KR101837020B1 - Separation method for saponins from Sapindus mukorossi and cosmetic composition containing the same - Google Patents

Abstract

The present invention relates to a method for separating saponin from soapberry with high purity. More specifically, the present invention relates to a method for separating high purity saponin at an optimum yield by continuous processes, comprising the following processes: a hot water pretreatment process for eliminating acid components and sugar components which cause yellowing and darkening during purification by pretreating the soapberry with hot water to separate saponin from the soapberry; an extraction process for extracting the saponin component exhibiting intermediate polar surfactant activities as much as possible by carrying out reflux extraction with a 40-60% ethyl alcohol solvent; and a separation and purification process for completely removing inactive pigment layers to acquire only a high amount of active saponin. According to the present invention, the soapberry extract containing the isolated high purity saponin can be applied and processed into external skin applications and cosmetic ingredients which are excellent in cleaning effects, acne improvement effects, antioxidation effects, whitening effects, wrinkle ameliorating effects, anti-inflammatory effects, antibacterial effects, atopic alleviation effects, and ultraviolet protection effects.

Description

Description: TECHNICAL FIELD The present invention relates to a method for separating high-purity saponins from fruit-bearing fruits and a cosmetic composition containing the saponins from Sapindus mukorossi and a cosmetic composition containing the same.

The present invention relates to a method for separating saponins from fruit juice with high purity and a cosmetic composition containing the saponin. More particularly, the present invention relates to a method for separating saponin from fruit juice by heat treatment of heat- The extraction process that extracts the saponin component exhibiting the intermediate polar surface activity by the reflux extraction with a 40-60% ethyl alcohol solvent, and the extraction process which completely removes the pigment layer which is not active, The present invention also relates to a method for separating high purity saponin at an optimum yield by a continuous process including a separation and purification step of obtaining saponin only. The present invention relates to a cosmetic which is excellent in cleaning effect, acne improvement effect, antioxidant effect, whitening effect, wrinkle improving effect, anti-inflammatory effect, antibacterial effect, atopy improvement effect and ultraviolet ray protection effect It can be applied as an external preparation for skin.

Human skin has various physico-chemical changes in the aging process and it is divided into intrinsic aging and photo-aging depending on the cause. In other words, aging may occur due to activation of free radicals due to ultraviolet rays, stress, disease state, environmental factors, wounds, and aging. When such conditions become worse, the antioxidant defense network existing in the living body is destroyed, Thereby promoting adult diseases and aging. More specifically, lipids, proteins, polysaccharides and nucleic acids, which are major components of the skin, are oxidized to destroy skin cells and tissues, resulting in skin aging. In particular, the oxidation of proteins is caused by severing collagen, hyaluronic acid, elastin, proteoglycan, and fibronectin, which are connective tissues of the skin, resulting in severe hyperinflammation reaction and hindering skin elasticity. Causing mutations, cancer, and immune system degradation.

Therefore, it is necessary to protect the cell membrane by destroying the free radicals mediated by free radicals, ultraviolet rays, and inflammation that occur during the metabolism of the body, and to proliferate cells by regenerating already damaged cells by active metabolism, Healthy skin can be maintained. In addition to free radicals, MMP (matrix metalloproteinase) is involved in senescence, but the synthesis and degradation of extracellular matrix such as collagen is controlled in vivo, but its synthesis is reduced as aging progresses and the collagen- And the elasticity of the skin is lowered and wrinkles are formed. In addition, decomposition enzymes such as MMP may be activated by ultraviolet irradiation. Therefore, it is required to develop a substance capable of controlling the activation of MMP expression in cells or inhibiting its activity. Until now, the raw materials used as cosmetic materials mostly inhibited only the enzyme activity.

Next, skin changes due to pigmentation. Melanin, melanoid, carotene, and hemoglobin are the most important factors affecting skin color. The most important factors affecting biosynthesis are melanin, ultraviolet light and hormone secretion. Melanin absorbs or scatters ultraviolet light and plays a major role in preventing skin damage from ultraviolet rays. It does not have a specific maximum absorption wavelength and absorbs light in all areas. In addition, melanin is excellent in the function of removing reactive oxygen species, but sometimes melanin itself generates active oxygen, and other substances are reduced or oxidized by catechol or quinone in the melanin structure, and melanin itself is free radical It may also show properties.

The cause of atopic dermatitis has not yet been clearly identified, and sometimes it is expressed as skin dryness, eczema, etc. Atopic dermatitis is caused by a number of factors. Several external environmental factors associated with aggravation of atopic dermatitis are well known, resulting in increased serum IgE, increased IL-4 and IL-5, and increased INF-r. Skin abnormalities that occur in atopic dermatitis include decreased skin moisture content, reduced natural moisturizing factors, and increased transdermal water loss.

Sapindus mukorossi ) is distributed in China, Taiwan, Japan and Korea. It is a deciduous broad-leaved arboreous tree that grows mainly in Jeju Island, Jeolla-do and Kyungsang-do. Leaves are alternate phyllotaxis and odd numbered compound leaf. Leaflets are 9 ~ 13, long oval or long oval lanceolate. It is 7 ~ 14cm × 3 ~ 4.5cm in length and width. It has many wrinkles, flat edges, small petiole length of 2 ~ 6mm and height up to 20m. The flower is bloomed in May-June with a male and female flower. The conical flower has a short hairs with a length of 20 ~ 30cm and a female flower with a diameter of 4 ~ 5mm. There are 4-5 calyx pieces and petals, 8-10 stamens in male flower, and female pistil has 1 pistil. Fruit is round, hairless, and yellowish brown. The seeds are black, one in each, and mature in mid-October. It is planted with the number of trees, the number of the trees, the number of the trees, the number of the ecological parks. Wood is used as an appliance material, seeds are used to make beads and toys, and berries are used as a substitute for soap. Seeds, roots, bark, leaves, pulp, seeds and so on. When you use it as a medicine, you can use it as a bath or as a powder. Put powder on the trauma. The bark and fruit skin of the buds of the moths are rich in saponin, which is a natural surfactant, and they are used for laundry as a substitute for soap.

Korean Patent No. 10-1087734 discloses a method for purifying saponin from a mushroom tree, a dichloromethane fraction of a mushroom tree, and a hair cosmetic composition comprising the same. The present invention discloses a method for purifying saponin from saponin from fruit-bearing fruit, which comprises extracting perilla using functional ethanol and then fractionating the saponin with dichloromethane to obtain 25 to 35% of saponin, and a hair cosmetic composition comprising the saponin , This method is refined using a harmful organic solvent called dichloromethane because it is difficult to exclude acid and saccharides which cause galling and blackening during refining. The result of the refining is also poor in purity and is 25-35% Of relatively low saponins.

Korean Patent Laid-Open Publication No. 10-2016-0052066 discloses a method for extracting saponin by ultrasonication with an inorganic powder and a detergent composition for a mixture of a natural product and a surfactant. In this method, The yield is low by adding powder and extracting and filtering, and it is difficult to exclude the acids and saccharides which cause galling and blackening, so that the saponin is dark and relatively low in saponin because of low purification.

