KR100580858B1 - A cosmetic composition containing mixing extract of Shipjangsaeng-related natural substancePinus sylvestris, Ganoderma lucidum Karst., blueberry, Cervus elaphus L., Amyda sinensis, Germanum, Apis mellifera L., Panax ginseng C.A.Meyer, salmon egg and Citrus Medica Limonum and the method of manufacturing thereof - Google Patents

Abstract

The present invention relates to ten natural products related to ten longevity: Pinus sylvestris , Ganoderma lucidum Karst. , Cervus elaphus L. , Panax ginseng CA Meyer , Citrus Medus Limonum ), Amyda sinensis , Germanum , Royal jelly ( Apis mellifera L. ), Vaccinium Vitis-Idaea and Onchorhynchus pets), Pinus sylvestris , Ganoderma lucidum Karst. , Cervus elaphus L. , red ginseng ( Panax ginseng CA Meyer ) and lemon ( Citrus Medica Limonum ) are extracted by supercritical fluid extraction technique and then mixed with the rest of the natural products to reduce wrinkles, And to provide a cosmetic composition having anti-aging effect and the like.

Supercritical Fluid Extraction Technique * Octopus * Wrinkle Reduction * Elasticity Enhancement * Moisture Recovery * Anti-Aging * Cosmetic Composition

Description

TECHNICAL FIELD The present invention relates to a cosmetic composition containing a natural product related to ten longevity (Lycopersicon esculentum, mushrooms, bergamot, antler, jar, germanium, royal jelly, red ginseng, salmon roe and lemon) and the method of manufacture thereof. The present invention also relates to a method for the manufacture of a medicament for the treatment and / or prophylaxis of gastrointestinal disorders,

FIG. 1 shows the skin elasticity test results of the nutritional cream containing the natural mixed extract of the present invention of the present invention.

The present invention relates to ten natural products related to ten longevity: Pinus sylvestris , Ganoderma lucidum Karst. , Cervus elaphus L. , Panax ginseng CA Meyer , Citrus Medus Limonum ), Amyda sinensis , Germanum , Royal jelly ( Apis mellifera L. ), Vaccinium Vitis-Idaea and Onchorhynchus pets), Pinus sylvestris , Ganoderma lucidum Karst. , Cervus elaphus L. , red ginseng ( Panax ginseng CA Meyer ) and lemon ( Citrus Medica Limonum ) were obtained by supercritical fluid extraction technique and mixed with the remaining mixed extracts and applied to cosmetics Thereby improving cosmetic composition such as anti-wrinkle, elasticity-enhancing, and moisturizing-restoring action.

As the largest organ of our body, which occupies about 16% of our total body volume, the skin acts as an important barrier to protect our body from the many deadly harmful factors that directly touch the outside environment and invade the body have.

However, as our skin grows older, it becomes damaged due to various pollutants and stresses, which causes damage to the cells and cell proliferation, resulting in loss of elasticity and wrinkles. Originally, normal healthy skin has built up a defense system to protect the skin from these external damage factors, but by sustained external stimuli these defense systems are collapsed, resulting in loss of protective ability of the skin, The skin elasticity is reduced, and the skin aging phenomenon such as wrinkle generation occurs.

In addition, loss of moisturizing ability and turn over function of the skin is a kind of phenomenon caused by aging of the skin, which is associated with wrinkle formation and skin elasticity reduction. As the age increases, the amount of collagen, mucopolysaccharide and hyaluronic acid in the matrix, which plays an important role in the elasticity of the skin, decreases and the amount of insoluble collagen increases, It becomes difficult to maintain moisture, and the skin becomes rough. In addition, sebaceous glands and sweat glands decrease in function, sebum and sweat are gradually reduced, sebaceous membranes are not properly formed, and the protective ability for skin moisturization is lost. In the end, the skin becomes dry, translucence, And the elasticity of the skin protein is reduced and wrinkles are produced. In other words, the loss of moisture in the skin is a major symptom of aging, together with the reduction of skin elasticity and wrinkle formation, and is closely related.

In addition, it is known that wrinkle formation is caused by the production of reactive oxygen species, which is the main cause of skin damage when human skin cells are exposed to ultraviolet rays (UV-B and UV-A).

Accordingly, there is a need to develop cosmetics that can protect skin cells from the external environment such as ultraviolet rays, activate cellular proliferative activity, and impart smooth moisturizing power. The inventors of the present invention have found that 10 natural products To obtain a cosmetic composition capable of meeting such a need.

A conventional technique for extracting these ten effective components is as follows.

The mixed extract is added to a solvent such as butylene glycol or propylene glycol and then extracted by heating at 50 to 95 ° C for 3 to 20 hours in a distillation apparatus equipped with a cooling condenser so as to prevent evaporation of the solvent, And immersed at 5 to 37 DEG C for 1 to 15 days to extract an active ingredient.

Also, since the same effect is obtained when each of the natural product extracts is extracted and then mixed, the same method is also applicable. The solvent is evaporated using a distillation apparatus equipped with a cooling condenser, and the above-mentioned five natural product mixed extracts are completely concentrated under reduced pressure to obtain an extract.

However, such a general solvent extraction method has a problem that the natural active ingredient is destroyed due to evaporation, pyrolysis and oxidation because it is heated at a high temperature.

Therefore, the inventors of the present invention have found that, among the ten substances that have been proven through long clinical experience, the extracts obtained by extracting the five components through the supercritical fluid extraction method and then mixed with the remaining mixed extracts are compared with the mixed extracts obtained by extracting all the 10 substances by the general solvent extraction method It has been confirmed that it has excellent effects on alleviation of skin wrinkles, elasticity enhancement and moisturizing ability through cytoprotective effect from ultraviolet rays, cell proliferation ability and excellent moisturizing ability. When it is introduced into cosmetics, it is more effective in relieving wrinkles, improving elasticity, And the present invention has been completed.

