KR101533215B1 - Anti-aging and Anti-inflammation and Anti-oxidation cosmetic composition including Pulsatilla koreana Plant Placenta Cell Cultures Extracts - Google Patents

Abstract

[0001] The present invention relates to a cosmetic composition containing a tuberculous cell culture or an extract thereof of a plant of the family Ranunculaceae, and more particularly to a cosmetic composition for skin improvement comprising an extract of a tuberculosis cell .
The present invention relates to a cosmetic composition comprising a plant extract of Smyrna plant cell or an extract thereof, which is excellent in the ability to synthesize skin collagen without toxicity to skin cells, and has a pore reduction, whitening, suppression of sebum secretion, moisturizing, anti-inflammation and acne improvement.

Description

(Anti-aging and Anti-inflammation and Anti-oxidation cosmetic composition including Pulsatilla koreana Plant Placenta Cell Cultures Extracts)

[0001] The present invention relates to a cosmetic composition containing a tuberculous cell culture or an extract thereof of a plant of the family Ranunculaceae, and more particularly to a cosmetic composition for skin improvement comprising an extract of a tuberculosis cell .

The skin protects all the organs in the body from changes in temperature and humidity and from external stimuli such as ultraviolet rays. However, constant physical and chemical stimuli received from the outside deteriorate the skin function and accelerate aging of the skin. Such aging of the skin accompanies the deterioration of the skin constituents as a whole, resulting in the skin being easily split.

In addition, a decrease in fibroblasts and impaired activity of these cells are also an important part of the skin aging process. In order to prevent such skin aging phenomenon and to maintain healthier and more beautiful skin, cosmetics containing physiologically active substances obtained from various plants, animals and microorganisms have been conventionally used to maintain the inherent functions of the skin, We have been striving to control aging effectively. However, existing cosmetic raw materials are still insufficient to satisfy satisfactory effects or efficacies, and have a problem of causing skin side effects.

In addition, inflammation is one of the defense mechanisms that try to minimize or minimize the reaction when cells or tissues are damaged by causes such as stress or ultraviolet rays, causing cell reactions with nerves, blood vessels, tissues, blood, etc. Overexposure can result in itching, fever, redness, pain, skin irritation such as tissue damage, and skin trouble.

Such skin inflammation and skin troubles can regulate stimulation and inflammation by regulating cytokines and inhibiting the expression of inflammatory mediators to normalize skin.

Removal of inflammation to eliminate inflammation, the action of reducing vital signs and symptoms is called anti-inflammatory. To date, substances used for antiinflammatory purposes are nonsteroidal system such as flufenamicacid, ibuprofen, benzydamine, indomethacin, and steroids such as prednisolone ), Dexamethasone, etc. In addition, neuropeptide antagonists, bradykinin antagonists, and COX inhibitors have been proposed. However, most of them have problems in terms of safety to skin and stability in cosmetic preparation, (Patent Publication No. 10-2008-0020853).

On the other hand, acne (Acne vulgaris) is a skin disease caused by the narrowing or clogging of the pores and the loss of sebum. Hair follicles contain sebaceous glands that secrete oil and moisturize the hair. If the sebaceous gland secretion increases or the entrance of the hair follicle becomes clogged, the proliferation of acne bacterium becomes active and causes acne. There are many causes of acne, but the proliferation of the male hormones androgen, acne causing bacteria, Propionibacterium acnes and the increase of sebum secretion are the most important. In patients with acne, androgens stimulate the sebaceous glands to increase sebum secretion, and these increased sebum tangle with dead skin cells and block the fascial entrance. This clogged follicle entrance provides an environment in which the anaerobic bacterium, Propionibacterium acnes, can grow, and the lipases secreted by these bacteria degrade the sebum, which causes the hair follicles to irritate the hair follicle, eventually causing inflammation. Prevention and treatment of acne can be achieved through excessive suppression of sebum secretion and inflammation relief. Therefore, if the cause is removed in a complex manner, the acne can be effectively inhibited. Particularly, acne has been increasing in recent years due to external stress, air pollutants, hormone imbalance, and so on. This phenomenon is becoming more prominent every year, and many methods related to acne have been suggested. A method for preventing and treating acne by applying salicylic acid, triclosan and trihydroxy palmiic acid to a cosmetic composition as a method for improving skin troubles associated with acne by opening pores and promoting sebum secretion has been disclosed (Korean Patent Laid-Open Publication No. 2004-0089072) discloses a method of suppressing sebum secretion by reducing sebum secretion amount in sebaceous glands by taking oligosaccharide (JP-A-05-294836). In addition, a method for preventing acne by applying a violet extract having antibacterial activity to Propionibacterium acnes has been proposed (Korean Patent Publication No. 2002-0082616)

On the other hand, there has never been a functional study on a plant of the family Ranunculaceae as a material of the above-mentioned cosmetic composition.

Korean Patent Publication No. 2004-0089072 Japanese Patent Application Laid-Open No. 05-294836 Korean Patent Publication No. 2002-0082616

The present inventors tried to prepare a cosmetic composition having a skin-improving ability. The inventors of the present invention have succeeded in separating the embryonic stem cells from the seedlings of Ranunculaceae and culturing them through plant tissue culture, The present invention has been accomplished by confirming that there is the skin moisturizing, whitening, anti-inflammation, sebum suppression, acne improvement and wrinkle-reducing effects, and the like.

Accordingly, it is an object of the present invention to provide a cosmetic composition for skin improvement containing as an active ingredient a cultured tubercle cell of a plant of the family Ranunculaceae or an extract thereof.

It is still another object of the present invention to provide a method for producing a cosmetic composition comprising the above-mentioned Ranunculaceae cell culture or an extract thereof as an active ingredient.

In order to achieve the above object, the present invention provides a cosmetic composition for improving skin comprising Ranunculales plant tubercle cell culture or its extract as an active ingredient.

In the present invention, the buttercups are selected from the group consisting of Paeonia, Nelumbo, Nymphaea, Berberidaceae, Circaeasteraceae, Kingdoniaceae, (Eupteleaceae, Lardizabalaceae, Menispermaceae, Papaveraceae, Fumariaceae), Pteridophyllaceae, or Ranunculaceae plants, which are plant parts of the plant, And the like.

In the present invention, the buttercups of the buttercups include Anemone, Anemone coronaria, Poeny, Paeonia lactiflora, Hepatica asiatica Nakai, Pulsatilla koreana, Lotus, Nelumbo nucifera, (Christmas rose, Helleborus niger), Paeonia suffruticosa, and Nymphaea teragona.

In the present invention, the cosmetic composition may be used for wrinkle prevention or improvement, anti-inflammation, pore reduction, sebum secretion suppression, acne improvement, whitening, antibacterial or skin moisturizing.

The present invention also relates to a method for producing a plant cell, comprising the steps of: (a) And (b) a step of preparing a culture containing the culture obtained in the step (a) or an extract thereof, and a method for producing a cosmetic composition for skin improvement comprising the extract to provide.

The present invention relates to a cosmetic composition containing an extract of a tubercle plant of the family Ranunculaceae according to the present invention, which is excellent in the ability to produce skin collagen without toxicity to the skin cells, and has excellent skin moisturizing, anti-inflammatory, anti- Lt; / RTI >

TECHNICAL FIELD The present invention relates to a cosmetic composition for skin improvement containing a culture of tubercle cells of the plant Aspergillus or extract thereof as an active ingredient and a process for producing the same.

In one aspect, the present invention relates to a cosmetic composition for improving skin comprising as an active ingredient a cultured teacquard cell culture or a extract thereof. Specifically, the tubercle cell culture or extract thereof of the buttercups plant may be contained in an amount of 0.1 to 10% by weight based on the total weight of the composition. For example, in the following examples, the case of 2% by weight was experimented. However, in the case of 10% by weight or more, 20% or 30% or more, It is obvious to a person skilled in the art that the skin improvement effect, including acne improvement, sebum suppression, anti-inflammation, wrinkle improving function.

In the present invention, the Ranunculales plant is a plant according to the classification system belonging to the dicotyledonous plant river, and includes the following plants: Paeoniaceae, Nelumbonaceae, Nymphaeaceae, Berberidaceae, Circaeasteraceae, Kingdoniaceae, Eupteleaceae, Lardizabalaceae, Menispermaceae, Papaveraceae, and so on. Papaveraceae, Paleo and Fumariaceae), Pteridophyllaceae, or Ranunculaceae plants.

In the present invention, the buttercups may be selected from the group consisting of Anemone, Paeonia, Hepatica, Pulsatilla, Nelumbo, Helleborus, And may include Nymphaea plants.

In one embodiment of the present invention, an anemone (Anemone, Anemone coronaria), Poeny (Paeonia lactiflora), Hepatica asiatica Nakai, Pulsatilla koreana, lotus (Lotus, Nelumbo nucifera, Christmas rose (Helleborus niger), Paeonia suffruticosa, and Nymphaea teragona. The present invention has been accomplished on the basis of these findings.

In the present invention, it is preferable to use the above-mentioned cultured tubercle cell culture (cultured product) itself, or to use the culture by filtration, or to crush the culture, or to dry the culture to obtain powder, It can be melted and used.

In the present invention, "extract" means an extract obtained by various known extraction methods such as pulverization of the tubercle cell culture extract of buttercups, or a cold water extraction method, a hot water extraction method and an ethanol extraction method.

The extraction method in the present invention is not particularly limited, and examples thereof include cold extraction, ultrasonic extraction, reflux extraction, hot water extraction and the like. Here, in the case of hot water extraction, the hot water extract is obtained by heating at 80 to 100 ° C for 8 to 48 hours in a hot water distiller.