Korean Patent Laid-Open Publication No. 10-2012-0054297 discloses an oral composition containing an alopecia extract as an active ingredient. The present invention relates to a composition for oral use, which is obtained by simply extracting the alfalfa fruit with an 80% ethanol solution, aging, ≪ / RTI >

Accordingly, it is an object of the present invention to provide a method for producing an extract containing a high content of saponin from a non-treated fruit and a cosmetic use of the functional cosmetic composition containing the saponin, More preferably, the present invention is to provide a cosmetic composition containing a high-content saponin which is excellent in detergency, acne improvement, antioxidation, skin whitening, wrinkle improvement, anti-inflammation, antibacterial effect, atopy improvement effect and ultraviolet protection effect.

Therefore, the present inventors have studied the possibility of applying various natural extracts, which have not yet been known, to cosmetics. As a result, they have found out that a high content of saponin can be extracted and purified from fruit juice to improve the cleansing effect, acne improvement effect, antioxidative effect, whitening effect, Antimicrobial effect, atopy improvement effect and ultraviolet ray protection effect, it is possible to expect an efficacy as a cosmetic and external preparation for skin.

In order to accomplish the above object, the present invention provides a method for preparing an extract of fruit juice containing a high amount of saponin and a method for producing the same, which comprises the step of washing, acne improvement, antioxidation, skin whitening, skin wrinkle improvement, And atopic dermatitis.

More preferably, the present invention relates to a process for hydrothermal treatment, which comprises the steps of hydrothermal pretreatment for eliminating an acid component and a sugar component which cause gelation and blackening at the time of purification by hydrothermal treatment of an aluminiatic fruit, reflux extraction with a 40 to 60% ethyl alcohol solvent, A method for separating high purity saponin including an extraction process for extracting the saponin component to the maximum extent and a separation and purification process for completely removing the pigment layer having no biological activity to obtain only a high amount of active saponin and a cleaning method, Provided is a functional cosmetic composition excellent in whitening, wrinkle improvement, anti-inflammation, antibacterial, atopic improvement, and ultraviolet protection effect.

The present invention

(A) a pretreatment step of eliminating an acid component and a sugar component which cause a galling and a blackening phenomenon at the time of refining by extracting hot water or refluxing the fruit of a non-treating fruit;

(B) an extraction step of extracting the saponin exhibiting an intermediate polar surfactant by reflux extraction with a 40-60% ethyl alcohol solvent, which is obtained in the step (a); And

(C) a purification step of removing high levels of saponin by removing the pigment layer and impurities from the ethanol-free fruit extract obtained in the step (b).

Further, the present invention relates to a method for separating high purity saponin from a non-fungal fruit characterized in that the step (c) is carried out using an adsorbent.

Also, the present invention relates to a method for separating high purity saponin from a non-fungal fruit characterized in that the step (a) is carried out by adding 1 to 10% by weight of an unaffected fruit to a solvent.

Also, the present invention relates to a method for separating high purity saponins from a non-fungal fruit characterized in that the step (b) is carried out by adding 1 to 10% by weight of an alcohol-free alcohol-free fruit extract pretreatment.

The present invention also relates to a method for separating high-purity saponins from fruit juice, which is characterized in that the high-purity saponin content obtained through the step (c) is 85% or more.

In addition, the present invention provides a cosmetic composition containing an alopecic fruit extract,

(A) a pretreatment step of eliminating an acid component and a sugar component which cause a galling and a blackening phenomenon at the time of refining by extracting hot water or refluxing the fruit of a non-treating fruit;

(B) an extraction step of extracting saponin exhibiting an intermediate polar surface activity by reflux extraction with a 40-60% ethyl alcohol solvent, which is obtained by the above process (a); And

(C) a step of removing the coloring matter layer and the impurities from the ethanol-free extract of the non-treated fruit obtained in the step (b) to obtain a saponin of 85% or more; To a cosmetic composition.

According to the method of the present invention, the non-specific fruit extract of the present invention can remove acid and sugar components, which cause pretreatment, 40 to 60% functional ethanol treatment, and pigment layer and impurity removal, So that the saponin content is very high and the browning phenomenon does not occur, so that it is suitable for use in cosmetics and external skin preparations.

The present invention also relates to a cosmetic composition characterized in that the cosmetic composition is a cleansing detergent, a hair cleansing agent or a cleansing agent for body cleansing.

Also, the present invention relates to a cosmetic composition, wherein the cosmetic composition is selected from cosmetic lotion, gel, water-soluble liquid, cream, essence, oil / water and w / o formulations.

The present invention also relates to an anti-aging functional cosmetic composition characterized in that the cosmetic composition has an antioxidative effect.

The present invention also relates to an anti-aging functional cosmetic composition characterized in that the cosmetic composition exhibits an effect of improving skin wrinkles.

The present invention also relates to a cosmetic composition characterized in that the cosmetic composition is a whitening functional cosmetic composition.

The present invention also relates to a cosmetic composition characterized in that the cosmetic composition is an antibacterial functional cosmetic composition for improving acne symptoms and the like.

The present invention also relates to a cosmetic composition characterized in that the cosmetic composition is an anti-inflammatory functional cosmetic composition for improving atopy symptoms and the like.

Hereinafter, a method for producing the high-content saponin-free fruit extract of the present invention and a cosmetic composition containing the same will be described in more detail.

First, the high purity saponin separation method of the present invention starts with a pretreatment process. The optimum pretreatment process according to the present invention is performed by adding the fruit juice to the water solvent at a ratio of 0.5 to 20.0% (w / w) to the amount of 0.5 to 20.0% (w / w) Heated for 5 hours, extracted with hot water or refluxed, filtered and dried to dry.

The ethanol extracting process of the present invention is carried out at a temperature of 40 to 60 ° C for extracting a high content of saponin from the pre-treatment extract of the fruit with no acid component and saccharin components which cause galling and blackening phenomenon through the pretreatment process. % Ethanol. It can be applied by various methods such as heat extraction and room temperature extraction. When the ethanol concentration is out of this range, it is difficult to obtain a high amount of saponin because a large amount of polyphenol components having low surfactant ability is extracted.

The saponin-free fruit extract containing the high content of saponin obtained through the ethanol extraction process is subjected to a decoloring process. The decoloring process according to the present invention is carried out using activated carbon, Amberlite XAD resin, Dow X resin, suferrite DAX resin, -10, 20, 30, 40, and 50 resin is selected and decolorized. At this time, when the decoloring process is carried out without performing the pre-treatment process, refinement is not performed because of galling and blackening phenomenon.

The fruit of the above-mentioned passive fruit which has undergone the decolorization step is subjected to a filtration and concentration process to remove the decolorizing auxiliary carrier and the functional ethanol. The concentration process according to the present invention can be carried out by various methods such as heat drying, vacuum drying, spray drying or freeze drying And may be subjected to a filtration and concentration process to obtain an apple fruit extract containing a high amount of saponin in powder form.

The present invention relates to a process for producing an extract of saponin comprising a high content of saponin and a cosmetic composition containing the saponin and a cosmetic composition containing the saponin, .

