Cosmetic composition for improving skin containing taraxacum coreanum phytoplacenta culture extract that has moisturizing and soothing effects for extremely dry skin such as atopic dermatitis, and skin barrier strengthening effect

Abstract

The present disclosure relates to a Taraxacum Coreanum phytoplacenta cell culture extract that has moisturizing and soothing effects for extremely dry skin such as atopic dermatitis, and skin barrier strengthening effect. The Taraxacum Coreanum phytoplacenta cell culture or an extract thereof according to the present disclosure is non-toxic to skin cells, and has an efficacy of moisturizing, anti-aging, strengthening the skin barrier, improving wrinkles, soothing the skin, protecting the skin from UV rays, and protecting the skin from blue light, etc.

Claims

1. A method for preparing a composition containing a phytoplacenta cell culture of Taraxacum Coreanum or an extract of the phytoplacenta cell culture, the method comprising the steps of: (a) separating and culturing phytoplacenta cells from the phytoplacenta tissue in an ovary of Taraxacum Coreanum plant; and (b) preparing a composition containing the phytoplacenta cell culture obtained in the step (a) or an extract of the phytoplacenta cell culture obtained in the step (a); wherein step (a) includes culturing the phytoplacenta cells in the dark; and wherein the extract of the phytoplacenta cell culture in the step (b) is obtained by extracting the phytoplacenta cell culture of Taraxacum Coreanum obtained in the step (a) at 60° C. to 80° C. for 6 to 8 hours.

2. The method of claim 1, wherein step (b) includes drying, pulverizing, and mixing the phyplacenta cell culture of Taraxacum Coreanum plant obtained in the step (a) with purified water.

3. The method of claim 2, wherein step (b) after mixing the phytoplacenta cell culture of Taraxacum Coreanum plant performing ultrasonic extraction or hot water extraction.

4. A cosmetic composition comprising a phytoplacenta cell culture of Taraxacum Coreanum or the extract of the phytoplacenta cell culture and a carrier acceptable in cosmetic formulation, wherein the phytoplacenta cell culture of Taraxacum Coreanum or the extract of the phytoplacenta cell culture is present in an amount ranging from 0.1 to 10% by weight, based on the total weight of the composition.

5. A cosmetic composition according to claim 4 comprising an effective amount of the phytoplacenta cell culture or the extract of the phytoplacenta cell culture to moisturize skin.

6. A cosmetic composition according to claim 4 comprising an effective amount of the phytoplacenta cell culture or the extract of the phytoplacenta cell culture to improve a skin barrier.

7. A cosmetic composition according to claim 4 comprising an effective amount of the phytoplacenta cell culture or the extract of the phytoplacenta cell culture to protect skin from ultraviolet rays.

8. A cosmetic composition according to claim 4 comprising an effective amount of the phytoplacenta cell culture or the extract of the phytoplacenta cell culture to protect skin from blue light.

9. A cosmetic composition according to claim 4 comprising an effective amount of the phytoplacenta cell culture or the extract of the phytoplacenta cell culture to prevent or alleviate skin stress.

10. A cosmetic composition according to claim 4 comprising an effective amount of the phytoplacenta cell culture or the extract of the phytoplacenta cell culture to protect skin from fine dust.

11. A cosmetic composition according to claim 4 comprising an effective amount of the phytoplacenta cell culture or the extract of the phytoplacenta cell culture to soothe the skin.

12. A cosmetic composition according to claim 4 comprising an effective amount of the phytoplacenta cell culture or the extract of the phytoplacenta cell culture to improve or prevent skin wrinkles.

13. The cosmetic composition according to claim 4, wherein the phytoplacenta cell culture of Taraxacum Coreanum or the extract of the phytoplacenta cell culture is present in an amount ranging from 1 to 10% by weight, based on the total weight of the composition.

14. The method of claim 1, wherein the extract of the phytoplacenta cell culture in the step (b) is obtained by extracting the phytoplacenta cell culture of Taraxacum Coreanum obtained in the step (a) at 60° C. to 80° C. for 6 to 8 hours at a pressure ranging from 0.03 to 0.3 MPa.