Accordingly, an object of the present invention is to provide a natural mixture of tenacious livestock by preparing the extract by extracting with a supercritical fluid extraction method, extracting ginseng, Ganoderma lucidum, antler, lemon and red ginseng, and then mixing the extract with a citrus tree, Zara, germanium, royal jelly and salmon extract will be.

It is also an object of the present invention to provide a method of extracting a part of the above-mentioned complex-related mixed extracts through a supercritical fluid extraction method, thereby reducing skin wrinkles, elasticity enhancement and moisturizing through the protection of cells from ultraviolet rays, The present invention provides a cosmetic composition which is superior in safety and stability in that it can be more advanced when extracting all of the above 10 kinds of extracts by a general solvent extraction method.

In order to achieve the above object, the cosmetic composition according to the present invention is characterized in that a mixed extract of 10 natural products related to ten luteal, i.e., extract, gruel, mushroom, bilberry, antler, jara, germanium, royal jelly, red ginseng, salmon roe and lemon And 0.1 to 10% by dry weight based on the total weight of the composition.

Preferably, the dry weight ratio of each of the 10 extracts to the mixed extract is 5-15%, 10-15%, 15-15%, 15-15%, 5-15% of red ginseng, 10-15% 15%, bilberry 1 ~ 5%, jara 15 ~ 20%, germanium 0.0001 ~ 0.0003%, royal jelly 10 ~ 15% and salmon egg 3 ~ 10%.

In order to overcome the aging, wrinkling, elasticity and moisturizing of human skin as the skin age, the present inventors have been studying about ten longevity which means longevity and eternal life. In other words, ten kinds of ten kinds of ten kinds of material consisting of pine, firefly, crane, deer, turtle, rock, water, mountain, sea and cloud are expressing long life immortality which did not die forever, I think this is related to aging.

In the present invention, the natural products associated with the tenacious lice were Lycoptera, Ganoderma lucidum, Bilberry, Deer, Zara, Germanium, Royal jelly, Red ginseng, Salmon roe and Lemon. For pine, deer and turtles that can be directly obtained from these livestock, pine trees, antler and jar are used as such, and those that do not exist or are transmitted only to images are selected as natural products to which the efficacy can be coped. In other words, in the sense that it does not grow old, it is used for the prevention of wrinkles and strengthening of immune component, and it was used as a manure. Symbol of nobility and cleanliness, In addition, germanium has been selected to enhance the skin resistance as a means of strengthening and firmness of rocks. The moisturizing effect of water can be shown by using royal jelly which has moisturizing and elasticity effect, and provides the function of resting and vitality Red ginseng, known to have antioxidant and antioxidant properties by taking, can combine the vitality of the sun with intense energy or cell regeneration. It can be used to stimulate the circulatory function of the cloud, Lemon was selected for association with keratinization. The above ten natural products are natural products having different efficacy and are related to the tenacious. The present inventors extracted the five natural products from the natural products using the supercritical fluid extraction method, And effectively extracted and mixed with the remaining 5 natural products. Thus, it was confirmed that they effectively exhibited cell protection action, cell proliferation action and moisturizing action from ultraviolet rays.

As a result, cosmetic compositions containing the above-mentioned complex mixture of the herbal extracts were prepared and directly used on human skin. As a result, compared to conventional wrinkle-preventing and moisturizing cosmetic preparations, the cosmetic composition was improved in skin elasticity, wrinkle, The present invention has been accomplished on the basis of confirming that it is not only excellent in skin stability and safety as well as providing an anti-aging effect by the contribution of the supercritical fluid extraction method.

The method for preparing the natural extract according to the present invention, that is, the extract mixture of ginseng, Ganoderma lucidum, lemon, red ginseng, antler, jara, germanium, royal jelly, salmon roe,

1) mixing, grinding, mixing, and stirring a mixture of the tenant-associated ingredients with the solvent, ginger mushroom, antler, lemon and red ginseng;

2) placing the mixture in a supercritical water extraction tank and supplying supercritical fluid;

3) maintaining the pressure of the extraction tank at 330 atm and raising the temperature in the extraction tank to 42 ° C at a rate of 0.5 ° C / min;

4) maintaining the temperature and pressure of the extraction tank at 30 to 50 ° C, 250 to 400 atm for 50 minutes, and then reducing the pressure to normal pressure to obtain the first mixed extract;

5) extracting the second mixed extract with water, an organic solvent or a mixed solvent thereof, among the tenant-related mixed ingredients, extracting the zeolites, germanium, royal jelly, salmon roe and bilberry; And

6) mixing the first mixed extract and the second mixed extract at a weight ratio of 6.5: 3.5;

.

First, the leaves, gingko mushrooms, antler, lemon and red ginseng were extracted with supercritical fluid extraction method. Supercritical fluids are defined as "fluids above critical temperature and pressure" and have unique properties that distinguish them from conventional solvent extraction methods. When a material in a supercritical state changes its pressure near its critical point, many physical properties such as density, viscosity, diffusion coefficient, and polarity continuously change greatly from gas close to liquid state. When the liquid carbon dioxide is put into a closed container and heated, the boundary of the gas - liquid gradually disappears as temperature and pressure increase. Supercritical fluid is a heavy fluid that fills an enclosed space like a gas. Supercritical fluid technology is a technique that exploits the advantages of supercritical fluids, such as high solubility, rapid mass transfer and heat transfer, low viscosity, high diffusion coefficient, and fast penetration into micropores due to low surface tension.