In the cosmetic composition of the present invention, the content of the high-content saponin-containing fruit juice extract is preferably 0.0005 to 10% by weight based on the total weight of the cosmetic composition, and the content of the saponin- The cosmetic composition can be manufactured into various formulations such as wash-offs, cosmetics, essences, creams, packs, patches, bubble-forms, cleansing foams, skin adhesive gels, foundations, make- It can be applied to the manufacturing method. The above-mentioned high-dose saponin-free fruit extracts of the present invention can be applied in various forms such as liquid, cream, paste, and solid forms. And may contain various conventional adjuvants and carriers.

According to the method of the present invention, it is possible to separate saponin of high purity from the fruit juice of the fruit with optimum yield by a continuous process including a pretreatment step, an extraction step of reflux extraction with 40 to 60% ethyl alcohol solvent and a separation and purification step of obtaining only active saponin have.

In addition, according to the method of the present invention, it is possible to obtain a high-purity saponin-free fruit juice extract containing high purity saponins according to the present invention, which has excellent cleaning effect, acne improvement effect, antioxidant effect, whitening effect, wrinkle improving effect, anti- And can be applied to cosmetics and external skin preparations.

Fig. 1 is a photograph showing degree of browning of Formulation Example 1 and Formulation Comparative Examples 1, 2, 3 and 4 after 4 weeks of daylight condition.

In order to facilitate understanding of the present invention, the present invention will be described in more detail with reference to specific examples such as examples, comparative examples and test examples. It will be apparent to those skilled in the art that the present invention is not limited to the embodiments described below.

≪ Example 1 >

5% by weight of fruit juice was added to a water solvent and the mixture was pre-treated at 80 ° C for 2 hours in a reflux condenser equipped with a condenser. After filtration with an 80 mesh filter and a 1.2um filter, the filtrate was discarded to obtain an alum freeze dried product, which was dried for one day in a drying oven at 80 ° C. 5% by weight of dried extract of pre-treatment fruit of fruit juice was added to 10, 20, 30, 40, 50, 60, 70, 80, 90, and 99.5% ethanol aqueous solutions, respectively, and heated in a reflux extractor Lt; / RTI > Thereafter, the mixture was filtered through an 80-mesh filter and a 1.2-μm filter to obtain an extraction filtrate. 5% by weight of activated carbon was added to the extracted filtrate, and the mixture was stirred at room temperature for three hours. The filtrate was concentrated by using a vacuum concentrator (Rotary Evaporator NE-series, Eyela, Japan) at 65 ° C and 20 hPa for one hour. The filtrate was filtered through an 80 mesh filter and a 1.2 μm filter to obtain a filtrate. A concentrated powder was obtained. The result thus obtained was used in the following test examples.

≪ Comparative Example 1 &

5% by weight of fruit juice was added to 10, 20, 30, 40, 50, 60, 70, 80, 90, and 99.5% ethanol aqueous solutions respectively and extracted with a reflux extractor with a cooling condenser at 80 ℃ for 2 hours. Thereafter, 80 mesh filter and 1.2um filter were used for filtration to obtain an extraction filtrate. To the extraction filtrate thus obtained, 5 wt% of activated carbon was added and the mixture was stirred at room temperature for 3 hours. The filtrate was concentrated by using a vacuum concentrator (Rotary Evaporator NE-series, Eyela, Japan) at 65 ° C and 20 hPa for one hour to obtain a concentrated powder ≪ / RTI > The result thus obtained was used in the following test examples.

≪ Comparative Example 2 &

100 g of fruit juice was placed in a reflux extractor equipped with a reflux condenser, which was supplied with cooling water by tap water or a cooling circulator and the sublimated solvent was returned to the extractor, and 1 L of 95% methanol was added to the slurry. The temperature of the outer jacket of the extractor was fixed at 70 캜 and extracted for 4 hours to prepare a 95% methanol extract twice. The extract was cooled at room temperature, filtered through a filter paper (Watman No.2, USA), concentrated under reduced pressure to 100 to 250 atm with a rotary vacuum concentrator (IEA, Japan) to recover the alcohol component in the filtrate, Crude extracts were prepared. Since the crude extract is not pure saponin, a dichloromethane solvent (concentration 95%) is added to remove the peeled water and other extracted components (polysaccharide, phenol compound, etc.) extracted by alcohol, The other condensed extract components were separated. The dichloromethane-soluble fraction was concentrated under reduced pressure to 70 to 90 atm with a rotary vacuum concentrator (IEA, Japan) to prepare a saponin fraction. The saponin fraction thus prepared was centrifuged at 3000 rpm to remove non-saponin condensation components which were not removed, thereby obtaining a purified dichloromethane fraction containing pure saponin saponin.

≪ Comparative Example 3 &

After removing the seeds from the dried fruit of the mulberry tree, 100 g of the fruit powder of the mulberry fruit, 20 g of gray tourmaline powder (particle size: 30 nm) and 630 g of purified water were put into a 2-liter beaker and mixed. The mixture was stirred at 80 rpm at 20 rpm for four hours, and then dispersed for one hour at 20 Hz and 200 W using an ultrasonic wave extender (sonifier 450). After the dispersion was completed, the solution was filtered with a 0.45 mu m filter, and the filtered solution was centrifuged at 3,000 to 4,000 rpm for 10 minutes to extract 480 g of the solution. Drying oven The dried saponin powder was obtained by hot-air drying at 80 ° C for 7 hours. Secondary extraction was carried out by using the residue remaining in the filter to obtain a powdery saponin extract.

≪ Comparative Example 4 &

The fruit was extracted with 80% ethanol solution, filtered and aged for 5 days. The resulting aged material was filtered to remove impurities, concentrated under reduced pressure, and dried to obtain an exfoliated fruit extract.

[Test Example 1] Analysis of saponin content

The saponin content of the sample of the present invention was analyzed as follows. 0.5 mL of vanilin solution (8%, w / v) was added to 0.5 mL of the prepared sample, and 5.0 mL of sulfuric acid (72%, w / v) was mixed in an ice water bath. Then, it was heated at 60 DEG C for 10 minutes and cooled in an ice water bath for 10 minutes. The absorbance of the cooled sample was measured at 535 nm using a microplate reader, and its content was calculated in comparison with the absorbance of the standard sample saponin (Sigma).

sample menstruum Saponin content (%) Pretreatment
O
Example 1 10% functional ethanol 45 20% functional ethanol 48 30% functional ethanol 52 40% functional ethanol 88 50% functional ethanol 92 60% functional ethanol 87 70% functional ethanol 64 80% functional ethanol 62 90% functional ethanol 61 99.5% functional ethanol 60 Pretreatment
X
Comparative Example 1 10% functional ethanol 25 20% functional ethanol 27 30% functional ethanol 28 40% functional ethanol 55 50% functional ethanol 64 60% functional ethanol 57 70% functional ethanol 38 80% functional ethanol 35 90% functional ethanol 33 99.5% functional ethanol 33 Comparative Example 2 95% methanol, dichloromethane 30 Comparative Example 3 water 13 Comparative Example 4 80% ethanol 19

[Test Example 2] Measurement of cleaning force

The washing power of the sample of the present invention was measured as follows. After 5 cm of each sample was prepared, 5 mL of water was applied to each sample, and 1 mL was applied. After 30 seconds of massage, the sample was wiped with a napkin to determine the cleaning effect of the oil type pen. Respectively.

sample
Cleaning power Pretreatment O
Example 1 (50% functional ethanol solvent) +++ Pretreatment X
Comparative Example 1 (50% functional ethanol solvent) ++ Comparative Example 2
+ Comparative Example 3
- Comparative Example 4
+ +++; Completely cleared
++; 1/2 or more erased
+; 1/4 or more cleared
-; Less than 1/4 cleared

[Test Example 3] Measurement of acne improvement effect

In this experiment, the inhibitory effect of Propionibacterium acne causing acne was investigated. In 20 acne women, clinical trials were conducted for 4 weeks of acne improvement with sample cream. The results are shown in Table 3 below.