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

Alternatively, it can be produced by a method of extracting using water, an organic solvent, or a mixed solvent thereof. The extracted liquid can be used directly or by concentrating and / or drying. When extraction is carried out using an organic solvent, an organic solvent such as methanol, ethanol, isopropanol, butanol, ethylene, acetone, hexane, ether, chloroform, ethyl acetate, butyl acetate, dichloromethane, N, (DMSO), 1,3-butylene glycol, propylene glycol, or a mixed solvent thereof. The active ingredient of the herbal medicine may be extracted at room temperature or warmed under the condition that the active ingredient is not destroyed or minimized. Depending on the organic solvent to be extracted, the degree of extraction and the degree of loss of the active ingredient of the medicament may differ. Therefore, an appropriate organic solvent should be selected and used. Particularly, in the present invention, an ethanol extract, preferably a 20-50% ethanol extract, specifically 50% ethanol extract, is preferable, and an extraction method is obtained by mixing with 50% ethanol and stirring for 3 days.

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

In the present invention, facilitation of skin improvement refers to the improvement of skin protection and skin condition, skin whitening, prevention or improvement of skin aging and wrinkles, skin moisturization, suppression of sebum secretion, improvement of acne, reduction of pores, skin protection, It is a concept that includes mitigation, improvement of immunity disease, improvement of skin barrier function, alleviation of skin irritation, skin cell proliferation and regeneration ability, antioxidant ability and collagen synthesis promoting ability.

In the present invention, the expression "comprising as an active ingredient" means a cosmetic composition which is capable of exhibiting skin improving effect, for example, antioxidant, pore reduction, skin moisturizing, sebum suppression, acne improvement, Quot; means an amount of effective amount that is capable of exhibiting collagen synthesis or iNOS inhibitory activity, and the like.

Accordingly, the cosmetic composition according to the present invention can be used for prevention of skin aging, improvement and prevention of skin wrinkles, and can be applied for increasing antioxidant, whitening, pore contraction, sebum suppression, antibacterial , anti-inflammation through iNOS inhibitory activity, skin inflammation prevention, alleviation, alleviation, and treatment.

The efficacy of such a tubercle cell culture extract extract of Ranunculaceae plants can be evaluated by comparing the extracts of flower buds, leaves, stem, root tissue-derived cell cultures, It has superior efficacy such as anti-inflammation, antioxidation, moisturizing, anti-wrinkle, whitening, pore contraction, sebum suppression, acne improvement, antibacterial, and polyphenol content and flavonoid content.

Plant tissue culture is carried out on small live plant tissue ( in in vitro ) and cultivating plant cells, organs and plants according to the purpose of culture. Plant tissue culture can be attributed to the basic characteristics of plants. Plant tissue culture technology is based on the use of differentiating totipotency to regenerate plants from plant somatic cells, a unique feature of plants. Namely, plants can be regenerated from these cells and tissues by culturing single cells (including protoplasts), leaves and root sections of plants in a medium supplemented with specific nutrients and growth regulators. The ability of these plants to have differentiating totipotency is a strategy for survival from harsh environments and herbivorous damage for the fixation and long-term survival of plants. Therefore, plants can be said to have the ability to regenerate lost organs again. The plant tissue cells isolated from the parent plant have the potential to be reproduced as a complete plant when cultured in a suitable culture environment, that is, plants having typical performance are of academic importance such as embryology, but they are also utilized for practical use of cosmetic raw materials .

The placenta of the plant is an important part that regulates seed breeding, such as an animal-derived placenta. It is a part of the ovary in the pistil (ovary, ovary). It is usually on the edge of the carpel, which forms the ovary. It also occurs in the base of the ovary, or in the shape of a column standing upright in the ovary from the base.

In another aspect, the present invention relates to a method for producing a transformed plant, comprising: (a) And (b) a step of preparing a composition containing the tubercle cell culture or an extract thereof of the buttercup garlic plant, or a method for producing a cosmetic composition for skin improvement comprising the extract of the tubercle cell of the bitter gourd plant .

In the step (a), specifically, a suitable medium may be selected for culturing the seedlings of the seedlings. If there is a medium commonly used in plant tissue culture in the technical field, it can be used without limitation. For example, the composition of the MS medium (1 L standard) is 1650 mg NH ₄ NO ₃ , 1900 mg KNO ₃ , 440 mg CaCl ₂ .2H ₂ O, MgSO 4, _{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, and Sucrose 30000 mg.

In the present invention, it is preferable that, in the step (b), a composition comprising the crude aspartame cell culture obtained as a result of the cultivation in step (a) is prepared as a cosmetic composition of a proper form, Can be obtained in the form of an extract through a known extraction method and can be prepared into a cosmetic composition in a suitable form.

For example, in the step (b), the cultured tubercle cell culture of Ranunculaceae plant obtained in step (a) is dried, powdered and mixed with purified water to prepare a composition,

the step of drying the pulverized tubercle cell culture obtained in step (a), pulverizing the pulverized tubular cell culture, mixing with purified water, and then preparing the composition by cold extraction, hot water extraction or ethanol extraction. As another example, the culture obtained in the step (a) may be dried by hot air and powdered to evaporate water.

In the present invention, physical and chemical elicitors can be used to increase the content of polyphenolic compounds, which are secondary metabolites. Specifically, it can be treated through the process shown in Experimental Example 2. Particularly, as a method for increasing the contents of carotenoids, flavonoids and phenolic compounds, which are secondary metabolites,

1. 100 to 300 μM, for example, 200 μM methyl jasmonic acid is added to MS medium.

2. Cell cultures of plant parts belonging to the Ranunculales are cultivated in a high frequency cell incubator and incubated for 3 to 10 minutes at a high frequency of 240 to 270 kHz, for example 240 or 270 kHz for one day, for example, Four times, for example three times, and repeats it for one week or two weeks.

It was confirmed that phenol and flavonoid contents were increased through the treatment with such an attractant, and in particular, it showed excellent efficacy in antioxidant, whitening, wrinkle improvement, anti-inflammation, moisturizing, reduction of pore, sebum suppression, antibacterial and acne improvement .

The cosmetic composition of the present invention may contain an acceptable carrier in cosmetic preparations. Herein, "an acceptable carrier for a cosmetic preparation" is a known or used compound or composition that can be contained in a cosmetic preparation, or a compound or composition to be developed in the future, which is toxic, instable or irritating It says nothing.

The carrier may be included in the cosmetic composition of the present invention in an amount of about 1% by weight to about 99.99% by weight, preferably about 90% by weight to about 99.99% by weight of the composition, based on the total weight thereof. However, since the ratio varies depending on the formulations described below in which the cosmetic composition of the present invention is prepared, and on the specific application site (face, neck, etc.) of the cosmetic composition and its preferable application amount, And should not be construed as limiting the scope of the invention.

Examples of the carrier include an alcohol, an oil, a surfactant, a fatty acid, a silicone oil, a wetting agent, a moisturizer, a viscous modifier, an emulsifier, a stabilizer, an ultraviolet scattering agent, an ultraviolet absorber, a coloring agent and a perfume. The compounds / compositions which can be used as the above-mentioned alcohol, oil, surfactant, fatty acid, silicone oil, humectant, humectant, viscous modifier, emulsifier, stabilizer, ultraviolet scattering agent, ultraviolet absorber, The person skilled in the art can select and use the appropriate substance / composition.

As an embodiment of the present invention, the cosmetic composition according to the present invention may contain glycerin, butylene glycol, propylene glycol, polyoxyethylene hydrogenated castor oil, ethanol, triethanolamine, etc., Preservatives, dyes, coloring agents, purified water and the like can be contained as needed.

The cosmetic composition according to the present invention can be prepared in various forms such as lotion, essence, gel, emulsion, lotion, cream (water-in-oil type, water-in-water type, multiphase), solution, suspension (Soft capsules, hard capsules) with a coating such as anhydrous products (oil and glycol), gel, mask, pack, powder, or gelatin.

The skin of the present invention includes not only a face but also a scalp and a whole body. The cosmetic composition applicable to such scalp is a shampoo, a rinse, a treatment, a hair removal agent, etc., and a body cleanser And can be manufactured in various forms as an application of the composition.

The method for preparing a cosmetic composition containing the extract of Laotiania cell culture extract according to the present invention is not limited to the above-described production method, and any person skilled in the art may make modifications , The cosmetic composition containing the extract of the tubercle cell cultured according to the present invention can be prepared.

In particular, the above-mentioned cosmetic composition can be produced in the form of a general emulsified formulation and a solubilized formulation, using a conventionally known manufacturing method, in addition to the manufacturing method specifically disclosed in the present invention.

When the cosmetic composition is manufactured from a cosmetic composition, the cosmetic composition of the emulsified formulation includes nutritional lotion, cream, essence and the like. It can also be prepared in the form of adjuvants which can be applied topically or systemically, which are conventionally used in the field of dermatology by containing a dermatologically acceptable medium or base.

In addition, suitable cosmetic formulations include, for example, solutions, gels, solid or kneaded anhydrous products, emulsions obtained by dispersing an oil phase in water, suspensions, microemulsions, microcapsules, microgranules or ionic forms (liposomes) May be provided in the form of a follicular dispersing agent of the type, cream, skin, lotion, powder, ointment, spray or conical stick. It can also be prepared in the form of a foam or an aerosol composition further containing a compressed propellant.

