the 1970s, and the study of diseases related to the cycle is being actively conducted. Jeffrey C. Hall, Professor, Maine University, Michael Rosbash, Brandeis University, and Michael W. Young, Professor Rockefeller University, have a model animal, Drosophila, in a 24-hour day and night cycle. He was honored with the 2017 Nobel Prize for his work on identifying the mechanisms by which circadian genes appear. The winners followed up the genes in the 1980s, after the earliest existence of a gene associated with the clock, revealed the basic mechanism of the clock. The molecular network of these biological clock genes contains clock genes that develop as they adapt to day and night changes. Clock genes called CLOCK (Clock circadian regulator) and PER1 (Period circadian protein) are present in organs such as the heart, lungs, fat, blood vessels and kidneys, as well as brain tissues such as the cerebellum, midbrain and hypothalamus. .

Their research has shown that the biological cycle is a biological phenomenon that occurs when the concentration of a protein (PER) expressed by a gene called pyriod changes every 24 hours. As the PER protein produced by the pyriod gene increases and continues to accumulate in the cytoplasm outside the nucleus, the PER protein now binds to other enzymes and enters the nucleus to inhibit the expression of the pyriod gene. The cycle of the livelihood clock appears as a molecular oscillation that increases and decreases the concentration of PER protein. Later, as other genes (proteins) were discovered that regulate the stability and functionality of these basic mechanisms, the mechanism of the 'bio clock', which is precisely and finely regulated, became known. At this stage, the dimer of CLOCK and BMAL1 protein triggers the expression of Period (PER) and Cryptochrome (CRY) family of feedback genes located in the lower stages as transcription factors, and the gene products burn higher level transcription factors. It forms a series of molecular circulatory structures that activate. These molecular networks are known to affect a variety of physiological processes by triggering the circadianity of genes regulated downstream (Stratmann and Schibler, 2006, J Biol Rhythms 21: 494). In other words, the circadian rhythm is controlled to be repeated every day through the associated feedback of the CLOCK / PER protein to activate the biological clock.

Skin cells, keratinocytes, melanocytes, and fibroblasts also have independent biological clock systems. The skin biological clock regulates skin physiological activity, which is the circadian rhythm of skin cell proliferation and differentiation, hydration and transepidermal water loss (TEWL), capillary blood flow, sebum production, temperature, surface pH, and wrinkle formation. Seems. If the rhythm of this process is broken, it will not produce the necessary physiological response at the right time, break the homeostasis of the skin, promote skin aging, and seriously cause skin cancer.

As human skin ages, it changes with a variety of internal and external factors. Internally, the secretion of various hormones that regulate metabolism is reduced, and the function and activity of cells of immune cells is reduced, thereby reducing the biosynthesis of immune proteins and constituent proteins necessary for the living body. Externally, due to the destruction of the ozone layer, the amount of ultraviolet rays reaching the surface of the sunlight increases and environmental pollution is intensified, resulting in the increase of free radicals and active harmful oxygen, which reduces the thickness of the skin and increases wrinkles. Not only does it reduce elasticity, but it also causes various changes such as frequent skin problems.

As skin aging progresses, keratinocytes and fibroblasts may divide, reduce collagen synthesis, increase MMP production, and increase melanin signaling, resulting in increased wrinkles and reduced skin elasticity. And blemishes, freckles and blotch are to be increased. In particular, the hormone balance is broken around 40s, and the moisture content of the stratum corneum, which is important for cell regeneration, collagen synthesis, and skin moisturization, is significantly lowered. It is suggested that this is due to the decrease in the stratum corneum lipid composition, the decrease in the natural moisturizing factor, the decrease in the amount of glycerol in the dry skin. As a result, in aging skin, the amount of water supplied from the lower epidermis to the stratum corneum is insufficient, resulting in severe skin dryness, which causes inflammation and pruritus. On the other hand, the wrinkles of the skin are due to the imbalance of collagen synthesis and degradation, and the type I collagen synthesis and the degradation enzyme MMP-1 are usually balanced in the skin. However, in photo-aged skin, the synthesis of type I and III collagen is decreased, and the activity of MMP-1 is increased. These MMPs (matrix metalloproteinases) are proteolytic enzymes that degrade the extracellular matrix and are known to be involved in wound healing or tissue regeneration during normal conditions.

The present inventors have made efforts to discover materials having various functionalities such as skin vitality, skin vitality improvement, skin wrinkle improvement, skin regeneration, skin barrier function enhancement, wrinkle improvement and the like, and thus, Edelweiss callus culture or its extracts, seasides, Lingo callus culture or its extract, Roxamfire callus culture or its extract, Apple cell culture or its extract, Grape cell culture or its extract, Desert rose leaf cell extract, Lavender leaf cell extract, Chinese yeast tree leaf cell Extract, lotus leaf cell culture powder, myrrh leaf cell extract, Arabian jasmine leaf cell extract reveals that it has a biological clock control effect, confirmed that it can be a variety of functional cosmetic material, and completed the present invention.

Accordingly, an object of the present invention is an Edelweiss callus culture or its extract, Seaside Eringo callus culture or its extract, Loxamfire callus culture, or extract, Apple cell culture or its extract, Grape cell culture or its extract Skin rejuvenating cosmetic composition containing any one or more of desert rose leaf cell extract, lavender leaf cell extract, Chinese chestnut leaf cell extract, lotus leaf cell culture powder, myrrh leaf cell extract, Arabian jasmine leaf cell extract To provide a composition.

Another object of the present invention, the Edelweiss callus culture or its extract, Seaside Eringo callus culture or its extract, Loxam fire callus culture, or extract, apple cell culture or its extract, grape cell culture or its For skin improvement containing any one or more of extracts, desert rose leaf cell extracts, lavender leaf cell extracts, Chinese yeast tree leaf cell extracts, lotus leaf cell culture powder, myrrh leaf cell extracts, Arabian jasmine leaf cell extracts It is to provide a method for producing a cosmetic composition.

In order to solve the above problems, the present invention is an Edelweiss callus culture or its extract, Seaside Eringo callus culture or its extract, Loxam fire callus culture, or extract, apple cell culture or its extract, grape cell culture or The extract, desert rosette leaf cell extract, lavender leaf cell extract, Chinese yeast tree leaf cell extract, lotus leaf cell culture powder, myrrh leaf cell extract, Arabian jasmine leaf cell extract any one or more thereof and a method for preparing the same, the extract It provides a cosmetic composition containing.

Hereinafter, the present invention will be described in more detail.

In one aspect, the present invention, Edelweiss callus culture or its extract, Seaside Eringo callus culture or its extract, Loxam fire callus culture, or extract, Apple cell culture or its extract, Grape cell culture or its extract Skin rejuvenating cosmetic composition containing any one or more of desert rose leaf cell extract, lavender leaf cell extract, Chinese chestnut leaf cell extract, lotus leaf cell culture powder, myrrh leaf cell extract, Arabian jasmine leaf cell extract It relates to a composition. Specifically, the callus culture or its extract is not limited to the content, but may contain 0.1 to 10% by weight relative to the total weight of the composition.

In the present invention, the description of each plant of each plant cell to be an active ingredient is as follows.