15. The method of claim 1, wherein the extract of the phytoplacenta cell culture in the step (b) is obtained by extracting the phytoplacenta cell culture of Taraxacum Coreanum obtained in the step (a) at 60° C. to 80° C. for 6 to 8 hours at a pressure ranging from 0.05 to 0.1 MPa.

16. A cosmetic composition comprising a phytoplacenta cell culture of Taraxacum Coreanum or the extract of the phytoplacenta cell culture and a carrier acceptable in cosmetic formulation, wherein the Phytoplacenta cell culture of Taraxacum Coreanum or the extract of the phytoplacenta cell culture is present in an amount ranging from 0.1 to 10% by weight, based on the total weight of the composition, wherein the phytoplacenta cell culture of Taraxacum Coreanum or the extract of the phytoplacenta cell culture is prepared by the method of claim 12.

17. The cosmetic composition according to claim 16, wherein the Phytoplacenta cell culture of Taraxacum Coreanum or the extract of the phytoplacenta cell culture is present in an amount ranging from 1 to 10% by weight, based on the total weight of the composition.

Description

BEST MODE

(1) Hereinafter, the present disclosure will be described in more detail through the following examples. However, these examples are for illustrative purposes only and the scope of the present disclosure is not limited to these examples.

Example 1: Preparation of Composition According to the Present Disclosure

Example 1-1: Surface Disinfection of Plants

(2) For plant cell production according to the present disclosure, the flower bud of Taraxacum Coreanum was carefully peeled off with tweezers to separate the phytoplacenta cells. Thereafter, the phytoplacenta cells were immersed in 70% ethanol for 30 seconds, and sterilized by shaking it in 2% sodium hypochlorite for 5 minutes, and then, was washed with sterile water.

Example 1-2: Induction of Plant Phytoplacenta Cell

(3) Sterilized phytoplacenta cells were cut at once using a sharp knife, and early plant cells were induced by culturing the cells in the dark in a basal MS medium (Murashige and Skoog 1962, Duchefa, Cat. M0221) under growth conditions at 25° C. and 70% humidity wherein 100 mg of myoinositol as plant growth regulators, 0.4 mg of thiamine, 0.1 mg of zeatin, 30 g of sucrose are included in the basal MS medium (based on 1 L). Each medium was adjusted to pH 5.8 with 1N NaOH. Subcultures were performed at two-week intervals.

Example 1-3: Plant Phytoplacenta Cell Culture and Mass Production According to the Present Disclosure

(4) The Taraxacum Coreanum phytoplacenta cells cultured through aseptic subculture were then constantly controlled at temperature 25° C., humidity 70%, air supply 0.1 vvm in a liquid medium of the same composition except for agar using a bioreactor, and ware mass-produced by culturing and proliferating at intervals of 14 days.

Example 1-4: Preparation of Plant Phytoplacenta Cell Composition According to the Present Disclosure

(5) The cultured Taraxacum Coreanum phytoplacenta cells were harvested, the moisture was sufficiently removed with a clean tissue, and then the obtained cells were dried at 60° C. for 2 days. In order to obtain a composition according to the present disclosure, 20 g of Taraxacum Coreanum phytoplacenta cells were extracted with 10 L of purified water at 60 to 80° C. for 6 to 8 hours under reduced pressure at low temperature, and filtered.

Experimental Example 1: Skin Cell Non-Irritating Effect Confirmation Test

(6) First, in order to find out whether the composition according to the present disclosure of Example 1 was stimulated by toxicity for each treatment concentration in skin cells, an MTT assay for measuring cell viability was performed.

(7) For this, HaCaT cells (keratinocytes) and Detroit cells (fibroblasts) were each dispensed to 96-well plate with DMEM (Dubelcco's Modified Eagle's Medium) containing 10% Fetal Bovine Serum (FBS) and 1% antibiotic-antimycotic and incubated for 24 hours in an incubator under the conditions of 5% CO.sub.2 and 37° C. Thereafter, the cells were treated with the composition as a control to which purified water was added and a test substance to be a final concentration of 1, 5, and 10%, and further cultured for 24 hours. Then, 5 mg/mL of a solution of MTT(3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide) was added to each well in an amount of 4 μL, and incubated for 4 hours to allow the reaction. After removing the culture medium, 100 μL of a solution of dimethyl sulfoxide (DMSO) was added to dissolve the cells, and the absorbance was measured at 540 nm.