The cosmetic composition of the present invention may further contain at least one selected from the group consisting of fatty substances, organic solvents, solubilizers, thickeners and gelling agents, softening agents, antioxidants, suspending agents, stabilizers, foaming agents, perfumes, Or any other conventionally used in cosmetics such as nonionic emulsifiers, fillers, sequestering and chelating agents, preservatives, vitamins, barrier agents, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic active agents, ≪ RTI ID = 0.0 > cosmetics < / RTI > such as botanical ingredients, or adjuvants conventionally used in the field of dermatology. And, the above ingredients can be introduced in amounts commonly used in the field of dermatology.

The cosmetic composition according to the present invention is useful as an anti-inflammatory agent for skin diseases, such as skin moisturization, reduction of pores, suppression of sebum secretion, improvement of acne, antibacterial, antioxidant, prevention of skin aging due to ability to promote collagen synthesis, And functional cosmetics that can function as a therapeutic / preventive agent.

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these embodiments are only for illustrating the present invention and that the scope of the present invention is not construed as being limited by these embodiments.

Particularly, in the following examples, the efficacy of the extract of Ranunculus Acanthopanacis thunbergii cell line was confirmed, but it will be apparent to those skilled in the art that such an effect is also obtained in the culture itself, not in the extract.

Preparation of Extracts by Ranunculales according to the Present Invention

(1) Cell separation

1.1 Isolation of Lawn Cells from Ranunculales

Ranunculales For the cultivation, the buds of anemones, peonies, nymphs, chrysanthemums, lotus flowers, Christmas roses, peonies and water lilies are sterilized for 5 minutes in 70% ethanol and 0.5% sodium hypochlorite, I washed it several times with water. Carefully, the ovary wall was removed using tweezers, and the thyroid cells were separated and cultured in MS medium (Duchefa, Cat # M0256) containing 5% sucrose for 2 to 3 weeks.

1.2 Isolation of Ranunculales Flower Cells

For the cultivation, Ranunculales Typically, the petals of anemone, peony, nectar, chrysanthemum, lotus, Christmas rose, peony, and water lily are sterilized for 1 minute with 70% ethanol and 0.5% sodium hypochlorite, I washed it several times. Carefully cut with a tweezers and a sharp knife at once, and cut flower parts were dipped in MS medium (Duchefa, Cat # M0256) containing 5% sucrose and cultured for 2 to 3 weeks.

1.3 Isolation of Leaf Cells of Ranunculales

Ranunculales For the cultivation, the leaves of anemone, peony, nori, hiragami, lotus, Christmas rose, peony, and water lily were sterilized for 1 minute with 70% ethanol and 0.5% sodium hypochlorite, I washed it several times. Carefully cut with a tweezers and a sharp knife at once, and the cut leaves were dipped in MS medium (Duchefa, Cat # M0256) containing 5% sucrose and cultured for 2 to 3 weeks.

1.4 Isolation of Ranunculales Stem Cells

For the cultivation, Ranunculales Typically, stems of anemone, peony, nectar, chrysanthemum, lotus, Christmas rose, peony, and water lily are sterilized for 1 minute with 70% ethanol and 0.5% sodium hypochlorite, I washed it several times. Carefully cut with a tweezers and a sharp knife at once, and the cut stems were streaked on MS medium (Duchefa, Cat # M0256) containing 5% sucrose and cultured for 2 to 3 weeks.

1.5 Isolation of Ranunculales Root Cells

For the cultivation, Ranunculales Typically, the roots of anemone, peony, nori, cherry, lotus, Christmas rose, peony, and water lily are sterilized for 1 minute with 70% ethanol and 0.5% sodium hypochlorite, I washed it several times. Carefully cut with a tweezers and a sharp knife at once, and the cut root was dipped in MS medium (Duchefa, Cat # M0256) containing 5% sucrose and cultured for 2 to 3 weeks.

(2) Separation of Ranunculales (Ranunculales) Cultivation and mass production of placenta, flowers, leaves, stems and root cells

Ranunculales (A), as well as flowers, leaves, stems, and root cells were aseptically subcultured in MS medium containing 5% sucrose at intervals of 2 to 3 weeks, and then cultured in a balloon type bioreactor Samsung Scientific) was used to mass-produce liquid culture of the same composition except for agar at a temperature of 25 ° C and a constant air supply of 0.1 vvm at 14 day intervals.

(3) Production of Ranunculales (Ranunculales) Cultured Extract of Placenta, Flower, Leaf, Stem and Root Cell

Growing Ranunculales Typically, we harvest the anemone, peony, nectar, chrysanthemum, lotus flower, Christmas rose, peony flower, lily flower, flower, leaf, stem and root cells and remove enough moisture with a clean tissue Lt; RTI ID = 0.0 > 60 C < / RTI > for 2 days. 10 g of purified water is placed in a container, and the mixture is heated at 80 to 100 ° C. for 8 to 48 hours in a hot water distiller, An extract was obtained. After extraction, the mixture was filtered with a mesh to remove the solids, and anemone, peony root, nori, hiruma, lotus, Christmas rose, peony flower, and watery cell culture extracts were prepared.

Experimental Example 1: Influence on the cell survival rate of cell culture extracts of Ranunculales according to the present invention

Human keratinocyte (HaCaT) was cultured in an ATCC (American Type Culture Collection) in order to examine the proliferative activity of skin cell proliferation of Ranunculales by Ranunculales according to the present invention . The cells were inoculated into a 24-well culture plate at a concentration of 1 × 10 ⁴ cells / ml. The medium was DMEM (Dubelco's Modified Eagle Medium, BRL, USA) containing 10% FBS. After culturing in DMEM containing 10% FBS for 48 hours and culturing by 25-30% of the surface area of the culture vessel, FBS-free DMEM containing 10% Ranunculales placenta tissue culture extract prepared in Example 1 And then cultured for another 24 hours. 50 μl of a solution of 2.5 mg / ml of 3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide (MTT, Sigma M5655, USA) (DMSO, Sigma D2650, USA) solution was added to each well and the mixture was stirred for 20 minutes to dissolve the formazan crystals. Then, 100 μl of each of the formazan crystals was dissolved in 96 wells And the absorbance at 570 nm was measured by Enzyme-Linked Immunosorbent Assay (ELISA). The degree of proliferative activity of skin cells was calculated as a percentage based on the absorbance of the control group using pure water according to the following formula.

[Equation 1]

Cell viability effect (%) = (absorbance at the time of extract treatment / absorbance of control group) x 100

Treatment concentration part Cell survival rate effect (%)
(Cell Viability) Control group 100 2 % anemone Taurus 107 Flower 100 leaf 95 stem 98 Root 102 Peony Taurus 96 Flower 82 leaf 80 stem 75 Root 68 hepatica Taurus 90 Flower 70 leaf 75 stem 72 Root 78 pasqueflower Taurus 110 Flower 99 leaf 100 stem 102 Root 100 year Taurus 120 Flower 110 leaf 100 stem 120 Root 99 Christmas Rose Taurus 109 Flower 106 leaf 100 stem 99 Root 97 peony Taurus 90 Flower 87 leaf 89 stem 65 Root 60 training Taurus 115 Flower 105 leaf 102 stem 108 Root 110

Using the human keratinocyte, the cell culture extract of Ranunculales representative plant parts was treated to determine the suitable range for skin application, that is, the cell viability cell survival effect was confirmed. In the case of the cell culture extracts of 2% high-density buttercups seedlings, the roots of the peony, the flowers and stems of the larvae, the stem and roots of the peony were slightly toxic compared with the control group, Survival rates were not affected.

Experimental Example 2: Identification and content test for each extract

A physical and chemical elicitor was used to increase the content of polyphenolic compounds, a secondary metabolite in plant cells, by proliferating the plant belonging to Ranunculales belonging to the family Ranunculales and proliferating the cells.

 Methods for increasing the content of carotenoids, flavonoids and phenolic compounds, which are secondary metabolites, by site of plant cells belonging to Ranunculales include,

1. For cell culture of plant parts belonging to Ranunculales, 200 μM methyl jasmonic acid (MeJA) is cultured in MS medium.

2. Cell cultures of plant parts belonging to Ranunculales in Ranunculales were cultured in a high frequency cell incubator and treated with radio frequency (RF) at 240 or 270 kHz for 3 times at intervals of 5 minutes for one day and repeated for one week or two weeks.

The following experiments were carried out to confirm the total phenolic content and total flavonoid content of phenolic substances, flavonoids, and antioxidative effects of the major components of cell culture extracts of Ranunculales according to the present invention.

Total phenol content measurement

After adding 10 ml of distilled water to 1 ml of the extract, 2 ml of Folin-Ciocalteu phenol reagent (Sigma) was added and mixed, followed by reaction at room temperature for 5 minutes. 2 ml of 20% sodium carbonate was added to the reaction mixture, and the mixture was incubated at room temperature for 1 hour. Then, the absorbance was measured at 680 nm using a microplate reader (UVT-06685, Thermo max, USA). At this time, gallic acid (Sigma, USA) was used as the indicator substance, and the calibration curve of the indicator substance was prepared and the absorbance value of each sample was substituted and shown in the table.