(1) Edelweiss callus culture extract, Leontopodium Alpinum Callus Culture Extract, LEONTOPODIUM ALPINUM

Edelweiss is a plant belonging to the Asteraceae family and lives in the highlands of Europe and Asia, including the Alps. It is 10 ~ 20cm high and covered with white wool. The leaves are relatively high in the roots, slightly on the stems, and linear. At the end of the stem, artillery runs around, spreads out all over, with some headshots in the center.

(2) Seaside Eringo callus culture filtrate, Eryngium Maritimum Callus Culture Filtrate, ERYNGIUM MARITIMUM

The Seaside Eeringo genus Eryngium is derived from the old Greek name eryggion. It is 2 years or perennial plant that grows about 60cm in height. The upper branches diverge a lot. Lower leaves are circular heart-shaped, divided into three to five, with bed rests. The upper leaves have no foliar and cover the stems. There are about 6 guns and a slightly wide ovate with two or three crabs on both sides. At the end is a bed. Flowers bloom from summer to autumn, and inflorescences are round and metallic pale purple.

(3) Crucmum Maritimum Callus Culture Filtrate

Rocksampire is an edible wild plant belonging to the genus Amphitaceae, which is a special plant that lives mainly in the rocky creeks of Europe's pristine beaches. It is an extreme plant that is resistant to water, high salinity, temperature and wind. Flowers grow at the end of stems and grow in greenish yellow with small petals and five petals. When the plant is broken, you can smell the fragrant lemon. For centuries, it has been used as a food ingredient and as a medicinal product. Drinking seeds, roots and leaves in wine helps with jaundice and urinary tract pain.It is also effective for scurvy caused by lack of vitamin C. I took it on.

(4) Apple Cell Culture Extract, Malus Domestica Fruit Cell Culture Extract, MALUS DOMESTICA

The fruit of the dicotyledonous rosewood rose and deciduous arbor plant is 5 ~ diameter round and its color is usually red or yellow. Origin is known as Balkans. Alkaline foods are low in calories and contain many healthy ingredients. Dietary fiber helps to prevent arteriosclerosis by releasing harmful cholesterol that builds up in your blood vessels and increasing your beneficial cholesterol.

(5) Vitis Vinifera (Grape) Fruit Cell Extract, VITIS VINIFERA

Grapes are fleshy, juicy, seed-bearing berries that ripen in black from July to August. Its origin is known as the Black Sea Coast and the Kafka region in western Asia, and our country has a record of it in the collections of the Goryeo period.

(6) Desert Rose Leaf Cell Extract, Adenium Obesum Leaf Cell Extract, ADENIUM OBESUM

A dicotyledon evergreen shrub, one of the succulents found in desert climates such as tropical Africa and Arabia. The name of the flower comes from Aden, one of the native areas of Yemen. It is called 'Desert Rose' because the flower is pretty like a rose.

(7) Lavandula Angustifolia (Lavender) Leaf Cell Extract, LAVANDULA ANGUSTIFOLIA

The name Lavandula comes from the Latin word for "wash," because the Romans loved to put lavender in water when bathing or washing. It is a perennial plant of the nectaraceae with purple star-shaped small flowers on the stalks, producing essential oils with fragrant oil glands among the hairs covering the flowers, leaves and stems.

(8) Chinese Chaste Tree Leaf Cell Extract, Vitex Negundo Leaf Cell Extract, VITEX NEGUNDO

It is a shrub of genus Verbena in southern Europe, whose leaves are hairy and lush with fragrant pale purple flowers.

(9) Lotus leaf cell culture powder, Nelumbo Nucifera Leaf Cell Culture Powder, NELUMBO NUCIFERA

The kite is native to southern Asia and northern Australia. It is a plant that grows in mud and is clean and noble, and has been friendly to people in many countries. Root stock is thick and stretches to the side, with many nodes, especially in autumn, with thick tips. Flowers bloom in July-Month, red or white, hanging on one end of stalk, diameter 15 ~, with thorns on stalk. The petals are upside down, with several stamens. Calyx is large and flat, about 10cm in diameter, and fruit is nut. Seeds in the calyx hole.

(10) Myrrh Leaf Cell Extract, Commiphora Myrrha Leaf Cell Extract, COMMIPHORA MYRRHA

Myrrh is also called myrrha. Pale yellow or dark brown large and small lumpy substance. Myrrh is a rubber resin obtained from olive myrrh or lamiaceae of olive trees.

(11) Arabian Jasmine Leaf Cell Extract, Jasminum Sambac (Jasmine) Leaf Cell Extract, JASMINUM SAMBAC

India is native to China, Taiwan, the Philippines, India, and Indonesia, mainly in the tropics and subtropics. The leaves are opposite or turned one by one, oval or broad oval. The edges are flat and pointed and shiny.

In the present invention, each of the samples may be included in the composition of the present invention in the form of plant cell culture, extract, filtrate and the like.

 In the present invention, 'callus' is an oily tissue in which a cell recovers division ability and prevents and enlarges the wound when the plant is injured. The tissue cut out of the plant is cultured in an auxin-containing medium, It refers to undifferentiated tissue or cell mass generated when wounding certain kinds of plants or treating wound sites with auxin, and can be permanently divided and proliferated by passage in fresh medium. These callus are amorphous cells or masses of cells that have lost their ability to cause normal organogenesis or tissue differentiation, and are mostly flow cells, and broadly include plant tumor tissues caused by infections such as agrobacterium. In other words, the cells in the plant are immediately oriented as they proliferate and are assembled regularly into organs as well as specific tissues. However, callus, an undifferentiated cell mass formed through tissue culture, can be interpreted to proliferate arbitrarily by releasing control that has been maintained until then when various plant growth hormones are stimulated from the outside.

When the callus obtained by dedifferentiation is put into a liquid medium having the same composition and shaken, the cells are separated and continue to proliferate as a suspended state. The callus or cultured cells are essentially different from the differentiated cells in which the whole plant ages and dies by being able to divide and proliferate permanently by passage in fresh medium. Callus or cultured cells, which have been passaged for several generations, are applied to a solid medium from which various plant growth hormones have been removed. This regenerated plant is derived from the division and proliferation of a single cultured cell, and the source of the cultured cell is derived from axons or aquatic tissues.

In the present invention, the "callus" can be any one derived from any part of the plant, but in one embodiment, the petals, calyx, fruit, ovary, part of the throne tissue, or cut cotyledon from germinated seedlings of the seed It may be induced callus.

In the present invention, "callus culture" or "plant cell culture" or "plant cell callus culture" is a culture of callus in the medium. The medium can be used without limitation as long as there is a medium generally used for callus culture in the art.

In the present invention, the skin improvement for skin vitality, skin vitality, skin convergence, wrinkle improvement, skin barrier strengthening or improvement, skin regeneration, moisturizing, maintaining elasticity, skin barrier function protection, improvement, anti-inflammatory, It includes various skin-activating functions such as atopy, antioxidant, anti-aging, pore contraction, sebum control and acne improvement.