The supercritical fluid extraction method according to the present invention uses carbon dioxide as the supercritical fluid and the extraction process is as follows:

1) Mixing, grinding, mixing and stirring with a solvent, ginger mushroom, antler, lemon and red ginseng;

2) placing the mixture in a supercritical water extraction tank and supplying supercritical fluid;

3) maintaining the pressure of the extraction tank at 330 atm and raising the temperature in the extraction tank to 42 ° C at a rate of 0.5 ° C / min; And

4) maintaining the temperature and pressure of the extraction bath at 30 to 50 DEG C for 50 minutes at 250 to 400 atmospheres, and then reducing the pressure to normal pressure to obtain a mixed extract.

By the extraction in the supercritical condition as described above, not only the effective ingredient can be effectively obtained but also each effective ingredient is effectively expressed after extraction.

The mixture of the extracts of the leaves, gentian mushrooms, antler, lemon and red ginseng according to the present invention is prepared and then mixed with the extracts obtained through the general extraction method of germanium, royal jelly, salmon roe, and cinnabar.

 The extraction method using the general extraction method of Jara, germanium, royal jelly, salmon roe and bilberry is as follows.

A mixture obtained by mixing dry weight ratio of 1 to 5% by weight of cinnabar and 15 to 20% by weight of ginger, 0.0001 to 0.0003% by weight of germanium, 10 to 15% by weight of royal jelly and 3 to 10% At least one solvent selected from the group consisting of water, ethanol, methanol, butanol, ethyl acetate, butyleneglycol, propylene glycol and glycerin is added as an extraction solvent in an amount of 5 to 15 times the volume of the mixed extract. As the extraction method, extraction is carried out by heating at 50 to 95 ° C for 3 to 20 hours in a state where a cooling condenser is installed to prevent the solvent from evaporating, or by dipping at 5 to 37 ° C for 1 to 15 days to extract the active ingredient Is available. Also, since the same effect is obtained when each of the natural product extracts is extracted and then mixed, the same method is also applicable. The solvent mixture evaporated by using a distillation apparatus equipped with a cooling condenser is recovered, and the natural product mixed extract is completely concentrated under reduced pressure from bergamot, Jara, germanium, royal jelly and salmon roe, and the concentration thereof is 0.01 to 9% Is taken from 0.3 to 6% and mixed with 5 natural mixed extracts extracted by supercritical fluid extraction method.

The above supercritical fluid extraction method will be described in detail as follows.

Butyleneglycol and ethanol were mixed, and the content of each was 62.7% by weight of the pulverized natural product, 33.9% by weight of butylene glycol, and 3.4% by weight of ethanol %, And stirred for 1 hour.

The mixture is introduced into a supercritical extraction tank and carbon dioxide is supplied to the extraction tank using a high pressure gas pump to increase the pressure. When the pressure of the extraction tank reaches 330 atmospheres, the supply of carbon dioxide is stopped. At this time, the temperature inside the extraction tank is similar to room temperature because the long pipe (1m) is connected between the pressurization pump and the extraction tank, so that the temperature near the pressurization pump is warmed up but the temperature in the extraction tank does not change.

Using a temperature controller, the temperature inside the extraction tank is raised to 42 ° C at a rate of 0.5 ° C / min using a hot wire wrapping the outside of the extraction tank. The pressure of the extraction tank due to the elevated temperature is partially discharged by the pressure regulator attached to the outlet of the extraction tank and the pressure is maintained at 330 atm.

The temperature and pressure of the extraction bath are maintained at 42 ° C and 330 atmospheres for 50 minutes. The high-pressure gas pump is restarted, and carbon dioxide is continuously supplied to the extraction tank at a flow rate of 20 L / min. At the same time, the pressure regulator attached to the exit side is used to release the extract and keep the temperature and pressure of the extraction tank constant. Emissions discharged through the pressure regulator are transferred to solid dry ice due to pressure and temperature drop. This may cause clogging of the tube on the final outlet side. To prevent this, keep the temperature of the pressure regulator at 70 ℃.

The extracted extract is decompressed at atmospheric pressure to release carbon dioxide into the atmosphere, and the active ingredient extract of five mixtures and butylene glycol are recovered together into a trap. The above procedure is carried out for 12 hours to obtain the extract and the extraction operation is terminated.

The preferred dry weight ratio of the extracts to the five mixed extracts by supercritical fluid extraction is 5-15% by weight of ginseng extract, 10-15% by weight of ganoderma lucidum, 15-25% by weight of antler extract, 15% by weight and lemon by 10-15% by weight, and the dry weight ratio of the remaining five extracts extracted by the general extraction method is 1 to 5% by weight of cinnabar and 15 to 20% by weight of ganoderma, germanium 0.0001 to 0.0003% by weight, 10-15% by weight of royal jelly, and 3 to 10% of salmon roe. The mixing ratio of the two mixed extracts is suitably 6.5: 3.5.

In addition, the present invention relates to a cosmetic composition containing a combination of a supernatant fluid extraction method and a supernatant fluid extraction method. By introducing the above-mentioned supernatant-related mixed extract into the skin, they can protect the skin from ultraviolet rays, The present invention relates to a cosmetic composition which is superior to that obtained by a general solvent extraction method.

The cosmetic composition according to the present invention is preferably contained in an amount of 0.1 to 10% by dry weight, preferably 0.5 to 8% by dry weight, based on the total composition, , 2 hours).

When the content of the complex-associated extracts is less than 0.1% by dry weight, it is not preferable to show effectiveness, and when the content is more than 10% by dry weight, there is a problem in formulation, and the optimum concentration considering the efficacy and stability is 0.5 to 8 Weight%.

Hereinafter, the constitution and effects of the present invention will be described in more detail through examples and test examples. It will be apparent, however, to those skilled in the art that these embodiments are for the purpose of illustrating the present invention and that the scope of the present invention is not limited to these embodiments.