Total flavonoid content measurement

2% AlCl36H2O dissolved in the same amount of methanol was mixed with 1.5 ml of the cell culture extract of Ranunculales at the site of Ranunculales. The mixture was incubated at room temperature for 10 minutes and then incubated with a microplate reader (UVT-06685, Thermo max, USA) Absorbance was measured at 367 nm. At this time, catechin (catechin, Sigma, USA) was used as the indicator material, and a calibration curve for the concentration of the indicator substance was prepared and the absorbance value of each sample was substituted and shown in the table.

process
density Incentive part Total phenol content
(mg ^-1 )
(gallic acid equivalents) Total flavonoid content (mg ^-1 )
(catechin equivalents) Control group 2 % anemone - Taurus 294 160 Flower 134 73 leaf 27 15 stem 16 9 Root 182 99 MeJA Taurus 401 218 Flower 214 116 leaf 134 73 stem 134 73 Root 267 145 RF Taurus 267 145 Flower 107 58 leaf 16 9 stem 16 9 Root 107 58 Peony - Taurus 331 180 Flower 27 15 leaf 53 29 stem 37 20 Root 11 6 MeJA Taurus 321 175 Flower 27 15 leaf 43 23 stem 27 15 Root 27 15 RF Taurus 428 233 Flower 134 73 leaf 150 81 stem 134 73 Root 134 73 hepatica - Taurus 321 175 Flower 27 15 leaf 32 17 stem 32 17 Root 43 23 MeJA Taurus 294 160 Flower 27 15 leaf 32 17 stem 32 17 Root 43 23 RF Taurus 283 154 Flower 32 17 leaf 27 15 stem 32 17 Root 32 17

pasqueflower - Taurus 380 207 Flower 64 35 leaf 32 17 stem 27 15 Root 203 111 MeJA Taurus 310 169 Flower 53 29 leaf 16 9 stem 16 9 Root 96 52 RF Taurus 481 262 Flower 160 87 leaf 139 76 stem 150 81 Root 310 169 year - Taurus 385 209 Flower 294 160 leaf 176 96 stem 182 99 Root 374 204 MeJA Taurus 321 175 Flower 278 151 leaf 160 87 stem 150 81 Root 267 145 RF Taurus 481 262 Flower 315 172 leaf 283 154 stem 289 157 Root 374 204 Christmas Rose - Taurus 267 145 Flower 225 122 leaf 32 17 stem 21 12 Root 187 102 MeJA Taurus 160 87 Flower 118 64 leaf 32 17 stem 21 12 Root 53 29 RF Taurus 321 175 Flower 294 160 leaf 80 44 stem 75 41 Root 160 87 peony - Taurus 449 244 Flower 11 6 leaf 27 15 stem 32 17 Root 91 49 MeJA Taurus 428 233 Flower 16 9 leaf 27 15 stem 27 15 Root 53 29 RF Taurus 374 204 Flower 16 9 leaf 21 12 stem 21 12 Root 27 15 training - Taurus 385 209 Flower 331 180 leaf 32 17 stem 64 35 Root 246 134 MeJA Taurus 363 198 Flower 321 175 leaf 27 15 stem 53 29 Root 160 87 RF Taurus 470 256 Flower 417 227 leaf 86 47 stem 102 55 Root 299 163

As a result of the above table, the cell culture extracts induced by MeJA induction agent in the Annamon spp. Showed high polyphenol content and flavonoid content in the anemone, and by the treatment with radiofrequency inducer, Highly induced cell culture extracts showed high polyphenol content and flavonoid content. By MeJA or by radiofrequency induction, the extracts of tuber cell cultures showed the highest contents of polyphenols and flavonoids compared to other parts of the seedlings.

EXPERIMENTAL EXAMPLE 3: Antioxidant Effect of Extract of Tacrolimus Extract from Lentinus edodes (Abyssinia sp.) (ABTS Radical Scavenging Activity)

The antioxidant activity of ABTS radicals was measured by the method of van den Berg et al., Which is a method using ABTS free radicals produced by the reaction with potassium persulfate, Respectively.

1.0 mM AAPH (2,2'-azo-bis 2-amidinopropanediol hydrochloride) was mixed with 2.5 mM ABTS (2,2'-azino-bis-3-ethylbenzenothiazolin-6-sulfonic acid) dissolved in 100 mM PBS buffer. Lt; 0 > C for 12 minutes. 2% Concentration The absorbance decreased at 734 nm while 20 μl of the extract of Example 1 and 980 μl ABTS solution were reacted in a 37 ° C water bath for 10 minutes, and the positive control group was 0.1% Trolox. The ABTS radical scavenging activity was calculated according to the following equation (2).

&Quot; (2) "

process
density Incentive part ABTS radical scavenging activity
(% of control) Control group 0 2 % anemone - Taurus 55 Flower 25 leaf 5 stem 3 Root 34 MeJA Taurus 75 Flower 40 leaf 25 stem 25 Root 50 RF Taurus 50 Flower 20 leaf 3 stem 3 Root 20 Peony - Taurus 62 Flower 5 leaf 10 stem 7 Root 2 MeJA Taurus 60 Flower 5 leaf 8 stem 5 Root 5 RF Taurus 80 Flower 25 leaf 28 stem 25 Root 25 hepatica - Taurus 60 Flower 5 leaf 6 stem 6 Root 8 MeJA Taurus 55 Flower 5 leaf 6 stem 6 Root 8 RF Taurus 53 Flower 6 leaf 5 stem 6 Root 6

pasqueflower - Taurus 71 Flower 12 leaf 6 stem 5 Root 38 MeJA Taurus 58 Flower 10 leaf 3 stem 3 Root 18 RF Taurus 90 Flower 30 leaf 26 stem 28 Root 58 year - Taurus 72 Flower 55 leaf 33 stem 34 Root 70 MeJA Taurus 60 Flower 52 leaf 30 stem 28 Root 50 RF Taurus 90 Flower 59 leaf 53 stem 54 Root 70 Christmas Rose - Taurus 50 Flower 42 leaf 6 stem 4 Root 35 MeJA Taurus 30 Flower 22 leaf 6 stem 4 Root 10 RF Taurus 60 Flower 55 leaf 15 stem 14 Root 30 peony - Taurus 84 Flower 2 leaf 5 stem 6 Root 17 MeJA Taurus 80 Flower 3 leaf 5 stem 5 Root 10 RF Taurus 70 Flower 3 leaf 4 stem 4 Root 5 training - Taurus 72 Flower 62 leaf 6 stem 12 Root 46 MeJA Taurus 68 Flower 60 leaf 5 stem 10 Root 30 RF Taurus 88 Flower 78 leaf 16 stem 19 Root 56

As can be seen from the above table, it was confirmed that the tubular cell culture extract of 2% concentration of the butterfly seeds of the present invention showed the most excellent antioxidant ability than the cell culture extract derived from other sites. Especially, MeJA and RF induction agent showed higher antioxidant ability.

Experimental Example 4: Measurement of inhibition of melanogenesis using B16F10 malanoma cells

Melanin-producing cells, B16F10, were cultured in DMEM medium supplemented with 10% FBS and 1% antibiotics at 37 ° C, 5%, and CO2. B16F10 cells were suspended in DMEM medium containing 10% FBS at a concentration of 1.5 × 10 ³ cells / well in order to confirm the change of melanin production by 2% Suspension cells (5 ml) are placed in a tissue culture flask and allowed to attach for 1 day. After the addition, the test samples were incubated at 37 ° C for 5 days in 5% CO 2. The cultured B16F10 cells were washed with phosphate buffered saline (PBS) and trypsinized. Cells were collected in a corning tube, and 1 × 10 ⁶ cells / well of 1N NaOH solution 1 was added to dissolve the cells. Melanogenesis inhibition rates were determined by measuring the absorbance at 490 nm with a microplate reader.

&Quot; (3) "

Inhibition of melanogenesis (%) = [1 - (Absample / Abcontrol)] 100

Treatment concentration part Melanin Contents
(% of control) Control group 100 2 % anemone Taurus 91 Flower 100 leaf 95 stem 100 Root 98 Peony Taurus 86 Flower 100 leaf 101 stem 100 Root 100 hepatica Taurus 80 Flower 100 leaf 99 stem 95 Root 98 pasqueflower Taurus 80 Flower 100 leaf 98 stem 100 Root 100 Lotus Taurus 75 Flower 90 leaf 100 stem 85 Root 95 Christmas Rose Taurus 83 Flower 95 leaf 98 stem 100 Root 100 Peony flower Taurus 80 Flower 98 leaf 97 stem 100 Root 100 Training Taurus 85 Flower 95 leaf 98 stem 92 Root 90

In order to examine the effect of the cell culture extracts of Bacillus subtilis on the melanin synthesis of B16 / F10 melanoma cells, total melanin levels were measured after culturing for 3 days at a concentration of 2%. As a result, it was confirmed that the whitening effect of the cell culture extract of the 2% parsley seedlings by the site decreased the melanin synthesis only in the thalamus region, and the whitening effect was insufficient in the remaining region. In addition, in the case of the cells cultured by the treatment with 200 μM methyl jasmonic acid inducer, the extract of the tuber cell cultures decreased the melanin synthesis twice as much as that without the inducer.

Experimental Example 5: Procollagen synthesis enhancement test

Human fibroblasts (Human Skin Fibroblast) for 37 ℃, 5% in a CO ₂ incubator of 10% in the cell culture medium to maintain a constant humidity FBS, Penicillin (50U / ml) , Streptomycin DMEM (Dulbecco's containing (50 / ml) (Gibco, USA). Cells were seeded at a density of 1 × 10 ⁵ cells / ml in a 24-well plate and cultured for 24 hours. The cells were cultured in a 5% CO ₂ incubator at 37 ° C for 48 hours in a medium containing a control (DMEM medium) and a cell culture extract of the plant part of the seedlings obtained in Example 1 at a concentration of 2%. After 48 hours, the amount of newly synthesized collagen was measured by measuring 20 μl of each supernatant of the culture medium and measuring it using Procollagen Type I C-Peptide EIA Kit (PICP, Takara, Cat No. MK101). The amount of PICP was converted into ng / 1 x 10 < ⁵ > cells, calculated according to the following formula, and the results are shown in Table 5.