The present invention provides a method for preparing a cosmetic composition for skin improvement comprising a plant cell complex culture or extract thereof:

(a) isolating cells from each plant tissue and incubating by inducing callus; And (b) preparing a composition containing the culture obtained from step (a) or an extract thereof.

In the present invention, step (b) may comprise the step of preparing a composition by drying, powdering and mixing each plant cell callus culture obtained in step (a) with purified water.

In the present invention, the step (b) is to dry the respective plant cell callus culture obtained in step (a), powdered and mixed in purified water, and then ultrasonic extraction or hot water extraction to prepare a composition It may include.

In the present invention, the callus culture (culture) may be used, the culture may be used by filtration, the culture may be used by crushing, or the culture may be dried and powdered, and then dissolved in purified water.

In the present invention, "extract" refers to an extract obtained by powdering the extract of the callus culture, or by various conventionally known extraction methods such as cold water extraction, hot water extraction, ethanol extraction.

The extraction method in the present invention is not particularly limited, and for example, cold extraction, ultrasonic extraction, reflux extraction, hot water extraction and the like. Here, in the case of hot water extraction, heated to 80 ~ 100 ℃ for 8 to 48 hours in a boiling water distillation to obtain a hot water extract.

In the case of cold water extraction, for example, cold tissue (15-25 ° C.) is mixed with the pollen tissue culture itself or its dried powder and extracted for 3 days to obtain a cold water extract.

Or by extraction using water, an organic solvent, or a mixed solvent thereof. The extracted solution can be used directly or can be concentrated and / or dried. When extracted with an organic solvent, methanol, ethanol, isopropanol, butanol, ethylene, acetone, hexane, ether, chloroform, ethyl acetate, butyl acetate, dichloromethane, N, N-dimethylformamide (DMF), dimethyl sulfoxide (DMSO), 1,3-butylene glycol, propylene glycol, or a mixed solvent thereof may be used and extracted by room temperature or warming under conditions where the active ingredient of the crude drug is not destroyed or minimized. Depending on the organic solvent to be extracted, the degree of extraction and loss of the active ingredient of the drug may vary, so select an appropriate organic solvent. In particular, in the present invention, an ethanol extract, preferably 20-50% ethanol extract, in one embodiment 50% ethanol extract is preferred, and the extraction method is 3 days after mixing in 50% ethanol, as described in the Examples Obtained by stirring.

In the present invention, the extract may be used by concentration or dilution, and distillate of the extract may be used.

In the present invention, the meaning of "contained as an active ingredient", as a cosmetic composition can exhibit the skin improvement effect of anti-aging, including skin vitality, skin vitality, skin convergence, skin barrier strengthening, improvement, wrinkle improvement For example, as a cosmetic composition, the degree of collagen synthesis, elastin increase, skin convergence, wrinkle improvement, skin barrier strengthening, improvement, pore contraction, sebum control, acne improvement, whitening, anti-inflammatory, etc. It means containing an effective amount of.

In the present invention, the principle of skin astringent action refers to a phenomenon in which the skin contracts by forming crosslinks by binding to polymer flavonoids of the skin protein. Convergence basically means wrinkles or shrinks, and astringents constrict the skin and mucous vessels, and block the cell and lymph gaps to inhibit the secretion of mucus. In addition, since astringents have a property of binding to proteins, the degree of convergence can be determined according to the degree of binding of hemoglobin proteins to extracts. This astringent action can be said to form an insoluble film on the surface of the skin and mucous membranes externally or internally, thereby protecting the topical or densifying tissues, thereby reducing the permeability of the cell membrane. There are two types of astringent action on the skin, chemical agglutination and physical abortion. Chemical agglutination refers to the temporary contraction of sweat glands and pores by protein-coagulating substances such as tannins. Physical astringent, on the other hand, refers to the use of cold water on the skin or to the temporary contraction of sweat glands and pores by lowering the temperature of the skin through the volatilization of ethanol.

In addition, such a skin convergence is caused by the occurrence of more sebum due to stress or in the summer, such as in the summer, the skin troubles occur, and the areas where the skin trouble occurs becomes more and more red, skin soothing means to alleviate such skin troubles .

In the skin barrier protection function according to the present invention, the skin barrier (stratum corneum), which is the outermost layer of the epidermis, is mainly composed of non-nuclear flat corneocytes. Multi lamella lipid layer formed by intercellular lipids such as ceramide, cholesterol, and fatty acid synthesized by keratinocytes of skin barrier maintained through normal epidermal cell division and differentiation process prevents moisture from evaporating inside skin Play a role. On the other hand, of these intercellular lipids, omega hydroxy ceramides are chemically covalently linked to involucrin, a protein in the outer layer of corneocytes, to form a corneocyte lipid envelope (CLE). It acts to physically stabilize the intercellular lipids of the form to strengthen the barrier function.

The composition of the present invention is delivered to the stratum corneum through the skin coating to promote the differentiation of keratinocytes, not only has the effect of thickening the thickness of the epidermal layer, but also has an excellent effect of restoring damage to the skin barrier by damage to the skin barrier. It can be usefully used for the treatment and prevention of induced skin diseases. Skin diseases caused by damage to the skin barrier include atopic dermatitis (atopic dermatitis), dry skin (xeroderma), psoriasis, psoriasis, ichthyosis, acne and the like, but are not limited thereto.

In addition, the skin barrier protection mechanism was confirmed by increasing the amount of protein of occludin and claudin-1, which is a close junction protein. Filaggrin is one of several structural proteins expressed by keratinocytes in the differentiation stage and is involved in the differentiation of the epidermis from the basal layer of the epidermis to the stratum corneum. It is the main agent of the natural moisture factor (NMF), which is essential for the maintenance of skin tissue moisture. It is used as a major indicator of skin moisture retention and skin membrane function. The plant cell complex callus culture according to the present invention has excellent skin barrier protection, strengthening and improving function as the gene expression of filaggrin is greatly increased. May have

In addition, the plant cell complex callus culture or its extract according to the present invention can also be utilized for skin protection by ultraviolet irradiation.

In addition, the plant cell complex callus culture or its extract according to the present invention has an ability to increase elastin expression. Elastin (Elastin) is a structural material present in the extracellular matrix of connective tissue and is present in the skin to impart elasticity. Elastin is synthesized in cells with a polypeptide called Tropo elastin and then produced by cross-linking between tropoelastin extracellularly. Therefore, elastin has a second or three-dimensional elasticity by crosslinking of tropoelastin. Therefore, the plant cell complex callus culture or its extract according to the present invention can be utilized for skin elasticity enhancement and improvement.

Therefore, the cosmetic composition according to the present invention, in particular, has the ability to increase the procollagen, increase collagen biosynthesis, can be applied for preventing skin aging, skin wrinkle improvement / prevention, in addition to enhancing skin elasticity, skin barrier according to the enhancement of elastin expression It can be applied for reinforcement and improvement.

In another aspect, the present invention (a) edelweiss, seaside erringo, rocksam fire, apple, grape, desert rose leaf, lavender leaf, Chinese chestnut leaf, lotus leaf, myrrh leaf, Arabian jasmine each plant tissue Inducing and incubating callus from the; And (b) preparing a plant cell complex callus culture or a skin improvement cosmetic composition containing the extract comprising the step of preparing a composition containing each plant cell callus culture or its extract.