(8) TABLE-US-00001 TABLE 1 Treatment Viability of Viability of concentration HaCaT cell Detroit cell Sample (%) (%) (%) Control — 100.00 100.00 Experimental 1 101.26 98.01 group 5 100.46 96.93 10 97.96 96.73

(9) As a result, when the composition according to the present disclosure was treated by concentration of 1 to 10%, all cell viability compared to the control group was 95% or more, which did not cause intradermal toxicity. Accordingly, it was confirmed that the “Taraxacum Coreanum phytoplacenta cell culture extract” has a low possibility of causing irritation to skin cells and is harmless to skin cells.

Experimental Example 2: Antioxidant Effect Confirmation Test from DPPH Radical Scavenging Ability

(10) In order to examine the antioxidant effect of the composition according to the present disclosure due to inhibition of 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical generation, DPPH assay was performed.

(11) For this, 500 μL of a 0.1 mM DPPH solution and 100 μL of the composition as a control and a test substance were added to 400 μL of ethanol. After mixing each sample, it was reacted for 30 minutes in the dark at room temperature, and the absorbance was measured at 517 nm.

(12) TABLE-US-00002 TABLE 2 Treatment DPPH radical concentration scavenging ability Sample (%) (%) Control — 0.00 Experimental 1 3.61 group 5 11.68 10 23.84

(13) As a result, it was confirmed that when the composition according to the present disclosure was treated by concentration each, the DPPH radical scavenging ability was increased compared to the control group, thereby confirming that the “Taraxacum Coreanum phytoplacenta cell culture extract” had an antioxidant effect.

Experimental Example 3: Antioxidant Effect Confirmation Test from H.SUB.2.O.SUB.2 .Oxidative Toxicity

(14) In order to examine the antioxidant effect from the H.sub.2O.sub.2 oxidative toxicity of the composition according to the present disclosure, the change in cell viability by H.sub.2O.sub.2 was investigated.

(15) To this end, HaCaT cells were dispensed to a 48-well plate, and then cultured for 24 hours in cell culture conditions. Thereafter, 1 mM H.sub.2O.sub.2 was irradiated, and the cells were treated with the composition as a control and a test substance, and further cultured for 12 to 16 hours. In this case, 2 mM of N-acetylcy-L-steine (NAC) was used as a positive control. Thereafter, cell viability was measured using the MTT assay to confirm whether the skin cells were protected from damage caused by the induction of H.sub.2O.sub.2 oxidative toxicity.

(16) TABLE-US-00003 TABLE 3 Treatment Viability of concentration HaCaT cell Sample (%) H.sub.2O.sub.2 (%) Control — − 100.00 Control — + 60.33 Positive control — + 88.8 Experimental 1 + 66.56 group 5 + 76.73 10 + 79.46

(17) As a result, as cell death by H.sub.2O.sub.2 treatment was inhibited in cells treated with the composition according to the present disclosure by concentration each compared to the control (+H.sub.2O.sub.2), it was confirmed that the “Taraxacum Coreanum phytoplacenta cell culture extract” had an antioxidant effect from H.sub.2O.sub.2 oxidative toxicity.

Experimental Example 4: Wrinkle Improvement Effect Confirmation Test According to Increase in PIP Production

(18) In order to examine the wrinkle improvement effect of the composition according to the present disclosure, the biosynthesis amount of procollagen (Procollagen Type I C-Peptide, PIP), which is a precursor of collagen synthesis, was measured.

(19) To this end, Detroit551 cells were dispensed to a 48-well plate, and then cultured for 24 hours in cell culture conditions. After that, the cells were treated with the composition as a control and a test substance, and further cultured for 48 hours. Then, a portion of 20 μL of the diluted supernatant of the culture medium was taken, and it was placed in an antibody-coated microtiter plate together with 20 μL of a PIP standard in a PIP EIA Kit (Takara, Cat. MK101), and 100 μL of a peroxidase-labeled antibody solution was also added to each well and reacted at 37° C. for 3 hours. After removing the medium and washing 4 times with 200 μL of PBS, 100 μL of a substrate solution was added to each well and reacted at room temperature for 15 minutes under light blocking. After that, a 100 μL of a substrate solution (1N H.sub.2SO.sub.4) was added, and the absorbance at 450 nm was measured.