&Quot; (4) "

Increase rate of collagen biosynthesis (%) = [(amount of collagen in test group) / (amount of collagen in control group)] × 100

Treatment concentration part Increase in collagen biosynthesis
(% of control) Control group 100 2 % anemone Taurus 117 Flower 110 leaf 105 stem 108 Root 112 Peony Taurus 106 Flower 92 leaf 90 stem 85 Root 78 hepatica Taurus 100 Flower 80 leaf 85 stem 82 Root 88 pasqueflower Taurus 120 Flower 109 leaf 110 stem 112 Root 110 Lotus Taurus 130 Flower 120 leaf 110 stem 130 Root 109 Christmas Rose Taurus 119 Flower 116 leaf 110 stem 109 Root 107 Peony flower Taurus 100 Flower 97 leaf 109 stem 75 Root 70 Training Taurus 125 Flower 115 leaf 112 stem 118 Root 120

As can be seen from the above table, the amount of procollagen biosynthesis was greatly increased by treatment of the tuber cell culture extract of the plant part of the seedlings at 2% concentration of Example 1, and 200 μM methyl jasmonate jasmonic acid), the amount of procollagen biosynthesis increased 2.5 times more than that of untreated cells.

Experimental Example 6: Antiinflammatory effect experiment by inhibition of iNOS activity

The amount of nitric oxide (NO) produced from the Raw 264.7 cell line was determined using the Griess reagent as the NO ₂ ^- form present in the cell culture medium. In order to measure inhibition of NO production in 1 g LPS-activated Raw 264.7 cells, cells were cultured for 24 h in the experimental group treated with 2% concentration of cell culture extracts of Ranunculaceae, and the Griess reagent was added to the cell culture supernatant And 100 μl of Griess reagent (1% sulfanilamide in 5% phosphoric acid, 1% -naphthylamide in H ₂ O) were mixed and reacted on 96-well plates for 10 minutes and absorbance was measured at 540 nm. The concentration of NO ₂ ^- was obtained by measuring the absorbance by diluting sodium nitrate.

Treatment concentration part Cell Viability (%) NO Production (M) LPS treatment
(-) - Control group - 100 1.5 Control group 65 3.5




LPS treatment
(+)



2 % anemone Taurus 107 2.5 Flower 100 2.9 leaf 95 2.8 stem 98 2.7 Root 102 2.3 Peony Taurus 96 2.0 Flower 82 3.5 leaf 80 3.4 stem 75 3.5 Root 68 3.5 hepatica Taurus 101 1.9 Flower 70 3.0 leaf 75 3.2 stem 72 3.2 Root 78 3.3 pasqueflower Taurus 110 2.0 Flower 99 3.5 leaf 100 3.4 stem 102 2.8 Root 100 3.5 Lotus Taurus 120 1.5 Flower 110 2.3 leaf 100 2.0 stem 110 2.0 Root 99 2.4 Christmas Rose Taurus 109 2.7 Flower 106 3.0 leaf 100 3.5 stem 99 3.4 Root 97 3.5 Peony flower Taurus 90 1.9 Flower 87 3.4 leaf 89 3.3 stem 65 3.5 Root 60 3.4 Training Taurus 105 1.8 Flower 105 2.0 leaf 102 2.3 stem 100 2.1 Root 100 2.3

As shown in the above table, the activity of iNOS inhibited NO production and showed the most excellent anti-inflammatory effect compared to the control group by treatment with a tuber cell culture extract of 2% NaNO3. In addition, the cells were cultured in a high frequency cell incubator for 3 days at a frequency of 240 to 270 kHz for 5 days and cultured in a high frequency cell culture incubator. INOS activity was 1.8 times more than that of untreated cells, whereas other cell culture extracts were ineffective.

Experimental Example 7: Skin moisturizing effect experiment

1. Anemone

Raw material name Weight% (w / w) compare
Produce
Example 1 Produce
Example 1-1 Produce
Example 1-2 Produce
Example 1-3 Produce
Example 1-4 Produce
Example 1-5 Cetostearyl alcohol 1.0 1.0 1.0 1.0 1.0 1.0 Self emulsifying monostearic acid 1.0 1.0 1.0 1.0 1.0 1.0 Wax 0.5 0.5 0.5 0.5 0.5 0.5 Squalane 5.0 5.0 5.0 5.0 5.0 5.0 Isocetyl octanoate 3.0 3.0 3.0 3.0 3.0 3.0 Dimethylsiloxane 0.3 0.3 0.3 0.3 0.3 0.3 Sorbitan monostearate 0.5 0.5 0.5 0.5 0.5 0.5 Polyethylene glycol monostearate 8.0 8.0 8.0 8.0 8.0 8.0 glycerin 4.0 4.0 4.0 4.0 4.0 4.0 Propylene glycol 0.2 0.2 0.2 0.2 0.2 0.2 Carboxy polymer 0.22 0.22 0.22 0.22 0.22 0.22 Triethanolamine 0.25 0.25 0.25 0.25 0.25 0.25 antiseptic Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Spices Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount flash Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Anemone Laotian cell culture extract - 2 - - - - Anemone flower cell culture extract - - 2 - - - Anemone leaf cell culture extract - - - 2 - - Anemone stem cell culture extract - - - - 2 - Anemone root cell culture extract - - - - - 2 Purified water to 100 to 100 to 100 to 100 to 100 to 100

2. Peony

Raw material name Weight% (w / w) compare
Produce
Example 2 Produce
Example 2-1 Produce
Example 2-2 Produce
Example 2-3 Produce
Example 2-4 Produce
Example 2-5 Cetostearyl alcohol 1.0 1.0 1.0 1.0 1.0 1.0 Self emulsifying monostearic acid 1.0 1.0 1.0 1.0 1.0 1.0 Wax 0.5 0.5 0.5 0.5 0.5 0.5 Squalane 5.0 5.0 5.0 5.0 5.0 5.0 Isocetyl octanoate 3.0 3.0 3.0 3.0 3.0 3.0 Dimethylsiloxane 0.3 0.3 0.3 0.3 0.3 0.3 Sorbitan monostearate 0.5 0.5 0.5 0.5 0.5 0.5 Polyethylene glycol monostearate 8.0 8.0 8.0 8.0 8.0 8.0 glycerin 4.0 4.0 4.0 4.0 4.0 4.0 Propylene glycol 0.2 0.2 0.2 0.2 0.2 0.2 Carboxy polymer 0.22 0.22 0.22 0.22 0.22 0.22 Triethanolamine 0.25 0.25 0.25 0.25 0.25 0.25 antiseptic Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Spices Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount flash Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Peony cell culture extract - 2 - - - - Peony flower cell culture extract - - 2 - - - Peony leaf cell culture extract - - - 2 - - Peony stem cell culture extract - - - - 2 - Peony root cell culture extract - - - - - 2 Purified water to 100 to 100 to 100 to 100 to 100 to 100

3. Nori

Raw material name Weight% (w / w) compare
Produce
Example 3 Produce
Example 3-1 Produce
Example 3-2 Produce
Example 3-3 Produce
Example 3-4 Produce
Example 3-5 Cetostearyl alcohol 1.0 1.0 1.0 1.0 1.0 1.0 Self emulsifying monostearic acid 1.0 1.0 1.0 1.0 1.0 1.0 Wax 0.5 0.5 0.5 0.5 0.5 0.5 Squalane 5.0 5.0 5.0 5.0 5.0 5.0 Isocetyl octanoate 3.0 3.0 3.0 3.0 3.0 3.0 Dimethylsiloxane 0.3 0.3 0.3 0.3 0.3 0.3 Sorbitan monostearate 0.5 0.5 0.5 0.5 0.5 0.5 Polyethylene glycol monostearate 8.0 8.0 8.0 8.0 8.0 8.0 glycerin 4.0 4.0 4.0 4.0 4.0 4.0 Propylene glycol 0.2 0.2 0.2 0.2 0.2 0.2 Carboxy polymer 0.22 0.22 0.22 0.22 0.22 0.22 Triethanolamine 0.25 0.25 0.25 0.25 0.25 0.25 antiseptic Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Spices Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount flash Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Leucocyte cell culture extract - 2 - - - - Reticulata flower cell culture extract - - 2 - - - Leaf leaf cell culture extract - - - 2 - - Rectal stem cell culture extract - - - - 2 - Root cell culture extract - - - - - 2 Purified water to 100 to 100 to 100 to 100 to 100 to 100

4. Flowers

Raw material name Weight% (w / w) compare
Produce
Example 4 Produce
Example 4-1 Produce
Example 4-2 Produce
Example 4-3 Produce
Example 4-4 Produce
Example 4-5 Cetostearyl alcohol 1.0 1.0 1.0 1.0 1.0 1.0 Self emulsifying monostearic acid 1.0 1.0 1.0 1.0 1.0 1.0 Wax 0.5 0.5 0.5 0.5 0.5 0.5 Squalane 5.0 5.0 5.0 5.0 5.0 5.0 Isocetyl octanoate 3.0 3.0 3.0 3.0 3.0 3.0 Dimethylsiloxane 0.3 0.3 0.3 0.3 0.3 0.3 Sorbitan monostearate 0.5 0.5 0.5 0.5 0.5 0.5 Polyethylene glycol monostearate 8.0 8.0 8.0 8.0 8.0 8.0 glycerin 4.0 4.0 4.0 4.0 4.0 4.0 Propylene glycol 0.2 0.2 0.2 0.2 0.2 0.2 Carboxy polymer 0.22 0.22 0.22 0.22 0.22 0.22 Triethanolamine 0.25 0.25 0.25 0.25 0.25 0.25 antiseptic Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Spices Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount flash Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Acanthopanax senticosus cell culture extract - 2 - - - - Hwasung flower cell culture extract - - 2 - - - Leaf flower cultivation extract - - - 2 - - Stem Cell Culture Extract - - - - 2 - Root cell culture extract - - - - - 2 Purified water to 100 to 100 to 100 to 100 to 100 to 100