[Example 1] Production of mixed extracts of tenosynovis

First, 370 g of a mixture obtained by mixing 85 g of germ oil, 57 g of Ganoderma lucidum, 85 g of antler, 74 g of red ginseng and 69 g of lemon was pulverized and mixed with 200 g of butylene glycol and 20 g of ethanol. The mixture was stirred and allowed to stand for 1 hour.

The mixture was introduced into a supercritical extraction tank, and carbon dioxide was supplied to the extraction tank by using a high pressure gas pump. When the pressure reached 330 atm, the supply of carbon dioxide was stopped. The temperature inside the extraction tank was raised to 42 ° C at a rate of 0.5 ° C / min through a hot wire wrapping the outside of the extraction tank using a temperature controller. The temperature and pressure of the extraction tank were kept at 42 캜 and 330 50 for 50 minutes while partially discharging the carbon dioxide using a pressure regulator attached to the outlet of the extraction tank. Then, the high-pressure gas pump was restarted, and carbon dioxide was continuously supplied to the extraction tank at a flow rate of 20 L / min. At the same time, the extract was released using a pressure regulator attached to the outlet side, and the temperature and pressure of the extraction bath were kept constant at 42 ° C and 330 atm. While the temperature of the pressure regulator was maintained at 70 ° C, the effluent discharged through the pressure regulator was transferred to solid dry ice due to pressure and temperature drop. The extracted extract was decompressed to normal pressure, carbon dioxide was released into the atmosphere, and the active ingredient extract of butanol - related mixture and butylene glycol were collected together into a trap. The above procedure was carried out for 12 hours to obtain the extract, and the extraction operation was terminated to obtain 29.5 g (dry weight) of the natural extract.

In addition, 200 g of the mixed raw material obtained by crushing 17.14 g of fully dried squirrel, 97.12 g of Jara, 0.02 g of germanium, 57.14 g of royal jelly and 28.58 g of salmon roe was added to 2 kg of water, and the mixture was boiled for 3 hours in an extractor with a cooling condenser. Filtered with a mesh filter cloth and cooled to room temperature. After aging at 5-15 ° C for 7 days, the mixture was filtered through filter paper No. 2 of Wattsman. The filtrate was concentrated under reduced pressure to 70 캜 in a distillation apparatus equipped with a cooling condenser to obtain 15.88 g (dry weight) of a natural product extract.

The two mixtures thus obtained were mixed at a ratio of 6.5 (mixture obtained through supercritical fluid extraction): 3.5 (mixture obtained through general solvent extraction) to prepare a natural mixture of tenacity.

[Example 2] Preparation of mixed extracts of tenosynovis

Using the same method as in Example 1 except that butylene glycol and ethanol were not used together as a solvent for supercritical fluid extraction in Example 1, butylene glycol alone was used, supercritical fluid 26g (dry weight) of the natural extract was obtained by the extraction method, which was mixed with the remaining mixed extract at a ratio of 6.5: 3.5.

[Example 3] Preparation of mixed extracts of tenosynovis

In the same manner as in Example 1 except that butylene glycol and water were used instead of butylene glycol and ethanol as the solvents of the supercritical fluid extraction method in Example 1, 27g (dry weight) of natural extract was obtained and mixed with the other mixed extracts at a ratio of 6.5: 3.5.

[Example 4] Preparation of mixed extracts of tenosynovis

The same procedure as in Example 1 was repeated except that ethanol and water were used instead of butylene glycol and ethanol as solvents for the supercritical fluid extraction in Example 1, Dry weight), which was mixed with the remaining mixed extract at a ratio of 6.5: 3.5.

[Example 5] Preparation of mixed extracts of tenosynovis

23 g (dry weight) of a natural product extract was obtained in the same manner as in Example 1, except that butylene glycol used as a solvent for supercritical fluid extraction in Example 1 and ethanol were used alone instead of ethanol , And this was mixed with the remaining mixed extract at a ratio of 6.5: 3.5.

[Example 6] Production of mixed extracts of tenosynovis

24 g (dry weight) of a natural product extract was obtained in the same manner as in Example 1, except that water alone was used instead of butylene glycol and ethanol as a solvent for supercritical fluid extraction in Example 1 , And this was mixed with the remaining mixed extract at a ratio of 6.5: 3.5.

[Comparative Example 1]

370 g of a mixture obtained by pulverizing a mixture of 85 g of gherkin, 57 g of Ganoderma lucidum, 85 g of antler, 74 g of red ginseng, and 69 g of lemon, 17.14 g of mulberry, 97.14 g of geranium, 0.00058 g of germanium, 57.14 g of royal jelly, 3.7 kg of coke and 300 g of ethanol, stirred for 10 hours, and then heated at 95 ° C for 5 hours in a distillation apparatus equipped with a cooling condenser to obtain 10.5 g (dry weight) of the extract.

[Comparative Example 2]

9.4 g (dry weight) of a natural product extract was obtained in the same manner as in Comparative Example 1, except that butylene glycol and ethanol were not used as a solvent in Comparative Example 1 but only butylene glycol was used alone. ≪ / RTI >

[Comparative Example 3]

8.7 g (dry weight) of a natural product extract was obtained in the same manner as in Comparative Example 1, except that butylene glycol and water were used instead of butylene glycol and ethanol as the solvent in Comparative Example 1.

[Comparative Example 4]

7.5 g (dry weight) of a natural product extract was obtained in the same manner as in Comparative Example 1, except that ethanol and water were used instead of butylene glycol and ethanol as solvents in the above Comparative Example 1.

[Comparative Example 5]

The same procedure as in Comparative Example 1 was carried out except that butylene glycol used as the solvent in Comparative Example 1 and ethanol were used alone and the temperature of the heating condition was changed to 60 ° C. Weight).