(20) TABLE-US-00004 TABLE 4 Treatment Ability of concentration synthesizing PIP Sample (%) (%) Control — 100.00 Positive control — 122.60 Experimental 1 109.5 group 5 114.1 10 119.1

(21) As a result, as the amount of PIP increased in the cells treated with the composition according to the present disclosure by concentration each compared to the control, it was confirmed that the “Taraxacum Coreanum phytoplacenta cell culture extract” was effective in anti-wrinkle.

Experimental Example 5: Moisturizing and Skin Barrier Improvement Effect Confirmation Test According to Increase in Expression of AQP3 and FLG

(22) In order to examine the moisturizing and skin barrier improvement efficacy of the composition according to the present disclosure, changes in the expression levels of AQP3 (Aquaporin), a moisture/glycerol transporter, and FLG (Filaggrin), known as a natural moisturizing factor, were investigated.

(23) HaCaT cells were dispensed to a 96-well plate, and then cultured for 24 hours in cell culture conditions. After that, the cells were treated with the composition as a control and a test substance, and further cultured for 24 hours. Real-time PCR was performed to confirm the gene level of AQP3 and FLG, and the test sequence is as follows.

(24) For RNA isolation and cDNA synthesis, SuperPrepm cell lysis & RT Kit for qPCR (TOYOBO, Cat. SCQ-101) were used. The cells from which the medium was removed were washed once with PBS, and 50 μL of a cell lysis mixture was added and reacted for 5 minutes, followed by addition of a stop solution. 8 μL of the previously extracted lysate was added to 32 μL of the RT reaction mixture, and cDNA was synthesized using PCR at 37° C. for 15 minutes, 50° C. for 5 minutes, and 95° C. for 5 minutes. For comparative analysis of gene expression, the cDNA synthesized above was used as a template, and real-time PCR analysis was performed using Thunderbird™ SYBR qPCR Mix (TOYOBO, Cat. QPS-201). The primer used in the experiment was Qiagen's QuantiTect primer assays (GAPDH; Cat. QT01192646, AQP3; Cat. QT00212996, FLG; Cat. QT02448138), and the AQP3 and FLG mRNA expression levels of the samples were quantified by GADPH. Real-time qPCR conditions were first performed at 95° C. for 1 minute, followed by performing a total of 40 cycles at 94° C. for 15 seconds, 60° C. for 30 seconds, and 72° C. for 30 seconds per cycle.

(25) TABLE-US-00005 TABLE 5 Treatment Relative Expression concentration level (Fold) Sample (%) AQP3 FLG Control — 1.00 1.00 Experimental 1 1.49 1.83 group 5 2.37 1.77 10 2.11 1.66

(26) As a result, as the mRNA expression levels of AQP3 and FLG increased in cells treated with the composition according to the present disclosure by concentration each compared to the control group, it was confirmed that “Taraxacum Coreanum phytoplacenta cell culture extract” was effective in moisturizing and an improvement of the skin barrier.

Experimental Example 6: Anti-Inflammatory Effect Confirmation Test According to COX-2 Expression Reduction

(27) In order to examine the anti-inflammatory effect of the composition according to the present disclosure, a change in the expression level of COX-2 (Cyclooxygenase-2) involved in the inflammatory response was investigated.

(28) To this end, HaCaT cells were dispensed to a 96-well plate, and then cultured for 24 hours in cell culture conditions. Thereafter, an inflammatory response was induced through UVB irradiation, the cells were treated with the composition as a control and a test substance, and further cultured for 4 hours. Real-time PCR was performed to confirm the gene level of COX-2, and the test sequence is as follows.