5. Year

Raw material name Weight% (w / w) compare
Produce
Example 5 Produce
Example 5-1 Produce
Example 5-2 Produce
Example 5-3 Produce
Example 5-4 Produce
Example 5-5 Cetostearyl alcohol 1.0 1.0 1.0 1.0 1.0 1.0 Self emulsifying monostearic acid 1.0 1.0 1.0 1.0 1.0 1.0 Wax 0.5 0.5 0.5 0.5 0.5 0.5 Squalane 5.0 5.0 5.0 5.0 5.0 5.0 Isocetyl octanoate 3.0 3.0 3.0 3.0 3.0 3.0 Dimethylsiloxane 0.3 0.3 0.3 0.3 0.3 0.3 Sorbitan monostearate 0.5 0.5 0.5 0.5 0.5 0.5 Polyethylene glycol monostearate 8.0 8.0 8.0 8.0 8.0 8.0 glycerin 4.0 4.0 4.0 4.0 4.0 4.0 Propylene glycol 0.2 0.2 0.2 0.2 0.2 0.2 Carboxy polymer 0.22 0.22 0.22 0.22 0.22 0.22 Triethanolamine 0.25 0.25 0.25 0.25 0.25 0.25 antiseptic Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Spices Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount flash Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Tumor cell culture extract - 2 - - - - Yeast cell culture extract - - 2 - - - Yeast leaf cell culture extract - - - 2 - - Cultured stem cell culture extract - - - - 2 - Root cell culture extract - - - - - 2 Purified water to 100 to 100 to 100 to 100 to 100 to 100

6. Christmas Rose

Raw material name Weight% (w / w) compare
Produce
Example 6 Produce
Example 6-1 Produce
Example 6-2 Produce
Example 6-3 Produce
Example 6-4 Produce
Example 6-5 Cetostearyl alcohol 1.0 1.0 1.0 1.0 1.0 1.0 Self emulsifying monostearic acid 1.0 1.0 1.0 1.0 1.0 1.0 Wax 0.5 0.5 0.5 0.5 0.5 0.5 Squalane 5.0 5.0 5.0 5.0 5.0 5.0 Isocetyl octanoate 3.0 3.0 3.0 3.0 3.0 3.0 Dimethylsiloxane 0.3 0.3 0.3 0.3 0.3 0.3 Sorbitan monostearate 0.5 0.5 0.5 0.5 0.5 0.5 Polyethylene glycol monostearate 8.0 8.0 8.0 8.0 8.0 8.0 glycerin 4.0 4.0 4.0 4.0 4.0 4.0 Propylene glycol 0.2 0.2 0.2 0.2 0.2 0.2 Carboxy polymer 0.22 0.22 0.22 0.22 0.22 0.22 Triethanolamine 0.25 0.25 0.25 0.25 0.25 0.25 antiseptic Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Spices Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount flash Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Christmas Rose Tacrolimus Cell Culture Extract - 2 - - - - Christmas rose flower cell culture extract - - 2 - - - Christmas rose leaf cell culture extract - - - 2 - - Christmas Rose stem cell culture extract - - - - 2 - Christmas rose root cell culture extract - - - - - 2 Purified water to 100 to 100 to 100 to 100 to 100 to 100

7. Peony

Raw material name Weight% (w / w) compare
Produce
Example 7 Produce
Example 7-1 Produce
Example 7-2 Produce
Example 7-3 Produce
Example 7-4 Produce
Example 7-5 Cetostearyl alcohol 1.0 1.0 1.0 1.0 1.0 1.0 Self emulsifying monostearic acid 1.0 1.0 1.0 1.0 1.0 1.0 Wax 0.5 0.5 0.5 0.5 0.5 0.5 Squalane 5.0 5.0 5.0 5.0 5.0 5.0 Isocetyl octanoate 3.0 3.0 3.0 3.0 3.0 3.0 Dimethylsiloxane 0.3 0.3 0.3 0.3 0.3 0.3 Sorbitan monostearate 0.5 0.5 0.5 0.5 0.5 0.5 Polyethylene glycol monostearate 8.0 8.0 8.0 8.0 8.0 8.0 glycerin 4.0 4.0 4.0 4.0 4.0 4.0 Propylene glycol 0.2 0.2 0.2 0.2 0.2 0.2 Carboxy polymer 0.22 0.22 0.22 0.22 0.22 0.22 Triethanolamine 0.25 0.25 0.25 0.25 0.25 0.25 antiseptic Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Spices Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount flash Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Peony Laotian Cell Culture Extract - 2 - - - - Peony flower cell culture extract - - 2 - - - Peony leaf cell culture extract - - - 2 - - Peony stem cell culture extract - - - - 2 - Peony root cell culture extract - - - - - 2 Purified water to 100 to 100 to 100 to 100 to 100 to 100

8. Training

Raw material name Weight% (w / w) compare
Produce
Example 8 Produce
Example 8-1 Produce
Example 8-2 Produce
Example 8-3 Produce
Example 8-4 Produce
Example 8-5 Cetostearyl alcohol 1.0 1.0 1.0 1.0 1.0 1.0 Self emulsifying monostearic acid 1.0 1.0 1.0 1.0 1.0 1.0 Wax 0.5 0.5 0.5 0.5 0.5 0.5 Squalane 5.0 5.0 5.0 5.0 5.0 5.0 Isocetyl octanoate 3.0 3.0 3.0 3.0 3.0 3.0 Dimethylsiloxane 0.3 0.3 0.3 0.3 0.3 0.3 Sorbitan monostearate 0.5 0.5 0.5 0.5 0.5 0.5 Polyethylene glycol monostearate 8.0 8.0 8.0 8.0 8.0 8.0 glycerin 4.0 4.0 4.0 4.0 4.0 4.0 Propylene glycol 0.2 0.2 0.2 0.2 0.2 0.2 Carboxy polymer 0.22 0.22 0.22 0.22 0.22 0.22 Triethanolamine 0.25 0.25 0.25 0.25 0.25 0.25 antiseptic Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Spices Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount flash Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Suitable amount Extract - 2 - - - - Lily flower cell culture extract - - 2 - - - Lily leaf cell culture extract - - - 2 - - Water stem cell culture extract - - - - 2 - Root cell culture extract of water lily - - - - - 2 Purified water to 100 to 100 to 100 to 100 to 100 to 100

In order to examine the skin moisturizing ability during one application, the cosmetic composition was applied to the inside of the face and anterior chamber to 50 persons who were in the 30s to 40s who were thought to be dry skin or dry skin, The skin moisture was measured using a corneometer (CM820 courage Khazaka electronic GmbH, Germany). The coneometer measures the amount of moisture present in the epidermis of the skin by measuring the degree of ion of water using a sensor and numerically expressing the amount of water to calculate the amount of moisture, and the measurement method is as follows.

1) A coneometer probe was placed on the skin area to be measured.

2) When the probe is pressed onto the skin, the electrical conductivity of the skin is quantified through the sensor and displayed on the screen.

3) Measurement was repeated with different measurement sites.

4) After the measurement, the sensor was wiped with a tissue paper such as a kim wipe and then measured again.

The moisture content of the skin was measured using a coneometer at a constant temperature and humidity (40% humidity) under the constant temperature and humidity conditions before initiation of the application, and the changes were measured after 12 hours and 24 hours. The results are shown in the following table.

Skin moisture (%) sample Before application After 12 hours After 24 hours Comparative Preparation Example 1 30.9 38.5 31.4 Production Example 1-1 30.5 41.4 37.7 Production Example 1-2 30.3 39.9 36.3 Production Example 1-3 29.9 39.9 35.9 Production Example 1-4 30.2 40.3 36.5 Production Example 1-5 30.5 41.5 35.6 Comparative Production Example 2 30.7 38.5 31.4 Production Example 2-1 30.4 45.1 43.3 Production example 2-2 30.8 40.1 37.9 Production Example 2-3 30.1 41.2 37.8 Production example 2-4 30.2 41.5 37.5 Production example 2-5 30.3 40.3 37.1 Comparative Production Example 3 30.9 38.4 31.2 Production example 3-1 30.5 46.2 42.2 Production example 3-2 30.3 43.1 39.1 Production Example 3-3 29.9 42.8 38.2 Production example 3-4 30.2 42.1 38.7 Production Example 3-5 30.5 42.5 37.9 Comparative Production Example 4 30.6 38.6 31.6 Production Example 4-1 30.1 44.1 40.7 Production example 4-2 30.1 42.1 36.3 Production Example 4-3 30.1 41.9 36.9 Production Example 4-4 30.2 40.9 36.5 Production Example 4-5 30.3 40.5 37.7 Comparative Preparation Example 5 30.6 38.2 31.2 Production Example 5-1 30.2 48.9 44.5 Production Example 5-2 30.3 43.2 42.1 Production example 5-3 30.9 45.1 41.2 Production Example 5-4 30.2 44.9 41.9 Production Example 5-5 30.5 45.0 41.1 Comparative Preparation Example 6 30.7 38.4 31.5 Production example 6-1 30.4 45.2 43.1 Production Example 6-2 30.8 44.4 42.9 Production example 6-3 30.1 43.2 41.9 Production Example 6-4 30.2 44.5 40.8 Production Example 6-5 30.3 43.4 41.5 Comparative Preparation Example 7 30.9 38.3 31.8 Production Example 7-1 30.5 46.0 41.0 Production Example 7-2 30.3 41.2 42.1 Production Example 7-3 30.1 41.3 40.0 Production Example 7-4 30.2 42.5 39.9 Production Example 7-5 30.5 42.6 41.1 Comparative Preparation Example 8 30.7 38.2 31.0 Production Example 8-1 30.4 49.0 44.0 Production Example 8-2 30.8 43.2 42.1 Production example 8-3 30.1 46.1 41.2 Production Example 8-4 30.2 45.9 41.9 Production Example 8-5 30.3 46.0 41.1