[Comparative Example 6]

8.2 g (dry weight) of a natural product extract was obtained in the same manner as in Comparative Example 1, except that only water was used instead of butylene glycol and ethanol as a solvent in Comparative Example 1.

[Test Example 1] Test for protecting cells from ultraviolet rays

1) Test method

Human fibroblasts were placed in a well plate at 2 × 10 ⁵ cells / well, 0.2 ml of a DMEM medium containing 10% FBS was supplied, and then cultured in a CO ₂ incubator for 24 hours. Each of the natural product mixed extracts of Examples 1 to 6 and Comparative Examples 1 to 6 dissolved in DMSO was treated and irradiated with ultraviolet rays of 2 MED for 5 minutes and cultured for 24 hours. MTT assays were performed to determine cell viability. The MTT assay was performed by adding 0.1 ml of MTT solution to the cultured cells for 4 hours, removing the medium, adding 0.5 ml of DMSO, and measuring formazan formed at 570 nm using an ELISA reader Respectively. In addition, the absorbance of each extract was tested with DMSO, a solvent in which the solid content of the extract was dissolved, as a control, and the cytoprotective effect was determined from the difference in absorbance relative to the control. On the other hand, for comparison, the effect of green tea extract, which is a representative raw material of skin aging prevention, was measured.

2) Test result

The cytoprotective effect of the extracts obtained in Examples 1 to 6 and Comparative Examples 1 to 6 on UV light was measured according to the above method, and the results are shown in Table 1. From the above results, it can be seen that Examples 1 to 6 of the present invention are significantly superior to the extracts of Comparative Examples 1 to 6 prepared by the conventional extraction method and the green tea extract, which is a representative raw material for preventing skin aging, there was.

Cell Protection Effect from Ultraviolet (%) Example Comparative Example Green tea extract division effect division effect Extraction solvent effect Example 1 91 Comparative Example 1 71 Butylene glycol / ethanol 68 Example 2 99 Comparative Example 2 85 Butylene glycol 79 Example 3 94 Comparative Example 3 79 Butylene glycol / water 70 Example 4 90 Comparative Example 4 73 Ethanol / water 62 Example 5 74 Comparative Example 5 45 ethanol 42 Example 6 83 Comparative Example 6 59 water 46 The concentration of the sample contained in the reaction solution is 100 μg / ml. Cell protection effect (%) = absorbance of the test group / absorbance of the control group × 100 The value in parentheses is the absorbance (nm). For reference, the absorbance of the control group was 1.0449 nm

[Test Example 2] Test of cell protection effect from superoxide radical

1) Test method

Human fibroblasts were placed in a well plate at 2 × 10 ⁵ cells / well, 0.2 ml of a DMEM medium containing 10% FBS was supplied, and then cultured in a CO ₂ incubator for 24 hours. Each of the natural product mixed extracts of Examples 1 to 6 and Comparative Examples 1 to 6 dissolved in DMSO was treated and cultured for 30 minutes. Superoxide radicals were generated by adding 10 μl of 1.5 mM xanthine and 10 μl of 0.25 U xanthine oxidase to the cultured cells, and then cultured in a CO ₂ incubator for 24 hours. MTT assays were performed to determine cell viability. The MTT assay was performed by adding 0.1 ml of MTT solution to the cultured cells, culturing the cells for 4 hours, removing the medium, adding 0.5 ml of DMSO, and measuring the absorbance at 570 nm using an ELISA reader. In addition, the absorbance of each extract was tested with DMSO, a solvent in which the solid content of the extract was dissolved, as a control, and the cytoprotective effect was determined from the difference in absorbance relative to the control. On the other hand, for comparison, the effect of green tea extract, which is a representative raw material of skin aging prevention, was measured.

2) Test result

According to the above method, the cytoprotective effect of the extracts extracted from Examples 1 to 6 and Comparative Examples 1 to 6 on the superoxide radical was measured. The results are shown in Table 2. From the above results, it can be seen that Examples 1 to 6 according to the present invention are superior to the extracts of Comparative Examples 1 to 6 prepared by the conventional extraction method and the green tea extract, which is a representative raw material for preventing skin aging, .

 Cell Protection Effect from Superoxide Radical (%) Example Comparative Example Green tea extract division effect division effect Extraction solvent effect Example 1 80 Comparative Example 1 73 Butylene glycol / ethanol 69 Example 2 99 Comparative Example 2 91 Butylene glycol 82 Example 3 97 Comparative Example 3 85 Butylene glycol / water 77 Example 4 95 Comparative Example 4 81 Ethanol / water 76 Example 5 87 Comparative Example 5 63 ethanol 60 Example 6 89 Comparative Example 6 79 water 66 The concentration of the sample contained in the reaction solution is 100 μg / ml. Cell protection effect (%) = absorbance of the test group / absorbance of the control group × 100 The value in parentheses is the absorbance (nm). For reference, the absorbance of the control group was 1.0449 nm

[Test Example 3] Cell proliferation effect test

1) Test method

100 μl of a 5% fetal bovine serum (FBS) solution was added to a 96 μ-well plate, and DMEM medium containing 2% FBS was added to each of the natural product mixed extracts of Examples 1 to 6 and Comparative Examples 1 to 6 Was diluted to a constant concentration and diluted 100 쨉 l each time. Using a hemocytometer, fibroblasts from the foreskin tissue of newborns were dispersed in 100 μl of 4,000 cells per well, and then cultured for 7 days. After the incubation, the supernatant was removed, and then 200 μl of 5% phosphate buffered saline was added thereto. The resultant was washed with DMSO (150 μl / well) for 10 minutes, and absorbance was measured at 570 nm. The absorbance of each extract was measured with DMSO, a solvent in which the solid content of the extract was dissolved, as a control, and the cell proliferation effect was determined from the difference in absorbance relative to the control. On the other hand, for comparison, the effect of green tea extract, which is a representative raw material of skin aging prevention, was measured.