In step (a), specifically, an appropriate medium may be selected for inducing and culturing callus. If there is a medium generally used in the art, it can be used without limitation. In plants, MS medium and B5 medium are generally used.For example, the composition of MS medium (1 L standard) is NH ₄ NO ₃ 1650 mg, KNO ₃ 1900 mg, CaCl ₂ H ₂ O 440 mg, MgSO _{4 .7H 2 O 370 mg, KH} 2 PO 4 170 mg, KI 0.83 mg, H 3 BO 3 6.2 mg, MnSO 4 .4H 2 O 22.3 mg, ZnSO 4 .7H2O 8.6 mg, Na 2 MoO 4 .2H 2 O _{0.25 mg, CuSO 4, 5H 2} O 0.025 mg, CoCl 2 .6H 2 O 0.025 mg, FeSO 4 .7H 2 O 27.8 mg, Na 2 EDTA.2H 2 O 37.3 mg, Myoinositol 100 mg, Nicotinic acid 0.5 mg, Pyridoxine -HCl 0.5 mg, Thiamine-HCl 0.5 mg, Glycine 2 mg, Sucrose 30000 mg.

In the present invention, in the step (b), the composition comprising each of the callus cultures obtained through the culture of step (a) as prepared in a cosmetic composition of the appropriate form, or the culture is known extraction It can be obtained in the form of extract through the method to prepare a cosmetic composition in a suitable form.

For example, in step (b), each callus culture obtained in step (a) is dried, powdered and mixed in purified water to prepare a composition, or

The callus culture obtained in step (a) may be dried, powdered and mixed with purified water, followed by cold water extraction, hot water extraction or ethanol extraction to prepare a composition.

As another example, the culture obtained in step (a) may be prepared by drying the powder after hot air drying and evaporating moisture.

In the cosmetic composition, an acceptable carrier may be included in the cosmetic formulation. Herein, "acceptable carrier in cosmetic preparation" means a compound or composition already known and used that can be included in cosmetic preparations, or a compound or composition which will be developed in the future, which has no toxicity, instability or irritation that can be adapted to the human body upon contact with skin. Say nothing.

The carrier may be included in the cosmetic composition of the present invention in an amount from about 1% to about 99.99% by weight, preferably from about 90% to about 99.99% by weight of the composition. However, since the ratio depends on the formulation as described below in which the cosmetic composition of the present invention is prepared and its specific application site (face, neck, etc.), its preferred application amount, etc., the ratio is in any aspect to the present invention. It should not be understood as limiting the scope of.

Examples of the carrier include alcohols, oils, surfactants, fatty acids, silicone oils, wetting agents, moisturizers, viscosity modifiers, emulsions, stabilizers, ultraviolet scatterers, ultraviolet absorbers, colorants, fragrances, and the like. The alcohols, oils, surfactants, fatty acids, silicone oils, wetting agents, moisturizers, viscosity modifiers, emulsions, stabilizers, UV scattering agents, UV absorbers, colorants, compounds / compositions that can be used as fragrances are already known in the art. As such, those skilled in the art can select and use appropriate materials / compositions.

As one embodiment of the present invention, the cosmetic composition according to the present invention may include glycerin, butylene glycol, propylene glycol, polyoxyethylene hydrogenated castor oil, ethanol, triethanolamine, etc., in addition to the callus extract of the active ingredient, A preservative, a coloring agent, coloring matter, purified water, etc. can contain a trace amount as needed.

In addition, it may further include a conventionally known bioactive component or other plant cell culture or extract. For example, it may further include a plant cell culture or plant cell culture extract of a dodol or scrubber or a dodol / loofah hybrid.

Cosmetic compositions according to the invention can be prepared in various forms, for example, lotions, essences, gels, emulsions, lotions, creams (oil-in-water, water-in-oil, multiphase), solutions, suspensions (anhydrous and aqueous) , Anhydrous products (oil and glycol based), gels, masks, packs, powders, or gelled capsules (soft capsules, hard capsules) formulations and the like.

The skin in the present invention is a concept that includes not only a face but also a scalp and a whole body, and a cosmetic composition that can be applied to such a scalp, such as a shampoo, a rinse, a treatment, a hair regrowth, and a body cleanser that can be applied to the whole body. It can be prepared in various forms as the use of.

The manufacturing method of the cosmetic composition according to the present invention is not limited to the above-described manufacturing method, and those skilled in the art can also be prepared by a method of partially modifying the manufacturing method.

When the cosmetic composition is prepared, cosmetics of the emulsion formulations include nutrient cosmetics, creams, essences, etc., and cosmetics of the solubilized formulations include soft cosmetics. In addition, by containing a dermatologically acceptable medium or base, it can be prepared in the form of topical or systemically applicable adjuvants commonly used in the field of dermatology.

In addition, suitable formulations of cosmetics include, for example, emulsions, suspensions, microemulsions, microcapsules, microgranules or ionic (liposomes), nonionics obtained by dispersing an oil phase in a solution, gel, solid or pasty anhydrous product, aqueous phase, for example. It may be provided in the form of a vesicle dispersant in the form of a cream, skin, lotion, powder, ointment, spray or concealed stick. It may also be prepared in the form of a foam or in the form of an aerosol composition further containing a compressed propellant.

In addition, the cosmetic composition of the present invention may further comprise fatty substances, organic solvents, solubilizers, thickeners and gelling agents, emollients, antioxidants, suspending agents, stabilizers, foaming agents, fragrances, surfactants, water, ionic Or nonionic emulsifiers, fillers, metal ion sequestrants and chelating agents, preservatives, vitamins, blockers, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic active agents, lipid vesicles or any other commonly used in cosmetics It may contain adjuvants conventionally used in the cosmetic or dermatological fields, such as ingredients. In addition, the above components may be introduced in an amount generally used in the dermatology field.

The cosmetic composition containing the plant cell complex culture or extract thereof according to the present invention may provide skin vitality, enhance skin vitality, improve skin wrinkles, regenerate skin, protect skin barrier function, protect skin from ultraviolet light, atopy, anti-inflammatory, antioxidant, As a cosmetic composition having a function such as anti-aging, it can be used as a material of various functional cosmetics.

1 is a result confirming the toxicity of the plant cell complex composition according to the present invention.
Figure 2 is a result confirming the mRNA expression rate of the skin barrier-related genes of the plant cell complex composition according to the present invention.
3 is a result confirming the wrinkle improvement effect of the plant cell complex composition according to the present invention.
Figure 4 is a result confirming the anti-inflammatory effect of the plant cell complex composition according to the present invention.
5 is a result confirming the whitening effect of the plant cell complex composition according to the present invention.
Figure 6 is a result confirming the protective effect from the ultraviolet light of the plant cell complex composition according to the present invention.
Figure 7 is a result confirming the biological expression control related gene expression test results of the plant cell complex composition according to the present invention.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. In general, the nomenclature used herein is well known and commonly used in the art.