From the above results, it was found that when Comparative Production Examples 1 to 8 were applied, the amount of skin moisture was increased to some extent after 12 hours from application, but returned to the state before application after 24 hours. On the other hand, Examples 1-1 to 8-2, which are skin external composition compositions containing cell culture extracts of each part of ananomonas, peonies, nori, hiragana, kite, Christmas rose, peony and water lily of representative plants of 2% 5 was applied to the skin, it was confirmed that the moisturizing power lasts more than 24 hours and the skin moisture content increase effect is also higher than that of Comparative Production Examples 1 to 8.

In addition, 200 μM methyl jasmonic acid inducer treatment and high frequency cell culture were performed for 3 days at a frequency of 240 to 270 kHz for 5 days, , The extracts of Tacrolimus cell cultures showed 1.5 times more skin moisture content than those without Tacrolimus treatment.

Experimental Example 8: In vitro Test

In order to measure the pore-shrinking effect of the composition of the present invention, the composition prepared in the above example was tested for pore-shrinking effect using hemoglobin as a protein to replace skin. A hemoglobin solution (0.05 g / 50 ml) was prepared with 0.9% Phosphate Buffer Saline (0.1 mM, pH 7.4), 2 ml of the composition obtained in the above Example was added to 2 ml of the hemoglobin solution, The mixture was centrifuged at 3,500 rpm for 10 minutes, and 2 ml of purified water was added to 1 ml of the supernatant. The measurement was carried out at 407 nm in a UV-Visible spectrum. As a control, 2 ml of PBS (Phosphate Buffer Saline) was added in place of the composition of the above Example. In the case of the experimental group containing various concentrations of the composition, the shrinkage effect was measured by measuring the amount of hemoglobin settled. The higher the sedimentation rate (%) of hemoglobin was, the better the shrinkage effect of pores was judged.

The shrinkage effect was evaluated by measuring the sedimentation amount of hemoglobin. The higher the sedimentation rate (%) of hemoglobin was, the better the shrinkage effect was judged. As shown in the table below, the cell culture extracts of 2% parsley seedlings showed significant differences and showed pore shrinking effect. In addition, 200 μM methyl jasmonic acid inducer treatment and high frequency cell culture were performed for 3 days at a frequency of 240 to 270 kHz for 5 days, , The extract of Tacrolimus cell culture showed a pore-shrinking effect of 2.2 times better than that of the untreated case.

Treatment concentration part Precipitation of hemoglobin
(% of control) Control group 2.9 2 % anemone Taurus 24.8 Flower 15.3 leaf 11.1 stem 13.1 Root 7.9 Peony Taurus 19.3 Flower 12.3 leaf 15.7 stem 13.0 Root 11.1 hepatica Taurus 5.1 Flower 2.9 leaf 3.5 stem 3.0 Root 2.5 pasqueflower Taurus 28.0 Flower 20.1 leaf 17.9 stem 15.5 Root 13.1 Lotus Taurus 33.0 Flower 30.4 leaf 25.8 stem 21.1 Root 25.8 Christmas Rose Taurus 20.3 Flower 15.5 leaf 15.2 stem 8.8 Root 7.9 Peony flower Taurus 20.3 Flower 10.3 leaf 8.9 stem 5.3 Root 7.9 Training Taurus 32.4 Flower 16.2 leaf 8.9 stem 8.0 Root 5.5

Experimental Example 9: Effect of inhibiting sebum

Ten subjects were tested for secretion of sebum on skin using a skin oil analyzer (Sebum meter SM810) 4 weeks after using the cell culture extract of 2% parsley seedlings of the present invention. Experiments were performed at 22 ± 2 ° C and 40 ± 5% humidity. The measurement value for oily skin is 220 / / cm ² or more, and for normal skin is 100 to 220 / / cm ² . Purified water was used as a control, and the experimental results are shown in the table.

Treatment concentration part Sebum suppression effect 0 day (before use) 28 days (after use) Control group 230 225 2 % anemone Taurus 231 175 Flower 229 200 leaf 230 205 stem 232 201 Root 230 210 Peony Taurus 229 200 Flower 231 210 leaf 229 215 stem 230 195 Root 232 199 hepatica Taurus 230 210 Flower 231 205 leaf 232 205 stem 231 210 Root 231 190 pasqueflower Taurus 230 190 Flower 232 195 leaf 231 205 stem 231 210 Root 230 205 Lotus Taurus 230 155 Flower 231 175 leaf 231 180 stem 230 190 Root 231 185 Christmas Rose Taurus 231 175 Flower 230 178 leaf 230 180 stem 231 190 Root 230 191 Peony flower Taurus 230 179 Flower 231 199 leaf 231 205 stem 230 200 Root 229 210 Training Taurus 229 170 Flower 231 185 leaf 230 190 stem 229 190 Root 230 195

As shown in the above table, the extract of Tacrine cell culture of 2% parsley seedlings showed the best inhibitory effect on sebum secretion compared with the control group. In addition, 200 μM methyl jasmonic acid inducer treatment and high frequency cell culture were performed for 3 days at a frequency of 240 to 270 kHz for 5 days, , The extract of Tacrolimus cell culture showed 2.5 times more inhibitory effect on sebum secretion than those without treatment.

Experimental Example 10: Measurement of antibacterial activity

The antimicrobial test was carried out by the solid culture dilution method by the test for the antimicrobial activity of the extract of the tuberous cell culture of 2% parsley seedlings of the present invention of the present invention. Staphylococcus aureus KCTC 6910, Gram-negative bacteria Pseudomonas aeruginosa KCTC 1637, E. coli KCTC 1039, and Candida yeast, respectively, were purchased from Korea Research Institute of Bioscience and Biotechnology. albicans KCTC 7965) and aspergillus (Aspergillus nigerKCTC 6910).

Agar Serial Dilution Method

Tritic soy broth was used as the fungi, and the broth was inoculated on the medium and pre-cultured for 24 hours in a 37 ° C shaking incubator. Potato dextrose culture medium was used for culture of yeast, and the bacteria were inoculated on the medium and cultured for 2 days in a 25 ° C shaking incubator. Potato dextrose agar medium was used for culture of molds, and the bacteria were inoculated on the medium and pre-cultured for 7 days in a 25 ° C incubator.

More specifically, in the case of bacteria, bacteria are inoculated on a tryptic soy broth and cultured at 37 ° C for 24 hours. In the case of yeast, bacteria are inoculated on a potato dextrose medium and cultured for 2 days at 25 ° C. The filamentous fungus was inoculated on a potato dextrose agar medium and pre-cultured for 7 days in a 25 ° C. incubator. Spores of filamentous fungus formed on the surface of the medium were recovered by using a smear rod and diluted in sterilized saline. 2 ml of each of the extracts diluted to the appropriate concentration in 5% DMSO (dimethylsulfoxide) physiological saline solution was added to the sterilized Patriedish by the sample and the experimental species, and the control was added to 2 ml of each sample diluted to a proper concentration in 5% physiological saline solution , And 2 ml of a 5% DMSO physiological saline solution was added to the control. Then, 18 ml of a tryptic soya agar medium and a potato dextrose agar medium, which had been sterilized in each Patridish and cooled to 48 ° C, were added, stirred and allowed to stand and solidify. Each of the pre-cultivated test bacteria was applied to each petri dish at a final concentration of about 1 × 10 ⁶ CFU / ml for bacteria, 1 × 10 ⁵ CFU / ml for yeast, about 1 × 10 ⁵ CFU / ⁴ CFU / ml in each petri dish. Each petri dish was incubated at 37 ° C for 24 hours, and the yeast was incubated at 25 ° C for 3 days. The molds were cultured in a 25 ° C incubator for 7 days, and then cloin formation was observed in each compartment. The minimum sample concentration was defined as the minimum inhibitory concentration (MIC), and the results are shown in the table. At this time, the minimum inhibitory concentration means that the smaller the value, the higher the antibacterial effect.