2) Test result

According to the above method, the cell proliferation effects of the extracts obtained in Examples 1 to 6 and Comparative Examples 1 to 6 were measured, and the results are shown in Table 3. From the above results, it can be seen that Examples 1 to 6 according to the present invention are superior to the extracts of Comparative Examples 1 to 6 prepared by the conventional extraction method and the green tea extract, which is a representative raw material for preventing skin aging, .

Cell proliferation effect (%) Example Comparative Example Green tea extract division effect division effect Extraction solvent effect Example 1 150 Comparative Example 1 103 Butylene glycol / ethanol 101 Example 2 140 Comparative Example 2 127 Butylene glycol 109 Example 3 142 Comparative Example 3 111 Butylene glycol / water 110 Example 4 130 Comparative Example 4 121 Ethanol / water 115 Example 5 125 Comparative Example 5 102 ethanol 102 Example 6 130 Comparative Example 6 113 water 104 The concentration of the sample contained in the reaction solution is 100 μg / ml. Cell proliferation effect (%) = absorbance of the test group / absorbance of the control group × 100 The value in parentheses is the absorbance (nm). For reference, the absorbance of the control group was 1.2214 nm

[Test Example 4] Moisturizing effect test

1) Test method

5 g of the natural product mixed extract of Examples 1 to 6 was placed in a dish and allowed to stand under the conditions of a relative humidity of 41% and a temperature of 25 캜 in a desiccator containing CaCl ₂ , Were measured with time. As a comparative example, 1,3-butylene glycol, which is generally used in moisturizing cosmetics, was used.

2) Test result

According to the method described above, the moisturizing effect of the herbal mixed extract of the present invention was measured, and the results are shown in Table 4. From the above results, it was confirmed that the extract of the present invention has a high moisture content due to moisturizing, and that the effect is even better when compared with 1,3-butylene glycol, which is known as a moisturizing substance.

division Measurement time (hr) Moisture content (%) Example 1 120 57.2 Example 2 120 63.5 Example 3 120 60.7 Example 4 120 57.7 Example 5 120 55.5 Example 6 120 59.3 1,3-butylene glycol 120 43.2 Water content (%) = Residual sample amount (g) / Amount of initial sample (g) × 100

[Test Example 5] Safety Test of Natural Mixed Extract of Octopus

In order to test the safety of the natural mixed extract of Echinochloa liquor prepared in Example 1, a first skin irritation test, a first eye mucosal irritation test and a skin sensitization test were carried out on experimental rats, and the results are shown in Tables 5 to 7 (Testing Period: April 14, 2004 to July 20, 2004, Testing Institution: Biorand Biotechnology Research Institute). Table 5 below shows the results of the primary skin irritation test.

time Hong Ban  Minor species 30-60 minutes 0 0 24 hours 0 0 48 hours 0 0 72 hours 0 0  * Species Species: Albino Rabbit from New Zealand, weight and age at start of administration: 10 weeks, 2.5 kg ± 20 g * Test sample: Example 1

From the above results, it was confirmed that the natural herbal mixed extract of the present invention is a safe substance without irritation such as skin erythema and edema.

The following Table 6 shows the results of the first mucosal irritation test of the natural mixed extract of Novosome prepared in Example 1.

opinion 1 hours 24 hours 48 hours 72 hours Corneal opacity 0 0 0 0 Iris abnormality 0 0 0 0 Corneal edema 0 0 0 0  * Species Species: Albino Rabbit from New Zealand, weight and age at start of administration: 10 weeks, 2.5 kg ± 20 g * Test sample: Example 1

From the above results, it was confirmed that the natural mixed extract of Chinese herbal products prepared in Example 1 of the present invention is a safe material having no mucosal irritation such as corneal opacity, iris irregularity, or corneal edema.

In addition, the following Table 7 shows the results of the first skin sensitization test of the natural mixed extract of Novosome prepared in Example 1.

time Sensitization rate (%) Rating Sensitivity 24 hours 6 Ⅰ Very weak 48 hours 3 Ⅰ Very weak * Experimental animal species: Hartly male Guinea-Pig, * Weight and age at start of administration: 10 weeks, 300 ~ 500g * Test sample: Example 1

In the above results, the sensitization rate was calculated from the number of animals that showed a positive response to the test and the skin response thereof, which were scored by a conventional method. When the sensitization rate was 0 to 8% And the degree of sensitivity is determined to be very weak. Therefore, it was confirmed that the natural mixed extract of Chinese herbal products prepared in Example 1 of the present invention is a safe substance in skin sensitization.

In addition, in the human skin patch test, all of the reactions up to 30 minutes and 24 hours after the removal of the sample were negative, and no abnormality was observed even on the 7th day after the sample was removed.

Namely, from the above safety test results, it was proved that natural herbal extracts are very safe substances.

[Test Example 6] Measurement of physical stability of a natural mixed extract

In order to evaluate the degree of stability of the physical properties of the cosmetic composition when it was introduced into cosmetics, the natural mixed extract prepared in Example 1 was generally used for measuring physical properties of cosmetic ingredients at room temperature, 42 ° C and daylight The results are shown in Table 8 below.