Example  1: according to the invention Plant cells Complex  Composition preparation

Example  1-1: Surface disinfection of 11 plant samples

In order to produce the cells according to the present invention, Edelweiss, Seaside Erringo, Roxamfire, Apple, Grape, Desert Rose Leaf, Lavender Leaf, Chinese Chestnut Leaf, Lotus Leaf, Myrrh Leaf, Arabian Jasmine Leaf, and prepared To sterilize, it was immersed in 70% ethanol for 30 seconds, shaken with 2% sodium hypochlorite for 5 minutes, sterilized, and washed with sterile water.

Example  1-2: Plant cell induction from 11 samples

One) Sterilized edelweiss leaves were cut at once with a sharp knife to include plant growth regulators 1 mg / L 6-Benzylaminopurine, 0.3 mg / L 2,4-D, 3% Sucrose and 8 g / L In early MS cells (Murashige and Skoog 1962, Duchefa, Cat No. M0221) were cultured at 25 ° C., 70% humidity and growth conditions to induce early plant cells. Each medium was adjusted to pH 5.8 with 1N sodium hydroxide (NaOH). Subcultures were performed at two week intervals.

2) Sterilized seaside erygo leaves were cut at once with a sharp knife to remove plant growth regulators 50 mg / L indole-3-acetic acid (IAAA), 5 mg / L zeatin, 3 In early MS cells containing% Sucrose and agar 8 g / L (Murashige and Skoog 1962, Duchefa, Cat No. M0221), cancer cells were cultured at 25 ° C and 70% humidity in growth conditions to induce early plant cells. Each medium was adjusted to pH 5.8 with 1N sodium hydroxide (NaOH). Subcultures were performed at two week intervals.

3) Sterilized Roxamfire leaves are cut at once with a sharp knife and plant growth regulators 5 mg / L indole-3-acetic acid (IAA), 10 mg / L zeatin, 3% Sucrose And agar 8 g / L in a basic MS medium (Murashige and Skoog 1962, Duchefa, Cat No. M0221) incubated at 25 ℃, growth conditions of 70% humidity and induced early plant cells. Each medium was adjusted to pH 5.8 with 1N sodium hydroxide (NaOH). Subcultures were performed at two week intervals.

4) Sterilized apple leaves were cut at once with a sharp knife, plant growth regulators 5 mg / L indole-3-acetic acid (IAA), 10 mg / L zeatin, 3% Sucrose and In early MS cells containing 8 g / L of agar (Murashige and Skoog 1962, Duchefa, Cat No. M0221), cancer cells were cultured at 25 ° C and 70% humidity in growth conditions to induce early plant cells. Each medium was adjusted to pH 5.8 with 1N sodium hydroxide (NaOH). Subcultures were performed at two week intervals.

5) Sterilized grape leaves were cut at once with a sharp knife, and the basic MS medium containing 0.1 mg / L 2,4-D, 3% Sucrose and 8 g / L of plant growth regulators (Murashige and Skoog 1962, Duchefa, Cat No. M0221) was incubated at 25 ° C., 70% humidity and growth conditions to induce early plant cells. Each medium was adjusted to pH 5.8 with 1N sodium hydroxide (NaOH). Subcultures were performed at two week intervals.

6) Sterilized desert rose leaves are cut at once with a sharp knife and plant growth regulators 0.5 mg / L IAA (indole-3-acetic acid), 1 mg / L zeatin, 3% Sucrose And agar 8 g / L in a basic MS medium (Murashige and Skoog 1962, Duchefa, Cat No. M0221) incubated at 25 ℃, growth conditions of 70% humidity and induced early plant cells. Each medium was adjusted to pH 5.8 with 1N sodium hydroxide (NaOH). Subcultures were performed at two week intervals.

7) Sterilized lavender leaves were cut with a sharp knife in one step, and the basic MS medium containing plant growth regulators 0.1 mg / L 2,4-D, 3% Sucrose and 8 g / L agar (Murashige and Skoog 1962, Duchefa, Cat No. M0221) was incubated at 25 ° C., 70% humidity and growth conditions to induce early plant cells. Each medium was adjusted to pH 5.8 with 1N sodium hydroxide (NaOH). Subcultures were performed at two week intervals.

8) Sterilized Chinese chest tree leaves are cut at once with a sharp knife and plant growth regulators 0.5 mg / L IAA (indole-3-acetic acid), 1 mg / L zeatin, 3 In early MS cells containing% Sucrose and agar 8 g / L (Murashige and Skoog 1962, Duchefa, Cat No. M0221), cancer cells were cultured at 25 ° C and 70% humidity in growth conditions to induce early plant cells. Each medium was adjusted to pH 5.8 with 1N sodium hydroxide (NaOH). Subcultures were performed at two week intervals.

9) Sterilized lotus leaf was cut at once with a sharp knife to make plant growth regulators 1 mg / L 6-Benzylaminopurine, 0.3 mg / L 2,4-D, 3% Sucrose and agar 8 g / L Early plant cells were induced by cancer culture at 25 ° C., 70% humidity 70% growth in the included 1/2 MS medium (Murashige and Skoog 1962, Duchefa, Cat No. M0221). Each medium was adjusted to pH 5.8 with 1N sodium hydroxide (NaOH). Subcultures were performed at two week intervals.

10) Sterilized myrrh leaves were cut at once with a sharp knife to give plant growth regulators 0.5 mg / L IAA (indole-3-acetic acid), 1 mg / L Zeatine, 3% Sucrose and In early MS cells containing 8 g / L of agar (Murashige and Skoog 1962, Duchefa, Cat No. M0221), cancer cells were cultured at 25 ° C and 70% humidity in growth conditions to induce early plant cells. Each medium was adjusted to pH 5.8 with 1N sodium hydroxide (NaOH). Subcultures were performed at two week intervals.

11) Sterilized arabian jasmine leaves were cut at once with a sharp knife and plant growth regulators 0.5 mg / L IAA (indole-3-acetic acid), 1 mg / L zeatin, 3% Sucrose And agar 8 g / L in a basic MS medium (Murashige and Skoog 1962, Duchefa, Cat No. M0221) incubated at 25 ℃, growth conditions of 70% humidity and induced early plant cells. Each medium was adjusted to pH 5.8 with 1N sodium hydroxide (NaOH). Subcultures were performed at two week intervals.

Example  1-3: According to the invention Plant cells  Culture and Mass Production

Cells of edelweiss, seaside erygogo, rocksam fire, apple, grape, desert rose leaf, lavender leaf, chinese chestnut leaf, lotus leaf, myrrh leaf, arabian jasmine leaf, aseptically cultured Using a bioreactor, the liquid medium of the same composition except for agar was controlled at a temperature of 25 ° C., a humidity of 70%, and an air supply of 0.1 vvm.

Example  1-4: according to the invention Plant cells  Complex extract manufacturer

Eleven plant cells obtained by cultivation were harvested, dried with clean tissue, and dried at 60 ° C. for 2 days. In order to obtain the composition according to the present invention Edelweiss plant cells 40 g, seaside erygogo callus 20 g, Rocksam fire callus 20 g, apple callus 20 g, grape callus 40 g, desert rose leaf callus 40 g, lavender leaf callus 40 g, Chinese chestnut leaf callus 20g, lotus leaf callus 40g, myrrh leaf callus 20g and Arabian jasmine leaf callus 20g was thoroughly washed with purified water and mixed. Hot water extract of callus complex was obtained by heating to 80-100 ° C. for 8 to 48 hours using 20 L of purified water in a boiling water distillation machine. The obtained composition was centrifuged at 12000 rpm for 4 ° C. for 30 minutes to remove insoluble solids, and the supernatant was obtained as a callus complex extract.