Treatment concentration part Minimum inhibitory concentration (MIC, g / ml) S. aureus P. aeruginosa E. Coli C. albicans A. niger 2 % anemone Taurus 315 276 122 271 39 Flower 350 289 132 273 40 leaf 325 302 125 277 42 stem 370 310 130 276 46 Root 370 307 133 280 43 Peony Taurus 245 211 98 231 31 Flower 252 230 100 233 35 leaf 266 225 106 235 33 stem 272 221 101 250 34 Root 280 238 103 251 39 hepatica Taurus 363 297 154 298 49 Flower 366 305 157 300 52 leaf 383 310 160 305 56 stem 400 313 163 303 57 Root 403 316 169 301 59 pasqueflower Taurus 281 231 132 255 38 Flower 286 232 133 259 41 leaf 290 235 135 261 39 stem 310 238 140 263 39 Root 315 233 138 259 37 Lotus Taurus 497 439 298 351 198 Flower 508 440 299 353 199 leaf 515 441 297 354 199 stem 520 443 297 352 201 Root 525 439 296 353 198 Christmas Rose Taurus 377 298 169 401 121 Flower 380 299 169 418 120 leaf 400 300 172 405 121 stem 460 301 172 405 121 Root 425 298 171 406 120 Peony flower Taurus 534 513 331 399 273 Flower 550 515 331 400 273 leaf 558 513 333 405 275 stem 500 512 335 399 279 Root 525 515 333 401 278 Training Taurus 433 319 197 436 151 Flower 463 319 197 436 151 leaf 460 330 190 433 153 stem 450 320 190 436 152 Root 453 321 193 450 155

As shown in the table, all of the cell culture extracts of the 2% parsley herbaceous plant-derived plant-derived parts exhibited antibacterial effects. In particular, the cell culture extracts from the taiyoshi showed excellent antimicrobial activity. In addition, 200 μM methyl jasmonic acid inducer treatment and high frequency cell culture were performed for 3 days at a frequency of 240 to 270 kHz for 5 days, , The extract of Tacrolimus cell culture showed 1.5 times better antimicrobial effect than the untreated case.

Experimental Example 11: Effect of improving acne

Ten men and women who developed acne were asked to apply the lotion thoroughly over the face for 4 weeks in the morning and evening. The results of the evaluation of the acne healing effect were evaluated according to the degree of the person who participated in the experiment by evaluating the pre-test condition by 1-3 points with reference to the following criteria, and the results are shown in the table.

 ≪ Evaluation of state before test >

 1 = a little acne 2 = a little acne 3 = acne severe

 ≪ Determination of acne effect &

 -: Almost no effect, +: slight effect, ++: Much mitigated, +++: Completely healed

Treatment concentration part Pre-test condition Acne effect determination Effect after 2 weeks Effect after 4 weeks 2 % anemone Taurus Subject 1 2 + + Subject 2 One + ++ Subject 3 One - + Subject 4 2 - - Subject 5 2 + ++ Subject 6 2 + + Subject 7 3 ++ +++ Subject 8 2 - ++ Subject 9 3 + + Subject 10 2 + ++ Peony Taurus Subject 1 2 + ++ Subject 2 One + + Subject 3 One ++ +++ Subject 4 2 - ++ Subject 5 2 + + Subject 6 2 + ++ Subject 7 3 + + Subject 8 2 + ++ Subject 9 3 - + Subject 10 2 - - hepatica Taurus Subject 1 2 + ++ Subject 2 One + + Subject 3 One + ++ Subject 4 2 + ++ Subject 5 2 - + Subject 6 2 - - Subject 7 3 + ++ Subject 8 2 + + Subject 9 3 + ++ Subject 10 2 + ++ pasqueflower Taurus Subject 1 2 + ++ Subject 2 One + + Subject 3 One ++ +++ Subject 4 2 - ++ Subject 5 2 + + Subject 6 2 + ++ Subject 7 3 + + Subject 8 2 + ++ Subject 9 3 - + Subject 10 2 - - Lotus Taurus Subject 1 2 + + Subject 2 One + ++ Subject 3 One - + Subject 4 2 - - Subject 5 2 + ++ Subject 6 2 + + Subject 7 3 ++ +++ Subject 8 2 - ++ Subject 9 3 + + Subject 10 2 + ++ Christmas Rose Taurus Subject 1 2 + ++ Subject 2 One + + Subject 3 One ++ +++ Subject 4 2 - ++ Subject 5 2 + + Subject 6 2 + ++ Subject 7 3 + + Subject 8 2 + ++ Subject 9 3 - + Subject 10 2 - - Peony flower Taurus Subject 1 2 + + Subject 2 One + ++ Subject 3 One + + Subject 4 2 + ++ Subject 5 2 - + Subject 6 2 - - Subject 7 3 + + Subject 8 2 + ++ Subject 9 3 + + Subject 10 2 + ++ Training Taurus Subject 1 2 + + Subject 2 One + ++ Subject 3 One - + Subject 4 2 - - Subject 5 2 + ++ Subject 6 2 + + Subject 7 3 ++ +++ Subject 8 2 - ++ Subject 9 3 + + Subject 10 2 + ++

As can be seen from the above table, the improvement effect of acne was confirmed in the majority of subjects after using the soft lotion containing the extract of 2% parsley herb lyocellular cell culture for 4 weeks. In addition, 200 μM methyl jasmonic acid inducer treatment and high frequency cell culture were performed for 3 days at a frequency of 240 to 270 kHz for 5 days, , The extract of Tacrolimus cell culture showed an antimicrobial effect twice as much as that of the untreated case.

Preparation of cosmetic composition

Cosmetic products of the emulsified form such as nutritional lotion, cream, essence, etc., and solubilized form such as softened longevity were prepared with the cosmetic product containing the extract of Laotianaceae cell culture extract of Ranunculaceae of the present invention as an active ingredient.

Manufacturing example  2-1: Lotion

According to the following formulation, it was prepared according to the conventional lotion preparation method.

Raw material name Weight% (w / w) glycerin 5.0 Dipropylene glycol 3.0 Hyaluronic acid 0.5 Polyoxyethylene hardened castor oil 0.1 Polyethylene oleyl ethyl 0.1 ethanol 5.0 antiseptic 0.15 Beauty Suitable amount flash Suitable amount Extract of Ranunculus Thunb 2.0 Purified water to 100

Manufacturing example  2-2: Essence

According to the following prescription, it was prepared according to a conventional method for producing essence.

Raw material name Weight% (w / w) Cetostearyl alcohol 1.0 Self emulsifying monostearic acid 1.0 Wax 0.5 Squalane 5.0 Isocetyl octanoate 3.0 Dimethylsiloxane 0.3 Sorbitan monostearate 0.5 Polyethylene glycol monostearate 8.0 glycerin 4.0 Propylene glycol 0.2 Carboxy polymer 0.22 Triethanolamine 0.25 antiseptic Suitable amount Spices Suitable amount flash Suitable amount Extract of Ranunculus Thunb 7.0 Purified water to 100

Manufacturing example  2-3: Lotion

According to the following formulation, it was prepared according to a conventional lotion preparation method.

Raw material name Weight% (w / w) Cetostearyl alcohol 0.8 Self emulsifying monostearic acid 1.0 Wax 0.5 Stearic acid 0.5 Liquid paraffin 7.0 Squalane 5.0 Macadamia oil 3.0 Isocetyl octanoate 2.0 Dimethylsiloxane 0.3 Sorbitan monostearate 0.5 Polyethylene glycol monostearate 1.2 glycerin 4.0 Propylene glycol 4.0 Betaine 4.0 Carboxy polymer 0.12 Triethanolamine 0.15 antiseptic 0.25 Spices Suitable amount flash Suitable amount Extract of Ranunculus Thunb 5.0 Purified water to 100

Manufacturing example  2-4: Cream

According to the following formulation, it was prepared according to a conventional cream production method.

Raw material name Weight% (w / w) Cetostearyl alcohol 3.0 Self emulsifying monostearic acid 1.5 Chin type monostearate 1.5 Wax 0.5 Liquid paraffin 8.0 Squalane 7.0 Isocetyl octanoate 4.0 Refined jojoba oil 4.0 Dimethylsiloxane 0.3 Sorbitan monostearate 1.0 Polyethylene glycol monostearate 1.2 glycerin 6.0 Propylene glycol 4.0 Betaine 4.0 Xanthan gum 0.06 Triethanolamine 0.10 antiseptic 0.25 Spices Suitable amount flash Suitable amount Extract of Ranunculus Thunb 7.0 Purified water to 100

Manufacturing example  2-5: Gel

According to the following prescription, it was prepared according to a conventional gel preparation method.

Raw material name Weight% (w / w) glycerin 4.0 Propylene glycol 4.0 ethanol 10 Polyoxyethylene hardened castor oil 0.1 Carboxy polymer 0.30 Triethanolamine 0.30 antiseptic Suitable amount Spices Suitable amount flash Suitable amount Extract of Ranunculus Thunb 1.0 Purified water to 100

While the present invention has been particularly shown and described with reference to specific embodiments thereof, those skilled in the art will appreciate that such specific embodiments are merely preferred embodiments and that the scope of the invention is not limited thereby. It will be obvious. Accordingly, the actual scope of the present invention will be defined by the appended claims and their equivalents.

Claims (9)

A cosmetic composition for skin improvement comprising a cultured cell of a plant of the order Pulsatilla koreana (Pulsatilla koreana) or an extract thereof.
The composition according to claim 1, wherein the skin improving agent is wrinkle preventing or improving agent.
The composition according to claim 1, wherein the skin improving agent is a salt solution.
The composition according to claim 1, wherein the skin improving agent is for shrinking pores.
The composition according to claim 1, wherein the skin improving agent is a composition for suppressing sebum secretion.
The composition according to claim 1, wherein the skin improving agent is for improving acne.
The composition according to claim 1, wherein the skin improving agent is a whitening agent.
The composition according to claim 1, wherein the skin-improving composition is an antibacterial composition.
The composition according to claim 1, wherein the skin-improving agent is a skin-moisturizing agent.