1) Propionibacterium acne inhibition experiment

To determine the antimicrobial activity against Propionibacterium acnes according to the test group, the minimum inhibitory concentration was measured using a solid culture dilution method (Agar Serial Dilution Method).

The cells were inoculated into the medium and incubated for 2 days in a 37 ° C shaking incubator. Each test group was diluted in 5% DMSO (dimethylsulfoxide) physiological saline solution in sterilized Patriedish for each sample and experimental species And 2 mL of 5% DMSO physiological saline solution was added to the control group. After sterilization in each Patridish, 18 mL of the agar medium cooled to 48 DEG C was added thereto, stirred, and then allowed to stand and solidify.

Then, each of the pre-cultivated test bacteria was applied to each petri dish at a final concentration of about 1 × 10 ⁶ CFU / mL, cultured at 37 ° C. for 2 days, and colonies were observed in each compartment, The minimum sample concentration of the plate was defined as the minimum inhibitory concentration (MIC), and the results are shown in Table 3 below. At this time, the minimum inhibitory concentration means that the smaller the value, the higher the antibacterial effect.

2) Clinical trial to improve acne

The results of the clinical trial of the acne improvement effect of the test group cream for 20 weeks were shown in Table 3. The cream formulations used were as shown in Table 4. The creams were prepared using the samples of the respective examples and comparative examples and used.

sample Minimum inhibitory concentration (MIC, ug / mL) acne
4 week clinical evaluation Pretreatment O
Example 1 (50% functional ethanol solvent) 150 +++ Pretreatment X
Comparative Example 1 (50% functional ethanol solvent) 300 ++ Comparative Example 2
1200 + Comparative Example 3
> 5000 - Comparative Example 4
2400 - +++; Very good improvement
++; Good improvement
+; Slight improvement
-; no effect

number ingredient Formulation Example One Pro-type glycerin monostearate 2.0 2 Stearic acid 1.5 3 Cetearyl alcohol 2.2 4 Wax 1.0 5 Squalane 3.0 6 Hardened vegetable oil 1.0 7 Sorbitan stearate 0.6 8 Mineral oil 5.0 9 Polysorbate 60 1.5 10 Dimethicone 1.0 11 Trioctanoin 5.0 12 Betaine 3.0 13 Triethanolamine 1.0 14 glycerin 5.0 15 Sodium hyaruronate 3.0 16 In each of the examples and comparative examples 1.0 17 Distilled water Balance 18 Preservative, fragrance, pigment a very small amount

[Test Example 4] Measurement of antioxidant effect

The free radical scavenging test and the active oxygen scavenging test were carried out in order to confirm the antioxidative effect of the test group of the present invention.

The free radical scavenging test uses the fact that the absorbance of stable DPPH exhibits the maximum absorbance at 540 nm. As the free radical DPPH is erased by the sample and becomes purple to transparent color, that is, as the free radical scavenging rate increases, the absorbance at the 540 nm wavelength The test was conducted as follows.

1 mL of the above-mentioned sample was diluted to 1 mL of a 0.1 mM DPPH (2,2-diphenyl-1-picryl-hydrazyl radical, Sigma) solution, and the mixture was allowed to stand at 37 캜 for 15 minutes. -06685, Thermo max, USA) was used to measure the absorbance at a wavelength of 540 nm.

In the free radical scavenging test, 1 ml of DPPH and 1 ml of solvent were added to the control group, and 1 ml of methanol and 1 ml of the sample were added to obtain the respective color correction values for the sample and the control group.

The free radical scavenging ratios were calculated using the following equation (1). In Table 5, SC ₅₀ is the concentration of the sample required to remove 50% of the free radicals, and a smaller value indicates a higher antioxidant activity.

[Equation 1]

Free radical scavenging rate (%) = 100 - ((absorbance of sample / absorbance of control) x 100)

In the active oxygen scavenging test, the change of absorbance by the oxidation of nitroblue tetrazolium (NBT) by reactive oxygen species using active oxygen generation by enzyme reaction of xanthine / xanthine oxidase (Sigma) By measuring, the active oxygen scavenging ability can be known.

Add 0.1 mL of Na ₂ CO ₃ , 0.1 mL of xanthine, 0.1 mL of ethylenediamine tetraacetic acid (EDTA), 0.1 mL of BSA (bovine serum albumin, Sigma), 0.1 mL of NBT and 0.1 mL of the sample, add to a vortex mixer Mixer, Mini mix, USA) and allowed to stand at 25 ° C for 10 minutes. After 0.1 ml of xanthine oxidase was added, the mixture was reacted at 25 ° C for 20 minutes, and 6 mM CuCl ₂ was added to stop the reaction , And the absorbance at 540 nm was measured using a microplate reader (UVT-06685, Thermo max, USA).

In the test, the control solution was prepared by adding the third distilled water instead of the sample solution. The third control solution was added to the third solution in place of the xanthine oxidase solution to obtain the corrected color values of the extracted sample and the control.

The active oxygen scavenging rate is numerically calculated using Equation (2) and shown in Table 5 below. In Table 5, the IC ₅₀ is a sample concentration required to remove 50% of the active oxygen, and a smaller value means that the antioxidant activity is stronger.

&Quot; (2) "

Ratio of active oxygen scavenging (%) = 100 - ((absorbance of sample / absorbance of control) x 100)

Name of sample Antioxidative effect
Free radical scavenging rate
SC ₅₀ (%) Active oxygen scavenging rate
IC ₅₀ (%) Pretreatment O
Example 1 (50% functional ethanol solvent) 2 5 Pretreatment X
Comparative Example 1 (50% functional ethanol solvent) 5 10 Comparative Example 2
15 > 15 Comparative Example 3
> 15 > 15 Comparative Example 4
> 15 > 15

[Test Example 5] Measurement of whitening effect

In order to confirm the whitening effect, the degree of inhibition of the tyrosinase enzyme function was evaluated by the skin aging improving effect test for the test group of the present invention. Tyrosinase is an enzyme that helps the production of melanin by promoting the oxidation process of tyrosine in vivo. Melanin is a black polymer that melts in the body to form melanin. When it moves to the upper part of the skin, the skin turns black and produces spots and freckles. (Pomerantz S. H., J. Biochem., 24: 161-168, 1966) was used to determine the degree of suppression of the action of tyrosinase and to determine the whitening effect. The test method is as follows.