Experimental Example 1: Influence test on cell viability

In this experiment, the proliferation activity of the cells of the plant cell complex extract prepared in Example 1 was examined.

To this end, first, HaCaT (human keratinocyte, keratinocyte) and Detroit551 (human fibroblast, fibroblast) were dispensed in 96 well plate with DMEM medium containing 10% FBS and in a thermostat at 5% CO ₂ , 37 ℃. Incubated for 24 hours. After 24 hours, the cells were treated to the final concentration of 0.5, 1, 2% of the plant cell complex extract, and further incubated for 24 hours. Then 5 mg / mL MTT solution was added to each well 4 μL and incubated for 4 hours. After the culture was removed, the cells were lysed by adding 100 μL of DMSO (Dimethyl sulfoxide) solution, and then the absorbance was measured at 540 nm using a spectrophotometer.

The survival rate of the skin cells was calculated according to the following equation based on the absorbance intensity of the control group using purified water and expressed as a percentage, and the results were shown in FIG. 1.

Equation 1] cell survival rate (%) = [(experimental absorbance-control absorbance) / control absorbance] × 100

As shown in Figure 1, it was confirmed that the plant cell complex extract of the present invention does not show toxicity in three kinds of skin cell lines in the treatment concentration.

Experimental Example  2: antioxidant effect ( ABTS  Radical Scavenging Activity

Antioxidant activity using ABTS radical was measured by removing the ABTS free radicals produced by the reaction with potassium persulfate by the antioxidants in the extract and decolorizing the blue color of the index. Measured.

1.0 mM AAPH (2,2'-azo-bis 2-amidinopropanedeihydrochloride) was mixed with 2.5 mM ABTS (2,2'-azino-bis 3-ethylbenzenothiazolin-6-sulfonic acid) dissolved in 100 mM PBS buffer. The reaction was carried out for 12 minutes in a thermostat bath. 20 μl of plant cell complex extract and 980 μl ABTS solution were reacted for 10 min in a 37 ° C. water bath to obtain a final concentration of 0.5, 1, 2, or 5%, and the degree of absorbance at 734 nm was measured. % Was used. ABTS radical scavenging ability was calculated according to the following equation.

[Equation]

Processing material ABTS  Radical Scavenging power  ( % ) Control 0 Trolox 0.1% 52.67 Plant Cell Complex Extract 0.5% 31.33 Plant Cell Complex Extract 1% 34.67 Plant Cell Complex Extract 2% 47.33

As a result, as shown in the table, it was confirmed that the plant cell complex extract of the present invention has an antioxidant effect according to the scavenging ability of the ABTS radicals in the treatment concentration.

Experimental Example  3: skin barrier improvement gene expression test

In this experiment, in order to examine the skin barrier improvement effect of the plant cell complex extract prepared in Example 1, the expression change of the AQP3, a water / glycerol transporter and FLG known as a natural moisturizing factor was investigated.

HaCaT cells were dispensed into 96 well plates, and then cultured for 24 hours under cell culture conditions. Thereafter, the plant cell complex extract, a test substance, was treated with the cells to a final concentration of 1% with DMEM medium containing no serum, and further cultured for 24 hours. Real-time PCR was performed to confirm the gene levels of AQP3 and FLG, and the test sequence was as follows.

SuperPrep ^™ cell lysis & RT Kit for qPCR (TOYOBO, Cat. SCQ-101) was used for RNA isolation and cDNA synthesis. The cells from which the medium was removed were washed once with PBS, 50 μL cell lysis mixture (including a gDNA remover) was added and reacted for 5 minutes, and then a stop solution was added. 8 μL of the extracted mRNA was added to 32 μL of the RT reaction mixture, and cDNA was synthesized at 37 ° C. for 15 minutes, 50 ° C. for 5 minutes, and 95 ° C. for 5 minutes by PCR. To compare the expression of genes, the synthesized cDNA was used as a template and real-time PCR analysis was performed using Thunderbird ^TM SYBR qPCR Mix (TOYOBO, Cat. QPS-201). The primers used in the experiment were QuantiTect® primer assays (GAPDH; Cat. QT01192646, AQP3; Cat. QT00212996, FLG; Cat. QT02448138) from Qiagen. Real-time qPCR conditions were first subjected to a total of 40 cycles of 95 ℃ 15 minutes, then 94 ℃ 15 seconds, 60 ℃ 30 seconds, 72 ℃ 30 seconds per cycle.

As a result, as shown in Figure 2, in the cells treated with the plant cell complex extract 1% of the present invention, it was confirmed that the mRNA expression rate of AQP3 and FLG is increased, thereby improving the skin barrier.

Experimental Example  4: skin regeneration and wrinkle improvement effect test

In this experiment, in order to examine the skin regeneration and wrinkle improvement effect of the plant cell complex extract prepared in Example 1, collagen decomposition-related enzyme MMP-1 (Matrix Metalloproteinase-1) and procollagen precursor of the synthesis of collagen Biosynthesis amount of (Procollagen Type I C-Peptide, PIP) was measured.

<4-1> MMP-1 biosynthesis measurement

Detroit551 cells were dispensed into 48 well plates and incubated for 24 hours under cell culture conditions. Thereafter, the plant cell complex extract as a test substance was treated with the cells to a final concentration of 1% with DMEM medium containing no serum, and further cultured for 48 hours.

Take 100 μL of the diluted supernatant of the culture medium through the above procedure and place them in antibody coated microtiter plates with 100 μL of MMP-1 standard in MMP-1 Biotrak Activity Assay Kit (Amersham, Cat. RPN2629). The reaction was carried out for 3 hours at ℃. After the medium was removed and washed four times with 200 μL of PBS, 100 μL of antiserum was added to each well and reacted at 25 ° C. for 3 hours. Thereafter, the solution was removed, washed four times with 200 μL of PBS, and then 100 μL of peroxidase-labeled antibody solution was added to each well, and further reacted at 25 ° C. for 1 hour. After removing the solution, 100 μL of substrate solution was added to each well, and the reaction was performed at room temperature for 15 minutes in a shaded state. Thereafter, 100 μL stop solution (1 NH ₂ SO ₄ ) was added thereto and the absorbance at 450 nm was measured. After measuring the absorbance at 450 nm, a standard concentration curve of MMP-1 was prepared to determine the amount of MMP-1 in the sample.

<4-2> PIP biosynthesis measurement

Detroit551 cells were dispensed into 48 well plates and incubated for 24 hours under cell culture conditions. Thereafter, the plant cell complex extract as a test substance was treated with the cells to a final concentration of 1% with DMEM medium containing no serum, and further cultured for 48 hours.