(29) For RNA isolation and cDNA synthesis, SuperPrep™ cell lysis & RT Kit for qPCR (TOYOBO, Cat. SCQ-101) were used. The cells from which the medium was removed were washed once with PBS, and 50 μL of a cell lysis mixture was added and reacted for 5 minutes, followed by addition of a stop solution. 8 μL of the previously extracted lysate was added to 32 μL of the RT reaction mixture, and cDNA was synthesized using PCR at 37° C. for 15 minutes, 50° C. for 5 minutes, and 95° C. for 5 minutes. For comparative analysis of gene expression, the cDNA synthesized above was used as a template, and real-time PCR analysis was performed using Thunderbird™ SYBR qPCR Mix (TOYOBO, Cat. QPS-201). The primer used in the experiment was Qiagen's QuantiTect primer assays (GAPDH; Cat. QT01192646, COX-2; Cat. QT00040586), and the COX-2 mRNA expression levels of the samples were quantified by GADPH. Real-time qPCR conditions were first performed at 95° C. for 1 minute, followed by performing a total of 40 cycles at 94° C. for 15 seconds, 60° C. for 30 seconds, and 72° C. for 30 seconds per cycle.

(30) TABLE-US-00006 TABLE 6 Treatment concentration Relative Expression Sample (%) UVB level of COX-2(Fold) Control — − 0.129 Control — + 1.00 Experimental 1 + 0.88 group 5 + 0.67 10 + 0.54

(31) As a result, as the mRNA expression level of COX-2 decreased in the cells treated with the composition according to the present disclosure by concentration each compared to the control (UVB treatment), it was confirmed that the “Taraxacum Coreanum phytoplacenta cell culture extract” had an anti-inflammatory effect.

Experimental Example 7: Skin Cell Protective Effect Confirmation Test from UV Stimulation

(32) In order to examine the skin cell protective effect of the composition according to the present disclosure from UV stimulation, the change in cell viability was investigated.

(33) To this end, HaCaT cells were dispensed to a 96-well plate, and then cultured for 24 hours in cell culture conditions. After that, UV was irradiated, the cells were treated with the composition as a control and a test substance, and further cultured for 24 hours. Then, cell viability was measured by using the MTT assay to confirm whether the UV-induced damage to skin cells was protected.

(34) TABLE-US-00007 TABLE 7 Treatment Viability of concentration HaCaT cell Sample (%) UVB (%) Control — − 100.00 Control — + 40.46 Experimental 1 + 49.60 group 5 + 54.06 10 + 60.46

(35) As a result, as cell death by UVB treatment was inhibited in cells treated with the composition according to the present disclosure by concentration each compared to the control (+UVB), it was confirmed that “Taraxacum Coreanum phytoplacenta cell culture extract” had a cytoprotective effect from UV rays.

Experimental Example 8: Skin Cell Protective Effect Confirmation Test from Blue Light Induction

(36) In order to examine the skin cell protective effect of the composition according to the present disclosure from blue light stimulation, the change in cell viability was investigated.

(37) To this end, HaCaT cells were dispensed to a 96-well plate, and then cultured for 24 hours in cell culture conditions. After that, the cells were treated with the composition as the control and test substance, and after 2 hours, the cells were irradiated with blue light for 6 hours using Topview 5450 VRI SMD LED (ITSWELL, Cat. IWS-L5056-VRI-K3), and then the cells were further cultured for 24 hours. Then, cell viability was measured by using the MTT assay to confirm whether the blue light-induced damage to skin cells was protected.

(38) TABLE-US-00008 TABLE 8 Treatment Viability of concentration Blue HaCaT cell Sample (%) light (%) Control — − 100.00 Control — + 74.1 Experimental 1 + 82.01 group 5 + 88.43 10 + 89.62

(39) As a result, as cell death by blue light treatment was inhibited in the cells treated with the composition according to the present disclosure by concentration each compared to the control (+blue light), it was confirmed that “Taraxacum Coreanum phytoplacenta cell culture extract” had a cytoprotective effect from blue light.

Experimental Example 9: Anti-Aging Effect Confirmation Test According to TERT Expression Increase

(40) In order to examine the anti-aging efficacy of the composition according to the present disclosure, a change in the expression level of telomerase reverse transcriptase (TERT), which is known as an enzyme extending the length of telomeres at the ends of DNA strands, was investigated.