0.9 mL of the sample, 1.0 mL of 0.1 M phosphate buffer (pH 6.8) and 1.0 mL of 1.5 mM L-tyrosine solution were added, and the mixture was maintained at 37 DEG C for 10 minutes. After incubation at 37 ° C for 10 minutes, the absorbance was measured at 475 nm using a UV-vis spectrophotometer (Smartspec Plus, Biorad, USA) The inhibition rate against Nausea was measured.

As a control group for the inhibition of tyrosinase activity, a buffer solution was added in place of the sample solution, and the same method was used. A buffer solution was added instead of tyrosinase to obtain the respective color correction values for the sample and the control.

The test samples used in the tests were the test substances obtained in each of the examples and the comparative examples, and the tyrosinase inhibition rates were calculated numerically using the following equation (3). In Table 6, IC ₅₀ is the concentration of the sample required to inhibit tyrosinase activity by 50%, and the smaller the value, the higher the inhibition rate.

&Quot; (3) "

Inhibition rate (%) = 100 - ((absorbance of sample / absorbance of control group) x 100)

sample Inhibition rate of tyrosinase activity,
IC ₅₀ (mg / mL) Pretreatment O
Example 1 (50% functional ethanol solvent) 2.5 Pretreatment X
Comparative Example 1 (50% functional ethanol solvent) 5.4 Comparative Example 2
22.5 Comparative Example 3
> 50 Comparative Example 4
42.3

[Test Example 6] Elastase activity inhibition test

In order to test the skin wrinkle suppression and improving effect of the sample obtained in the present invention, the Elastase activity inhibition test was conducted. Elastin is a constituent of the matrix layer in the dermis of the skin. As the skin ages, elastin breaks down and the matrix layer in the dermis breaks down and wrinkles occur. (Ala) 3 p-nitroaniline (N-succinyl- (Ala) 3 p-nitroaniline, which is an elastase substrate, is used as a method for measuring the activity of Elastase, nitroaniline, Sigma) is used to measure the absorbance at a wavelength of 405 nm as a change in color caused by decomposition of p-nitroaniline. The buffer solution was pH 8.0, 0.267 M Trizma-HCl (Sigma), the substrate solution was 8.8 mM N-Succinyl- (Ala) 3 p-nitroaniline and the enzyme solution was used at a concentration of 10 ug / mL (Sigma) . 20 μL of the buffer solution and 20 μL of the substrate solution were mixed with 100 μL of the sample solution diluted in the third distilled water for each concentration. Then, 20 μL of the enzyme solution was added and reacted in a constant temperature water bath at 25 ° C. for 15 minutes to measure the amount of p-nitroaniline produced, (UVT-06685, Thermo max, USA) at a wavelength of 405 nm.

As a control group for measuring the activity of the Elastase, third distilled water was added instead of the sample, and the same method was used.

The sample used in the test is shown in Table 7 by numerically calculating the inhibition rate of Elastase using the following formula (4). In Table 7, IC ₅₀ is the concentration of the sample required to inhibit the enzyme activity by 50%, and the lower the value, the higher the inhibition rate.

&Quot; (4) "

Elastase activity inhibition rate (%) = 100 - (absorbance of sample / absorbance of control group) x 100)

sample Elastase activity inhibition rate,
IC ₅₀ (mg / mL) Pretreatment O
Example 1 (50% functional ethanol solvent) 3 Pretreatment X
Comparative Example 1 (50% functional ethanol solvent) 8 Comparative Example 2
> 20 Comparative Example 3
> 20 Comparative Example 4
> 20

[Test Example 5] Collagen synthesis effect test

In order to test the effect of improving the skin wrinkles of the samples obtained in the present invention, the collagen aggregation performance was measured. Collagen is a constituent component of the dermal matrix layer in the skin together with the elastin, and the skin gradually ages, and the components constituting the matrix layer in the dermis are decomposed to form wrinkles. The collagen is a constituent of the dermal matrix layer By confirming the synthesis effect, the effect of improving the skin wrinkles can be confirmed. Proceed as follows.

Human normal dermal fibroblasts were inoculated into 24-well microplates (1 x 10 ⁵ cells / well) and incubated for 24 hours at 37 ° C in a 5% CO ₂ incubator. Each sample was incubated in serum-free DMEM medium for 24 hours. The amount of collagen synthesis is measured by enzyme immunoassay as follows.

The medium cultured for 24 hours was dispensed in a 96-well microplate and coated overnight at 4 占 폚. After washing three times with wash buffer (PBS-T; 0.05% Tween 20 in phosphate buffered saline), blocking solution (5% skim milk, Fluka) was added and blocked at 37 ° C for 1 hour. The blocking solution was discarded and washed three times with washing buffer. The primary antibody (rabbit anti-collagen type I, Sigma) was diluted in PBS-T and added in an amount of 100 μl. The reaction was carried out at 37 ° C for 2 hours. After washing three times with washing buffer, 100 μl of secondary antibody (anti-rabbit IgG alkaline phosphatase conjugate, Sigma) was diluted in PBS-T and reacted at 37 ° C for 2 hours. After washing three times with washing buffer, 100 μL of alkaline phosphatase substrate solution (Sigma) was added, and the solution was developed at 25 ° C. The absorbance at 405 nm was measured using a microplate reader (UVT-06685, Respectively.

The control group for measuring the collagen synthesis effect is the reaction absorbance of the cell culture without treatment of the sample, and the collagen synthesis effect is numerically calculated using the following formula (5).

&Quot; (5) "

Collagen synthesis effect (%) = 100 + (absorbance of control group - absorbance of cell culture liquid of sample) / absorbance of control group 100

Call
la
Gen
synthesis
castle
Hyo
and
(%) sample Concentration (mg / mL) 0
One 10 100 Pretreatment O
Example 1 (50% functional ethanol solvent) 100.00 114.97 132.54 141.12 Pretreatment X
Comparative Example 1 (50% functional ethanol solvent) 100.00 105.44 113.67 121.41 Comparative Example 2
100.00 101.32 106.43 108.20 Comparative Example 3
100.00 101.33 99.42 105.34 Comparative Example 4
100.00 102.21 101.43 103.24

[Test Example 6] Evaluation of inhibition of MMP-1 expression after irradiation with ultraviolet light

Enzyme immunoassay (ELISA), which measures the concentration of MMP-1 after UV irradiation and sample addition, was conducted as follows to evaluate inhibition of MMP-1 expression after UV irradiation of the sample obtained in the present invention.