Take 20 μL of the diluted supernatant of the culture medium, and add 20 μL of PIP standard in PIP EIA Kit (Takara, Cat. MK101) to the antibody coated microtiter plate, respectively, and 100 μL of peroxidase-labeled antibody solution. Also added to each well, and reacted for 3 hours at 37 ℃. After the medium was removed and washed four times with 200 μL of PBS, 100 μL of substrate solution was added to each well and reacted at room temperature for 15 minutes in a shaded state. Then 100 μL stop solution (1 NH ₂ SO ₄ ) was added and the absorbance at 450 nm was measured. After measuring the absorbance at 450 nm, the PIP standard concentration curve was prepared to determine the amount of PIP in the sample.

As a result, as shown in Figure 3, in the cells treated with the plant cell complex extract 1% of the present invention, it was confirmed that the amount of MMP-1 is reduced, the amount of PIP is increased, which is effective in anti-wrinkle.

Experimental Example  5: anti-inflammatory effect

The amount of nitric oxide (NO) produced from the Raw 264.7 cell line was measured using Griess reagent as the NO ₂ ⁻ form present in cell culture. To measure the inhibition of NO production in Raw 264.7 cells activated with 1 μg LPS, the experimental and control groups were treated with medium containing the plant cell complex extract at the final concentration of 1.0% for 24 hours, followed by 24 hours of cell culture, followed by Griess reagent cell culture supernatant. 100 μl and 100 μl of Griess reagent (1% sulfanilamide in 5% phosphoric acid, 1% α-naphthylamide in H ₂ O) were mixed for 10 minutes in 96 well plates, and the absorbance was measured at 540 nm. The concentration of NO ₂ ^- was diluted with sodium nitrate to measure the absorbance to obtain a standard curve.

As a result, as shown in Figure 4, the cells treated with the plant cell complex extract 1% of the present invention was confirmed that the anti-inflammatory effect by reducing the NO content in the inflammatory response induced by LPS.

Experimental Example  6: inhibit melanin production Whitening effect  effect

B16F1 cells were put in 2 mL of 1 × 10 ⁵ cells / well in 6 well plates with DMEM medium containing 10% FBS, and incubated in a thermostat at 5% CO 2, 37 ° C. for 24 hours. After removing the medium, each well was treated with 2 ml of plant cell complex extract containing a final concentration of 1%. After treating α-MSH dissolved in DMEM medium to the final 10 nM, the treated cells were reacted for 72 hours in a thermostat at 5% CO2, 37 ℃. After removing the culture medium and washing three times with PBS (phosphate buffer saline), 100 μL of 1X RIPA buffer containing protease inhibitor was lysed and the cells were lysed and centrifuged at 15,000 rpm for 20 minutes. Was recovered. The recovered supernatant is added to a 96 well plate in 50 μL.

For quantification of protein, 150 μL of BCA solution (A: B = 50: 1) was added to each well, and then reacted for 30 minutes in an incubator at 37 ° C., followed by measurement of absorbance at 540 nm using an ELISA reader. It was. The amount of protein was determined by substituting the measured absorbance values into a standard calibration curve using albumin in the concentration range of 0-1 mg / mL.

Finally, for quantitation of melanin, Pellet was dried at 60 ° C. and 400 μL of 1N NaOH containing 10% DMSO was added and reacted in a 90 ° C. thermostat. 100 μL of the reacted solution was placed in a 96 well plate, and the absorbance was measured at 492 nm with an ELISA reader. The measured absorbance was substituted into the melanin standard calibration curve in the concentration range of 0-2.5 mg / mL to determine the amount of melanin.

As a result, as shown in FIG. 5, the cells treated with the plant cell complex extract 1% of the present invention was confirmed to have a whitening effect by inhibiting melanin production.

Experimental Example  7: Cytoprotective effect by UV irradiation

As a light source for UV irradiation, an ultraviolet crosslinker CL-1000 (Ultra-Violet Products, CA) that emits ultraviolet light having a wavelength of 302 nm was used. HaCaT cells, which are keratinocytes, were placed in a 24-well plate at a concentration of 1x10 ⁵ cells / ml, incubated for 24 hours in an incubator, and then the medium was removed and washed with phosphate buffer. 500 μL of phosphate buffer solution was added, and then irradiated with UV light at 10 mJ / cm², and then replaced with fresh cell culture medium without FBS. The plant cell complex extract was treated with a final concentration of 1% and further cultured for 24 hours. To this, 5 mg / ml of MTT (3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide, Sigma) was added, and after 3 hours of incubation in an incubator, the formed fomazan was dissolved in DMSO, After transferring to a 96well plate, the absorbance was measured with an ELISA reader at 595 nm, and the cell survival rate was compared with the control group without UV irradiation.

As a result, as shown in FIG. 6, it was confirmed that the cell survival rate decreased after UVB-induced cell treatment was increased by 1% treatment of the plant cell complex extract of the present invention.

Experimental Example  8: Gene expression test related to biological clock control

In this experiment, in order to examine the biological clock control effect of the plant cell complex extract prepared in Example 1, the expression changes of the related genes CLOCK and PER1 were investigated.

HaCaT cells were dispensed into 96 well plates, and then cultured for 24 hours under cell culture conditions. Subsequently, after induction of UVB, the test cell plant complex extract as a test substance with DMEM medium containing no serum was treated to cells to a final concentration of 1% and further incubated for 8, 16, 24 and 32 hours. Real-time PCR was performed to check gene levels of CLOCK and PER1. The test sequence is as follows.

SuperPrep ^™ cell lysis & RT Kit for qPCR (TOYOBO, Cat. SCQ-101) was used for RNA isolation and cDNA synthesis. The cells from which the medium was removed were washed once with PBS, 50 μL cell lysis mixture (including a gDNA remover) was added and reacted for 5 minutes, and then a stop solution was added. 8 μL of the extracted mRNA was added to 32 μL of the RT reaction mixture, and cDNA was synthesized under PCR conditions at 37 ° C. 15 minutes, 50 ° C. 5 minutes, and 95 ° C. 5 minutes. To compare the expression of genes, cDNA synthesized above was used as a template and real-time PCR analysis was performed using Thunderbird ^TM SYBR qPCR Mix (TOYOBO, Cat. QPS-201). The primers used in the experiment were QuantiTect® primer assays (GAPDH; Cat. QT01192646, CLOCK; Cat. QT00054481, PER1; Cat. QT00069265) from Qiagen. Real-time qPCR conditions were first subjected to a total of 40 cycles of 95 ℃ 15 minutes reaction, then 94 ℃ 15 seconds, 60 ℃ 30 seconds, 72 ℃ 30 seconds per cycle.

As a result, as shown in Figure 7, the expression pattern of irregular CLOCK and PER1 by UVB stimulation in the cells treated with the plant cell complex extract 1% of the present invention, the expression of CLOCK, PER1 of the cells not UVB stimulation It was confirmed that the recovery was close.

Experimental Example  9: skin energy activity enhancing effect test

In this experiment, in order to examine the skin energy activity enhancement effect of the plant cell complex extract prepared in Example 1, the activity of the energy (ATP) of the cells was measured.

Detroit551 cells were dispensed into 12 well plates and incubated for 24 hours under cell culture conditions. Thereafter, the plant cell complex extract, which is a test substance, together with the serum-free DMEM medium was treated to the final concentration of 0.5, 1, 2%, and further incubated for 24 hours.