    Date condition One 2 3 4 5 6 7 8 9 10 11 12 13 14 15 Room temperature - - - - - - - - - - - - - - - 40 ℃ - - - - - - - - - - - - - - - daylight - - - - - - - - - - - - - - -     Date condition 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 Room temperature - - - - - - - - - - - - - - - 40 ℃ - - - - - - - - - - - - - - - daylight - - - - - - - - - - - - - + + -; No color change, +; Start discoloration, ++; Much changed, +++; Complete discoloration * Experimental sample: The cosmetic introduced with Example 1

From the above results, it was proved that the mixed extract of Octagonal Bifidobacterium prepared in Example 1 of the present invention is very stable at room temperature and 40 ° C, and is discolored almost negligible in daylight, but is a very stable substance without further discoloration progression .

[Test Example 7] Elasticity increase test for human skin

The nutritional cream of Formulation Example 1 containing the natural mixed extract of the tenant of Example 1 and the nutrient cream of Comparative Formulation Example 1 containing the seaweed extract which was conventionally used for the cosmetic with the elasticity increasing substance were prepared, On skin elasticity. Among the women aged 20 to 55 years, 20 of the women with normal, oily, dry and complex skin had the nutritional cream of Formulation 1 on the left side of the face and the nutritional cream of Comparative Formulation 1 on the right side of the face. , And applied twice a day in the evening, and the elasticity of the skin was measured using a skin elasticity measuring instrument (Ballisto meter) after 1 week, 2 weeks, 3 weeks and 4 weeks. The results are shown in Table 9 and Fig.

≪ Formulation Example 1 and Comparative Formulation Example 1 >

The nutritional creams of Formulation Example 1 and Comparative Formulation Example 1 used in the present invention were prepared by a conventional method with the following composition.

Raw material Formulation Example 1 Comparative Formulation Example 1   Natural herbal extract (Example 1) 7.0 - Seaweed extract - 7.0 Sitosterol 4.0 4.0 Polyglyceryl 2-oleate 3.0 3.0 Ceramide 0.7 0.7 Setares-4 2.0 2.0 cholesterol 3.0 3.0 Dicetyl phosphate 0.4 0.4 Concentrated glycerin 5.0 5.0 Sunflower oil 22.0 22.0 Carboxyvinyl polymer 0.5 0.5 Triethanolamine 0.5 0.5 antiseptic a very small amount a very small amount Spices a very small amount a very small amount Purified water Balance Balance

Formulation Example 1 Comparative Formulation Example 1 T ₀ T ₁ T ₂ T ₃ T ₄ T ₀ T ₁ T ₂ T ₃ T ₄ Elasticity Popping 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 Amplitude first 1.7 4.2 5.2 6.3 7.1 1.9 3.5 4.4 4.7 5.3 second 1.3 3.4 3.9 5.2 6.0 1.5 1.9 2.7 3.0 3.6 third 0.0 2.0 2.5 3.6 4.3 0.0 0.0 1.9 2.1 2.3 fourth 0.0 0.0 1.3 2.9 3.3 0.0 0.0 0.2 0.3 0.9 Average 0.75 2.4 3.225 4.5 5.175 0.85 1.35 2.3 2.525 3.025 (Note) T _0; One week after, T _2;; before use, and then used T ₁ week 2, T _{3; 3} weeks of use, T _{4; 4} weeks after use

As can be seen from the above results, the nutritional cream of Formulation Example 1 containing the natural mixed extract according to the present invention is superior to the nutritional cream of Comparative Formulation Example 1, which contained the seaweed extract which was previously used as a skin elasticity- And the skin elasticity effect was improved by the improvement.

[Test 8] Wrinkle-improving effect test

The nutritional essence of Formulation Example 2 containing the natural intestinal mixture of Example 1 and the nutritional essence of Comparative Formulation Example 2 containing hyaluronic acid which had been conventionally used for the wrinkle improving cosmetic were compared with those of Formulation 2 To measure the improvement effect, Formulation Example 2 was applied twice a day to 20 left-eye margins and Comparative Example 2 was applied to the right-eye margin. After 45 days of each treatment, elasticity of the skin surface was measured by Cutometer SEM 474. Cutometer The skin elasticity measurement by SEM 474 measures the depth of the wrinkles. The smaller the value, the better the elasticity and the less wrinkles. The elasticity value was expressed as a percentage of the reduced value as compared with the measured value before the treatment, and the average value of 20 subjects was obtained and shown in Table 10.

≪ Formulation Example 2 and Comparative Formulation Example &

The nutritional essence of Formulation Example 2 and Comparative Formulation Example 2 used in the present invention was prepared by a conventional method with the following composition.

Raw material Formulation Example 2 Comparative Formulation Example 2  Natural herbal extract (Example 1) 7.0 - Hyaluronic acid - 7.0 Sitosterol 1.70 1.70 Polyglyceryl 2-oleate 1.50 1.50 Ceramide 0.7 0.7 Setares-4 1.2 1.2 cholesterol 1.5 1.5 Dicetyl phosphate 0.4 0.4 Concentrated glycerin 5.0 5.0 Sunflower oil 15.0 15.0 Carboxyvinyl polymer 0.2 0.2 Xanthan gum 0.2 0.2 antiseptic a very small amount a very small amount Spices a very small amount a very small amount Purified water Balance Balance

sample Skin elasticity (reduced wrinkle depth) Effect (%) 30 days 45 days Formulation Example 2 17.7 29.3 Comparative Formulation Example 2 10.4 21.5

[Test Example 9] Moisturizing effect test

The nutritional lotion of Formulation Example 3, which was formulated with natural herbal extract of Example 1, and the nutritional lotion of Comparative Formulation Example 3 containing 1,3-butyleneglycol, which was conventionally used in moisturizing cosmetics conventionally, , Formulation Example 3 and Comparative Formulation Example 3 were applied to the inside of 20 left arm to test skin moisturizing effect. After each application, the moisturizing power of the skin surface with time was measured by Corneometer CM 825. Corneometer CM 825 is used to measure skin moisture content. The higher the value, the better the moisturizing effect. The average value of 20 subjects was obtained and is shown in Table 11 and FIG. 2 below. The test was carried out at a temperature of 20 ° C and a relative humidity of 20%.