UVA is irradiated to human dermal fibroblasts at an energy of 5 J / cm < 2 > using a UV chamber. Ultraviolet irradiation dose and incubation time were established by preliminary experiment to maximize MMP expression level in fibroblasts. The negative control group was wrapped with silver foil and kept in the UVA environment for the same time. The UVA emission was measured using a UV radiometer. The UVA-irradiated cells remained in the previously dispensed medium, irradiated with UVA, exchanged with the medium containing the sample, cultured for 24 hours, Respectively. The primary antibody (MMP-1 (Ab-5) monoclonal antibody and MMP-2 (Ab-3) monoclonal antibody) was treated and reacted at 37 ° C for 60 minutes. The secondary antibody, anti mouse IgG conjugated) was reacted for about 60 minutes. Then, an alkaline phosphatase substrate solution (1 mg / ml-nitrophenyl phosphate in diethanolamine buffer) was reacted at room temperature for 30 minutes and absorbance was measured at 405 nm using a microplate reader. As a control group, a sample to which no sample was added was used.

Test group
Treatment concentration (%) MMP-1 expression inhibition rate (%) Pretreatment O
Example 1 50% functional ethanol solvent 0.1 45 Pretreatment X
Comparative Example 1 50% functional ethanol solvent 0.1 28 Comparative Example 2
0.1 15 Comparative Example 3
0.1 3 Comparative Example 4
0.1 12

[Experimental Example 7] Experiment to measure melanin formation inhibitory effect using B16F1 melanocyte

In order to test the whitening effect of the sample obtained in the present invention, the degree of inhibition of melanin formation on B16F1 melanocyte was examined as follows to determine the whitening effect. The B16F1 melanocyte used in this test example is a cell strain derived from a mouse, and is a cell that secretes a melanin pigment called melanin. During the artificial culture of these cells, samples were treated to compare the degree of reduction of melanin pigment. The B16F1 melanocyte used in this test example was purchased from ATCC (American Type Culture Collection, Accession No. 6323).

The melanin biosynthesis inhibitory effect of B16F1 melanocyte was measured as follows. B16F1 melanocytes were dispensed into 6-well plates at a concentration of 2 x 10 ⁶ cells per well, and after attaching the cells, the samples were treated at a concentration that did not induce toxicity and cultured for 72 hours. After incubation for 72 hours, cells were detached with trypsin-EDTA, and the number of cells was measured and centrifuged to recover the cells. Quantification of intracellular melanin was carried out with a slight modification of the method of Lotan ( Cancer Res. , 40: 3345-3350, 1980). The cell pellet was washed once with PBS, and 1 ml of homogenization buffer (50 mM sodium phosphate, pH 6.8, 1% Triton X-100, 2 mM PMSF) was added and vortexed for 5 minutes to disrupt the cells. After centrifugation (3,000 rpm, 10 min), 1N NaOH (10% DMSO) was added to the cell filtrate to dissolve the extracted melanin, and the absorbance of melanin was measured at 405 nm using a microplate reader. Melanin was quantitated to determine the melanin Production inhibition rate was measured. Melanogenesis inhibition rate (%) of B16F1 melanoma site was calculated by the equation 6], IC ₅₀ value is the concentration of a substance that inhibits melanin production by 50%.

&Quot; (6) "

Inhibition rate (%) = [(A-B) / A] x 100

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

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

Name of sample Treatment concentration (%) Melanin synthesis inhibitory effect (%) Pretreatment O
Example 1 50% functional ethanol solvent 0.1 43 Pretreatment X
Comparative Example 1 50% functional ethanol solvent 0.1 28 Comparative Example 2
0.1 11 Comparative Example 3
0.1 5 Comparative Example 4
0.1 8

[Test Example 7] Cytotoxicity mitigation effect by ultraviolet irradiation

In order to evaluate the mitigation effect of the cytotoxicity by ultraviolet irradiation of the sample obtained in the present invention, the following experiment was conducted.

Fibroblasts were placed in 24-well test plates at 1 × 10 ⁵ cells and adhered for 24 hours. Each well was washed once with PBS and 500 ul of PBS was added to each well. After irradiating the fibroblasts with 10 mJ / cm2 of ultraviolet light using an ultraviolet B (UVB) lamp (Model: F15T8, UVB 15W, Sankyo Dennki, Japan), PBS was removed and cultured in a cell culture medium (DMEM supplemented with 10% FBS ). Here, the sample to be evaluated was treated and cultured for 24 hours. After 24 hours, the culture medium was removed, and 500 μl of the cell culture medium and 60 μl of the MTT solution (2.5 mg / ml) were added to each well, followed by culturing in a 37 ° C. CO ₂ incubator for 2 hours. The medium was removed and 500 μl of isopropanol-HCl (0.04 N) was added. After shaking for 5 minutes, the cells were lysed and 100 쨉 l of the supernatant was transferred to a 96-well test plate, and absorbance at 565 nm was measured in a microplate reader. The cell viability was measured by the formula (7), and the cytotoxic relaxation rate by ultraviolet was calculated by the following formula (8).

&Quot; (7) "

Cell survival rate (%) = [(St-Bo) / (Bt-Bo)] X 100

Bo: Absorbance at 565 nm of wells reacting with cell culture medium

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

St: absorbance at 565 nm of the well that underwent chromogenic reaction in the sample-treated well

&Quot; (8) "

(%) = [1- (St-Bo) / (Bt-Bo)] X 100

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

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

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

Name of sample

Treatment concentration (%)

Cytotoxic Relaxation Rate (%)
Pretreatment O
Example 1 50% functional ethanol solvent 0.1 58 Pretreatment X
Comparative Example 1 50% functional ethanol solvent 0.1 31 Comparative Example 2
0.1 5 Comparative Example 3
0.1 2 Comparative Example 4
0.1 4

[Test Example 9] Inhibitory effect of inflammatory cytokine expression by ultraviolet irradiation

In order to evaluate the effect of inhibiting the expression of inflammatory cytokines expressed by ultraviolet irradiation of the sample obtained in the present invention, the following experiment was conducted.

Fibroblasts isolated from human epidermal tissue were placed in a 24-well test plate in an amount of 5 × 10 ⁴ and adhered for 24 hours. Each well was washed once with PBS and 500 占 퐇 of PBS was added to each well. The fibroblasts were irradiated with 10 mJ / cm 2 of ultraviolet light using an ultraviolet B (UVB) lamp (Model: F15T8, UVB 15W, Sankyo Dennki, Japan), and PBS was removed. Cell culture medium (DMEM without FBS) Was added. Here, the sample to be evaluated was treated and incubated for 5 hours. 150 [mu] l of the culture supernatant was taken to quantitate IL-l [alpha] to determine the effect of inhibiting the inflammatory cytokine expression of the sample. The amount of IL-l [alpha] was quantitated using Enzyme-linked Immunosorbent Assay and the production rate of IL-l [alpha] was calculated by [Equation 9].