(41) A change in the expression level of telomerase reverse transcriptase (TERT), which is known as an enzyme that extends the length of telomeres at the ends of DNA strands, was investigated.

(42) HaCaT cells were dispensed to a 96-well plate, and then cultured for 24 hours in cell culture conditions. After that, the cells were treated with the composition as a control and a test substance, and further cultured for 24 hours. Real-time PCR was performed to confirm the gene level of TERT, and the test sequence is as follows.

(43) For RNA isolation and cDNA synthesis, SuperPrep™ cell lysis & RT Kit for qPCR (TOYOBO, Cat. SCQ-101) were used. The cells from which the medium was removed were washed once with PBS, and 50 μL of a cell lysis mixture was added and reacted for 5 minutes, followed by addition of a stop solution. 8 μL of the previously extracted lysate was added to 32 μL of the RT reaction mixture, and cDNA was synthesized using PCR at 37° C. for 15 minutes, 50° C. for 5 minutes, and 95° C. for 5 minutes. For comparative analysis of gene expression, the cDNA synthesized above was used as a template, and real-time PCR analysis was performed using Thunderbird™ SYBR qPCR Mix (TOYOBO, Cat. QPS-201). The primer used in the experiment was Qiagen's QuantiTect primer assays (GAPDH; Cat. QT01192646, TERT; Cat. QT00073409), and the TERT mRNA expression levels of the samples were quantified by GADPH. Real-time qPCR conditions were first performed at 95° C. for 1 minute, followed by performing a total of 40 cycles at 94° C. for 15 seconds, 60° C. for 30 seconds, and 72° C. for 30 seconds per cycle.

(44) TABLE-US-00009 TABLE 9 Treatment concentration Relative Expression Sample (%) level of TERT(Fold) Control — 1 Experimental 1 1.38 group 5 1.67 10 1.81

(45) As a result, as the mRNA expression level of TERT increased in the cells treated with the composition according to the present disclosure by concentration each compared to the control, it was confirmed that the “Taraxacum Coreanum phytoplacenta cell culture extract” was effective in anti-aging.

Experimental Example 10: Anti-Stress Effect Confirmation Test According to LEA Expression Increase

(46) In order to examine the anti-stress effect of the composition according to the present disclosure, the expression change of LEA (Late embryogenesis abundant), which is known to protect cell damage against stress, was investigated.

(47) To this end, HaCaT cells were dispensed to 6-well plate, and then cultured for 24 hours in conditions for culturing cell. After that, the cells were treated with the composition as a control and a test substance, and then the cells were further cultured for 24 hours, and an In-Cell ELISA Colorimetric Detection kit (Invitrogen, Cat No. 62200) was used to measure the LEA protein expression level. The finished culture medium was removed and the cells were fixed by incubating at room temperature for 15 minutes with 100 μL fixing solution. After that, it was washed with PBS, and 100 μL of a permeabilization solution was added and reacted at room temperature for 15 minutes. After that, it was washed with PBS, and 100 μL of quenching solution was added and reacted at room temperature for 20 minutes. After that, it was washed with PBS, and 100 μL of a blocking solution was added and reacted at room temperature for 30 minutes. After removing the solution, 50 μL of LEA primary antibody was added and reacted at 4° C. for 24 hours. After removing the antibody, it was washed with PBS, and 50 μL of HRP-conjugated secondary antibody was added and reacted for 1 hour. After removing the antibody and washing with PBS, 100 μL of a TMB substrate solution was added and reacted at room temperature for 15 minutes while blocking the light. Then, 100 μL of a stop solution (1N H.sub.2SO.sub.4) was added, and the absorbance at 450 nm was measured.

(48) TABLE-US-00010 TABLE 10 Treatment Ability for concentration synthesizing LEA protein Sample (%) UVB (%) Control — − 68.57 Control — + 100.00 Experimental 1 + 103.21 group 5 + 108.85 10 + 111.95

(49) As a result, as the expression level of LEA was increased by UVB treatment in cells treated with the composition according to the present disclosure by concentration each compared to the control (+UVB), it was confirmed that “Taraxacum Coreanum phytoplacenta cell culture extract” had a stress relieving effect by protecting cell damage.