≪ Formulation Example 3 and Comparative Formulation Example 3 >

The nutritional lotions of Formulation Example 3 and Comparative Formulation Example 3 used in the present invention were prepared by a conventional method with the following composition.

Raw material Formulation Example 3 Comparative Formulation Example 3  Natural herbal extract (Example 1) 7.0 - 1,3-butylene glycol - 7.0 Sitosterol 1.70 1.70 Polyglyceryl 2-oleate 1.50 1.50 Ceramide 0.7 0.7 Setares-4 1.2 1.2 cholesterol 1.5 1.5 Dicetyl phosphate 0.4 0.4 Concentrated glycerin 5.0 5.0 Sunflower oil 10.0 10.0 Carboxyvinyl polymer 0.2 0.2 Xanthan gum 0.3 0.3 antiseptic a very small amount a very small amount Spices a very small amount a very small amount Purified water Balance Balance

Time (min) 5 10 15 20 25 30 Formulation Example 3 19.4 7.98 5.88 5.16 4.18 4.46 Comparative Formulation Example 3 20.4 7.62 4.12 3.74 3.62 3.02

Hereinafter, the cosmetic composition containing the natural extract of the present invention is shown as a formative example, but the present invention is not limited to these formulations.

≪ Formulation Example 4 > Lotion (skin lotion)

Raw material Formulation Example 4  Natural herbal extracts 7.0 glycerin 3.0 Butylene glycol 2.0 Propylene glycol 2.0 Polyoxyethylene (60) Hardened castor oil 1.0 ethanol 10.0 Triethanolamine 0.1 antiseptic a very small amount Pigment a very small amount Spices a very small amount Purified water Balance

<Formulation Example 5> Massage cream

Raw material Formulation Example 5  Natural herbal extracts 7.0 Wax 10.0 Polysorbate 60 1.5 Sorbitan sesquioleate 0.8 Liquid paraffin 40.0 Squalane 5.0 Caprylic / capric triglyceride 4.0 Carboxyvinyl polymer 0.2 glycerin 5.0 Butylene glycol 3.0 Propylene glycol 3.0 Triethanolamine 0.2 antiseptic a very small amount Pigment a very small amount Spices a very small amount Purified water Balance

The supercritical fluid extraction method according to the present invention can extract the natural active ingredient with almost no destruction. In addition, the content of the natural active ingredient in the extracts of the general solvent was very small, but the yield of the supercritical fluid extraction method was 3 to 4 times or more. Therefore, when the same amount of extract was applied, it could contain more natural active ingredient than ordinary solvent extract, and it was possible to obtain natural ingredient without destruction of the applied natural active ingredient. In addition, the cost of extraction is also low, which is very economical.

In the present invention, the extracts of germanium, royal jelly, salmon roe, and quercus mongolica were extracted with supercritical fluid extraction method and then mixed with the mixed extracts extracted by the general solvent extraction method at a ratio of 6.5: 3.5. It is possible to prevent skin aging, skin aging, skin aging, and moisturizing function, which are combined with cosmetics to reduce skin wrinkles, elasticity, and moisturizing ability through cytoprotection from ultraviolet rays, cell proliferation ability, And the like.

Claims (10)

1) mixing, grinding, mixing, and stirring a mixture of the tenant-associated ingredients with the solvent, ginger mushroom, antler, lemon and red ginseng; 2) placing the mixture in a supercritical water extraction tank and supplying supercritical fluid; 3) maintaining the pressure of the extraction tank at 330 atm and raising the temperature in the extraction tank to 42 ° C at a rate of 0.5 ° C / min; 4) maintaining the temperature and pressure of the extraction tank at 30 to 50 ° C, 250 to 400 atm for 50 minutes, and then reducing the pressure to normal pressure to obtain the first mixed extract; 5) extracting the second mixed extract with water, an organic solvent or a mixed solvent thereof, among the tenant-related mixed ingredients, extracting the zeolites, germanium, royal jelly, salmon roe and bilberry; And 6) mixing the first mixed extract and the second mixed extract at a weight ratio of 6.5: 3.5; &Lt; / RTI &gt; The method according to claim 1, wherein the first mixed extract comprises 5-15% by weight of ginseng, 10-15% by weight of Ganoderma lucidum, 15-25% by weight of antler, 5-15% by weight of red ginseng, 15 to 20% by weight of ginseng, 0.0001 to 0.0003% by weight of germanium, 10 to 15% of royal jelly, By weight and salmon eggs in a dry weight ratio of 3 to 10% by weight. The method according to claim 1, wherein the solvent of the supercritical fluid extraction method is at least one selected from the group consisting of butylene glycol, water, and ethanol. The method of claim 1, wherein carbon dioxide is used as the supercritical fluid. The method according to claim 1, wherein the organic solvent of the general solvent extraction method is at least one selected from the group consisting of ethanol, methanol, butanol, acetone, ethyl acetate, hexane, propanol, butylene glycol and propylene glycol. Way. 10. The herbal mixed extract prepared by the method according to claim 1. A cosmetic composition characterized by containing an intestinal mixed extract according to claim 6. 8. The cosmetic composition according to claim 7, wherein the cosmetic composition contains 0.1 to 10% by dry weight of the total composition of the total composition. The cosmetic composition according to claim 7, wherein the cosmetic composition is formulated with a lotion, a skin lotion, a nutrition lotion, a nutritional cream, a massage cream or a nutritional essence. [Claim 7] The cosmetic composition according to claim 7, wherein the extract is contained in ultraviolet rays for cell protection, cell proliferation or moisturizing.

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