&Quot; (9) "

Inhibitory rate of inflammatory cytokine expression (%) = [1- (St-Bo) / (Bt-Bo)] X 100

Bo: IL-1? Production in wells without UV irradiation

Bt: IL-1? Production in wells irradiated with ultraviolet light and not treated with the sample

St: IL-1α production in wells irradiated with ultraviolet light and treated with the sample

Name of sample
Treatment concentration (%) Inhibitory rate of inflammatory cytokine expression (%) Pretreatment O
Example 1 50% functional ethanol solvent 0.1 43 Pretreatment X
Comparative Example 1 50% functional ethanol solvent 0.1 38 Comparative Example 2
0.1 8 Comparative Example 3
0.1 5 Comparative Example 4
0.1 8

[Test Example 9] Antibacterial effect

The antimicrobial activity test for the sample obtained in the present invention was carried out by the solid culture dilution method. Experimental strains received pre-sale at the Korea Research Institute of Bioscience and Biotechnology, staphylococci (Staphylococcus aureus KCTC 6910), Pseudomonas aeruginosa as gram-negative Gram-positive bacteria (Pseudomonas aeruginosa KCTC 1637), Escherichia coli (E. Coli KCTC 1039), a yeast Candida yeast (Candida albicans KCTC 7965) and Aspergillus niger (KCTC 6910) were used as fungi. The experimental methods and results are as follows.

To determine the antimicrobial activity, the minimum inhibitory concentration was measured using the 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.

For more specific antibacterial test, 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 pre-cultured at 25 ° C for 2 days, The filamentous fungi were inoculated on a potato dextrose agar medium and pre-cultured for 7 days in a 25 ° C. incubator. Spores of filamentous fungi formed on the surface of the medium were recovered by using a smear rod and diluted in sterilized saline.

2 mL of each extract diluted to the appropriate concentration in 5% DMSO (Dimethylsulfoxide) physiological saline solution was added to the sterilized Patry Dish according to the samples and the experimental species, and each sample was diluted to a proper concentration in 5% physiological saline solution , And 2 mL of 5% DMSO physiological saline solution was added to the control group. Then, 18 mL of the tryptic soy agar medium and the 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.

In the case of bacteria, each pre-cultured bacterium was plated on each petri dish at a final concentration of about 1 × 10 ⁶ CFU / mL, 1 × 10 ⁵ CFU / mL for yeast, about 1 X 10 < ⁴ > CFU / mL. Bacteria were cultured in each petri dish at 37 ° C for 24 hours, yeast was cultured at 25 ° C for 3 days, the molds were cultured in an incubator at 25 ° C for 7 days, and colonies were observed in each compartment to determine the minimum The sample concentration was determined as the minimum inhibitory concentration (MIC), and the results are shown in Table 10. At this time, the minimum inhibitory concentration means that the smaller the value, the higher the antibacterial effect.

sample
Strain Minimum inhibitory concentration (MIC, ug / mL) Pretreatment O
Example 1 50%
Functional ethanol solvent Pretreatment X
Comparative Example 1 50%
Functional ethanol solvent Comparative Example 2 Comparative Example 3 Comparative Example 4 S. aureus 20 50 500 > 1,000 > 1,000 P. aeruginosa 100 250 > 1,000 > 1,000 > 1,000 E. Coli 30 100 > 1,000 > 1,000 > 1,000 C. albicans 100 500 > 1,000 > 1,000 > 1,000 A. niger 200 1,000 > 1,000 > 1,000 > 1,000

[Formulation Example 1 and Formulation Comparative Example 1]

The composition of the nutritional cream according to the sample of the Example was prepared as shown in Table 14 below.

number ingredient Formulation Example 1 Formulation Comparative Example 1 One Pro-type glycerin monostearate 2.0 2.0 2 Stearic acid 1.5 1.5 3 Cetearyl alcohol 2.2 2.2 4 Wax 1.0 1.0 5 Squalane 3.0 3.0 6 Hardened vegetable oil 1.0 1.0 7 Sorbitan stearate 0.6 0.6 8 Mineral oil 5.0 5.0 9 Polysorbate 60 1.5 1.5 10 Dimethicone 1.0 1.0 11 Trioctanoin 5.0 5.0 12 Betaine 3.0 3.0 13 Triethanolamine 1.0 1.0 14 glycerin 5.0 5.0 15 Sodium hyaruronate 3.0 3.0 16 Pretreatment O
Example 1 50% functional ethanol solvent 1.0 - 17 Distilled water Balance Balance 18 Preservative, fragrance, pigment a very small amount a very small amount

Among the components constituted by the compositions shown in Table 14, the aqueous components of raw materials 12 to 15 and 17 were completely dissolved by heating at 80 DEG C, and then raw materials 1 to 11 and 16 were heated to 80 DEG C to obtain a solution of the raw materials 12 to 15 and 17 And emulsified for 15 minutes at 3,000 rpm using a homomixer (Homo Mixer Mark II, Primix, JPN). The raw material 18 was then added, stirred for 5 minutes, and then cooled to room temperature.

[Test Example 10] Measurement of whitening effect of formulation application

The whitening effect of the nutritional cream prepared in Formulation Example 1 and Comparative Formulation Example 1 was measured by the following method. 30 women aged 25 years or older who had spots, freckles and pigmentosis were allowed to use the nutritional creams of Formulation Example 1 and Comparative Example 1 for 12 weeks, followed by the use of a Chromameter (CR-410, Minolta, Japan) To measure the change in skin color brightness (? L). It is calculated through the average value of 30 persons. The larger the brightness change value, the better the whitening effect. The experimental results are shown in Table 15 below.

medium
Formulation Example 1 Formulation Comparative Example 1
Skin color change (ΔL) 3.74 0.12

As shown in Table 15, it was confirmed that the nutritional cream of Formulation Example 1 to which the sample was added had much better whitening effect than the nutritional cream of Comparative Formulation 1, which was not added.

[Test Example 11] Measurement of wrinkle improvement effect

The wrinkle-improving effect of the nutritional cream prepared in Formulation Example 1 and Comparative Formulation Example 1 was measured by the following method. Ten women aged 30 or older undergoing skin aging were allowed to use the nutritional creams of Formulation Example 1 and Comparative Example 1 for 12 weeks on the crow feets, The wrinkle depth (μm) was measured by image analysis using a skin visiometer (SV-600, C + K, Germany) at weekly intervals and the results are shown in Table 16 below .

medium
Formulation Example 1
Formulation Comparative Example 1 Before application 318 ± 9 321 ± 11
After 12 weeks of application 271 ± 13 319 ± 13
Wrinkle depth reduction rate (%) 14.8 0.6

As a result, it was confirmed that the nutritional cream of Formulation Example 1 is superior to the nutritional cream of Formulation Comparative Example 1 in improving wrinkles.

[Test Example 12] Measurement of atopy improvement effect of application of formulation

The atopic improvement effect of the nutritional cream prepared in Formulation Example 1 and Comparative Formulation Example 1 was measured by the following method. Tape repeat stripping was used as a method of skin barrier damage. When the transepidermal water loss (TEWL) reaches 4.0 g / m ² / h by repeatedly stripping the tape to 20 persons over 25 years old, the nutritional cream of Formulation Example 1 and Comparative Example 1 . The transdermal water loss was measured by Tewameter 210, a vaporizer of C + K (Cologne, Germany). After 6 hours of application, the amount of percutaneous water loss was measured to measure the extent to which the amount of percutaneous water loss decreased, and the extent to which the skin barrier function was restored was measured. The recovery rate used in the efficacy evaluation was calculated as shown in [Equation 10] and the results are shown in Table 17 below.