Experimental Example 11: (In Vivo) Fine Dust Blocking Effect Confirmation Test

(50) The fine dust blocking effect of the essence formulation to which 1% of the composition according to the present disclosure was applied was investigated.

(51) First, the essence formulation was prepared, in the case of the experimental group, with 1 part by weight of a Taraxacum Coreanum phytoplacenta cell culture extract, 1 part by weight of betaine, 0.5 parts by weight of dipalmitoylhydroxyproline, 0.3 parts by weight of 1,2 hexanediol, 0.2 parts by weight of carbomer, 0.1 parts by weight of propylene glycol, 0.05 parts by weight of sorbitol, 0.02 parts by weight of sodium hydroxide, and 100 parts by weight of the remaining purified water, and a control of essence formulation is that 1 part by weight of a Taraxacum Coreanum phytoplacenta cell culture extract which is a composition according to the present disclosure was excluded from the experimental group.

(52) For the test, a square (2 cm×2 cm) was partitioned on the forearms of 20 subjects, the product was applied in an amount of 50 μL each, and the test site was exposed to a fine dust floating chamber equipped with a propeller for 1 minute. Substitute fine dust was carbon black (Korea Carbon Black Co., Ltd., Corax N220, particle size: 20 to 25 nm), and 1 g was suspended in the chamber. Measurements were carried out before adsorption of alternative fine dust and after adsorption of alternative fine dust. For photography, a digital camera (Canon, EOS70D) was used to take a picture of the forearm. The photographed images were analyzed using the image analysis program (Image-Pro® plus) to designate the Area of Interest (AO′) of the test site, and then analyze the difference in pixel values before and after adsorption of fine dust. The unit is an Arbitrary Unit (A.U.).

(53) TABLE-US-00011 TABLE 11 Treatment concentration Fine Pixel change Sample (%) dust level Control 1 — − 0.00 Control 2 — + 20.01 Experimental 1 + 14.65 group

(54) Improvement (%) of the effect of preventing the adsorption of the substituting fine dust in the experimental group compared to the control group=[1−(the change level of the substituting fine dust pixel in the experimental group)/(the change level of the substituting fine dust pixel in the control group 2)]*100

(55) As a result, the experimental group to which the composition according to the present disclosure was applied showed a 26.79% improvement in the prevention of fine dust adsorption compared to the control group 2. Therefore, “Taraxacum Coreanum phytoplacenta cell culture extract” is judged to be a product that helps preventing the adsorption of fine dust.

Experimental Example 12: (In Vivo) Skin Soothing Effect Confirmation Test

(56) The skin soothing effect according to the skin temperature reduction rate of the essence formulation to which 1% of the composition according to the present disclosure was applied, was investigated.

(57) For this purpose, a square (2 cm×2 cm) was partitioned on the forearms of 25 subjects, and the product was applied in an amount of 50 μL each, and then exposed to direct sunlight for 20 minutes before the test. Measurement was performed using a computerized infrared thermographic imaging sensor (DITI) to measure body heat at intervals of 0 minute (before sample application), 5 minutes (after sample application), and 10 minutes (after application of sample), and the temperature reduction rate was calculated.

(58) TABLE-US-00012 TABLE 12 Treatment Heat reduction rate (%) concentration After 0 After 5 After 20 Sample (%) minute minutes minutes Control — 7.1 3.1 0.9 Experimental 1 7.3 5.5 3.8 group

(59) As a result, it was confirmed that the continuous heat reduction rate was maintained even after 20 minutes compared to the control group in the experimental group to which the composition according to the present disclosure was applied. Therefore, it was confirmed that “Taraxacum Coreanum phytoplacenta cell culture extract” had a skin soothing effect.

(60) As described above, a specific part of the content of the present disclosure has been described in detail, and for one skilled in the art, it will be obvious that this specific technology is only a preferred embodiment, and the scope of the present disclosure is not limited thereby. Therefore, it will be said that the substantial scope of the present disclosure is defined by the appended claims and their equivalents.