50 μL of the diluted lysate of the cultured cells was taken, and 50 μL of ATP standard in ATP assay Kit (Abcam, Cat. Ab83355) was added to each 96 well plate and 50 μL of ATP reaction mix solution was added. In addition, it was added to each well and allowed to react at room temperature for 30 minutes in a shaded state. Then, after measuring the absorbance at 570 nm, the ATP standard concentration curve was prepared to calculate the amount of ATP in the sample. The degree of skin energy activity was calculated according to the following equation based on the absorbance intensity of the control group using purified water and expressed as a percentage.

Processing material Energy active effect (%) Control 0 Plant Cell Complex Extract 1% 44.77

As a result, as shown in the above table, it was confirmed that the cells treated with the plant cell complex extract 1% of the present invention had an excellent energy activity effect.

Experimental Example  10: skin vitality test

In this experiment, to examine the skin vitality-promoting effect of the plant cell complex extract prepared in Example 1, the test substance was applied to 20 20-45 year old female subjects having a BMI (Body Mass Index) of 21-27. After the shower was used once a day for a week and massage was used, and the effect was compared before and after 12 weeks of use. The evaluation items are shown in Table 3 below the number of subjects who responded that they felt the effect on skin cell activation and skin vitality by performing a survey.

Test substance Skin cell activation Skin vitality Control 2 0 Plant Cell Complex Extract 0.5% 6 5 Plant Cell Complex Extract 1% 11 13 Plant Cell Complex Extract 2% 13 17

As a result, as shown in the above table, the plant cell complex extract of the present invention hardly felt the effect of skin cell activation and skin vitality after 12 weeks in the control group when most of the respondents were used in the evaluation of feeling in the treatment concentration. It was confirmed that the plant cell complex extract responded that the skin cell activation and skin vitality were felt.

Experimental Example  11: acne improvement

10 people of acne developed men and women was applied to apply the 1% concentration of plant cell complex extract prepared in Example 1 to the entire face for 4 weeks in the morning and evening. Determination of the acne improvement effect is to evaluate the pre-test state to 1-3 points by reference to the following criteria according to the degree of the person who participated in the experiment, and then to evaluate the acne healing effect is shown in the table.

<Evaluation before the test>

1 = some acne 2 = some acne 3 = severe acne

<Acne effect judgment>

-: Almost no effect, +: Some effect, ++: Much eased,

+++: completely healed

Subject Pre-test status After 2 weeks After 4 weeks Subject 1 One - - Subject 2 3 + ++ Subject 3 One - + Subject 4 2 + ++ Subject 5 One + + Subject 6 2 + + Subject 7 2 + ++ Subject 8 2 - ++ Subject 9 One - + Subject 10 3 ++ +++

As a result, as shown in the table, after using the flexible lotion containing 1% of the plant cell complex extract of the present invention for 4 weeks, the majority of subjects could confirm the improvement of acne.

Experimental Example  12: sebum inhibitory effect experiment

In order to confirm the sebum inhibitory effect of the plant cell complex extract, the sebum secretion of the skin was measured after 4 weeks of 10 subjects using a skin meter (Sebum meter SM810). The experiment was conducted at 22 ± 2 ° C. and 40 ± 5% humidity. When the oily and the measured value is more than 220 ㎍ / cm ^2, when the normal skin is 100~220 ㎍ / cm ^2. Purified water was used as a control.

Processing material 0 days (before use) 28 days (after use) Control 232 225 Plant Cell Complex Extract 0.5% 231 220 Plant Cell Complex Extract 1% 230 191 Plant Cell Complex Extract 2% 227 180

As a result, as shown in the above table, the plant cell complex extract of the present invention was confirmed that the sebum secretion decrease after 4 weeks in the treatment concentration.

Experimental Example  13: Pore Shrinkage Effect Test (In vitro Test)

In order to measure the pore contraction effect of the plant cell complex extract composition of the present invention, the pore contraction effect was tested using hemoglobin as a protein to replace the skin of the composition prepared in the above example. Prepare a hemoglobin solution (0.05g / 50ml) with 0.9% Phosphate Buffer Saline (0.1mM, pH 7.4), add 2 ml of edelweiss plant cells and mycorrhizal cell culture extract to 2 ml of hemoglobin solution, and shake for 30 seconds. After centrifugation at 3,500 rpm for 10 minutes, 2 ml of purified water was added to 1 ml of the supernatant, which was measured by UV-Visible spectrum at 407 nm. 2 ml of PBS (Phosphate Buffer Saline) was added as a control.

Processing material Pore shrinkage effect ( % ) Control 0 Plant Cell Complex Extract 0.5% 28.78 Plant Cell Complex Extract 1% 35.74 Plant Cell Complex Extract 2% 39.15

As a result, as shown in the table, it was confirmed that the plant cell complex extract of the present invention has a pore contraction effect in the treatment concentration.

Claims (10)

Edelweiss callus culture or its extract, Seaside Eringo callus culture or its extract, Loxampire callus culture or its extract, Apple cell culture or its extract, Grape cell culture or its extract, Desert rose leaf cell culture Or any one or more of extracts, lavender leaf cell cultures or extracts, Chinese yeast tree leaf cell cultures or extracts, lotus leaf cell cultures or extracts, myrrh leaf cell cultures or extracts, Arabian jasmine leaf cell cultures or extracts Cosmetic composition for skin improvement containing as an active ingredient. The cosmetic composition according to claim 1, wherein the composition is for preventing or improving wrinkles. According to claim 1, wherein the composition is a cosmetic composition, characterized in that for protecting or enhancing the skin barrier function. According to claim 1, wherein the composition is a cosmetic composition, characterized in that for improving or preventing acne. According to claim 1, wherein the composition is a cosmetic composition, characterized in that for protecting the skin by ultraviolet radiation. The cosmetic composition according to claim 1, wherein the composition is for skin whitening. According to claim 1, wherein the composition is a cosmetic composition, characterized in that for anti-aging or antioxidant. The cosmetic composition according to claim 1, wherein the composition is for sebum suppression. The cosmetic composition according to claim 1, wherein the composition is for pore contraction. Edelweiss callus culture or its extract, Seaside Eringo callus culture or its extract, Loxamfire callus culture, or extract, Apple cell culture or its extract, Grape cell culture or its extract, comprising the following steps: Desert Rose Leaf Cell Culture or Extract, Lavender Leaf Cell Culture or Extract, Chinese Chaste Tree Leaf Cell Culture or Extract, Lotus Leaf Cell Culture or Extract, Myrrh Leaf Cell Culture or Extract, Arabian Jasmine Leaf Cell Culture Or a method for producing a cosmetic composition according to any one of claims 1 to 9 containing any one or more of the extracts:
(a) inducing and cultivating callus from the respective plant tissues of edelweiss, seaside erringo, rocksam fire, apple, grape, desert rose leaf, lavender leaf, chinese chestnut leaf, lotus leaf, myrrh leaf, arabian jasmine step; And (b) preparing a composition containing each of said plant cell callus cultures or extracts thereof.

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