&Quot; (10) "

B ( _t ) = (1- (B _{t = 6} - B _{t = 0} ) / (B _{t = d} - B _{t = 0} )

B _{t = 6} : Percutaneous water loss measurement value after 6 hours after skin barrier damage

B _{t = 0} : Percutaneous water loss measurement value before skin barrier damage

B _{t = d} : Percutaneous water loss measurement value immediately after skin barrier damage

Recovery rate (%) medium Formulation Example 1 Formulation Comparative Example 1 6 hour recovery rate
(Based on initial transdermal water loss) 53 11

As shown in the above results, the nutritional cream of Formulation Example 1 to which the sample of the present invention was added showed a remarkable increase in recovery effect after skin barrier damage as compared with the nutritional cream of Comparative Formulation 1, which was not added.

[Formulation Example 1 and Formulation Comparative Examples 1, 2, 3, 4]

The composition of the nutritional cream according to the sample of the Example was prepared as shown in Table 18 below.

number ingredient Formulation Example 1 Formulation Comparative Example 1 Formulation Comparative Example 2 Formulation Comparative Example 3 Formulation Comparative Example 4 One Pro-type glycerin monostearate 2.0 2.0 2.0 2.0 2.0 2 Stearic acid 1.5 1.5 1.5 1.5 1.5 3 Cetearyl alcohol 2.2 2.2 2.2 2.2 2.2 4 Wax 1.0 1.0 1.0 1.0 1.0 5 Squalane 3.0 3.0 3.0 3.0 3.0 6 Hardened vegetable oil 1.0 1.0 1.0 1.0 1.0 7 Sorbitan stearate 0.6 0.6 0.6 0.6 0.6 8 Mineral oil 5.0 5.0 5.0 5.0 5.0 9 Polysorbate 60 1.5 1.5 1.5 1.5 1.5 10 Dimethicone 1.0 1.0 1.0 1.0 1.0 11 Trioctanoin 5.0 5.0 5.0 5.0 5.0 12 Betaine 3.0 3.0 3.0 3.0 3.0 13 Triethanolamine 1.0 1.0 1.0 1.0 1.0 14 glycerin 5.0 5.0 5.0 5.0 5.0 15 Sodium hyaruronate 3.0 3.0 3.0 3.0 3.0 16-1 Pretreatment O
Example 1
50% functional ethanol solvent 1.0 - - - - 16-2 Pretreatment X
Comparative Example 1
50% functional ethanol solvent - 1.0 - - - 16-3 Comparative Example 2 - - 1.0 - - 16-4 Comparative Example 3 - - - 1.0 - 16-5 Comparative Example 4 - - - - 1.0 17 Distilled water Balance Balance Balance Balance Balance 18 Preservative, fragrance, pigment a very small amount a very small amount a very small amount a very small amount a very small amount

The aqueous components of the raw materials 12 to 15 and 17 were completely dissolved by heating to 80 DEG C among the components constituted by the composition of Table 18 and then the raw materials 1 to 11 and 16 were heated to 80 DEG C to obtain a solution of the raw materials 12 to 15 and 17 And emulsified for 15 minutes at 3,000 rpm using a homomixer (Homo Mixer Mark II, Primix, JPN). Thereafter, the raw material 18 was added, stirred for 5 minutes, and then cooled to room temperature.

[Test Example 13] Deformation measurement of cream

In order to measure the deformation of the nutritional cream prepared in Formulation Example 1 and Comparative Formulation Examples 1, 2, 3 and 4, physical properties and browning were measured after being left for a certain period of time according to storage conditions. Deformation (viscosity, browning, etc.) in a cream means that the product is unstable, easily phase separated, drops in value, and loses its meaning as a product. The experimental results are shown in [Table 19] and [Figure 1] below.

Name of sample
Daylight conditions 60 ℃ condition Early Four weeks Early Four weeks Viscosity color Viscosity color Viscosity color Viscosity color Formulation Example 1 62,500 White 62,000 White 62,500 White 62,500 White Formulation Comparative Example 1 58,400 Very light
yellow 52,300 Light
Brown 58,400 Very light
yellow 51,400 Light
Brown Formulation Comparative Example 2 44,700 Light
yellow 21,200 thick
Brown 44,700 Light
yellow 17,200 thick
Brown Formulation Comparative Example 3 52,200 Light
yellow 20,000 thick
Brown 52,200 Light
yellow 18,300 thick
Brown Formulation Comparative Example 4 55,400 Light
yellow 23,400 thick
Brown 55,400 Light
yellow 21,800 thick
Brown

Claims (13)

(A) a pretreatment step of eliminating an acid component and a sugar component which cause a galling and a blackening phenomenon at the time of refining by extracting hot water or refluxing the fruit of a non-treating fruit;
(B) an extraction step of extracting the saponin exhibiting the intermediate polar surface activity by reflux extraction with the ethanol-containing concentrated ethanol solvent having an ethanol content of 40 to 60% (v / v) obtained in the step (a); And
(C) a step of purifying saponin from the fruit juice which contains saponin by removing the pigment layer and impurities from the fruit juice extract obtained from step (b).
The method according to claim 1,
Wherein the step (c) is carried out using an adsorbent.
The method according to claim 1,
Wherein the step (a) is carried out by adding 1 to 10% by weight of a non-specific fruit to the solvent.
The method according to claim 1,
Wherein the step (b) is carried out by adding 1 to 10% by weight of the fruitless pretreatment extract to the functional ethanol solvent.
The method according to claim 1,
Wherein the saponin obtained through the step (c) has a saponin content of 85% by weight or more.
In a cosmetic composition containing an alopecic fruit extract,
The above-mentioned non-treated fruit extract
(A) a pretreatment step of eliminating an acid component and a sugar component which cause a galling and a blackening phenomenon at the time of refining by extracting hot water or refluxing the fruit of a non-treating fruit;
(B) an extraction step of extracting the saponin exhibiting the intermediate polar surfactant by reflux extraction with the alcoholic ethanol solvent having an ethanol content of 40 to 60% (v / v) obtained in the step (a); And
(C) a step of removing the pigment layer and impurities from the ethanol-free fruit extract obtained by the above step (b) to obtain saponin having a saponin content of 85% by weight or more, and having no browning phenomenon / RTI >
The method of claim 6,
Wherein the cosmetic composition is a cleansing detergent, a hair cleansing agent or a cleansing agent for body cleansing.
The method of claim 6,
Wherein the cosmetic composition is selected from cosmetics, gels, water-soluble liquids, creams, essences, oil-in-water (O / W) and water-in-oil (W / O) formulations.
The method of claim 6,
Wherein the cosmetic composition is an antioxidant functional cosmetic exhibiting an antioxidative effect.
The method of claim 6,
Wherein the cosmetic composition is a cosmetic composition exhibiting an effect of improving skin wrinkles.
The method of claim 6,
Wherein the cosmetic composition is a whitening functional cosmetic composition.
The method of claim 6,
Wherein the cosmetic composition is an antibacterial functional cosmetic composition.
The method of claim 6,
Wherein the cosmetic composition is an anti-inflammatory functional cosmetic composition.

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