COMPOSITION CONTAINING EXTRACT OF LILIUM HANSONII LEICHTLIN EX BAKER

Abstract

An extract of Lilium hansoni Leichtlin ex Baker, according to the present invention, exhibits a whitening effect by inhibiting the expression of tyrosinases and reducing the total amount of melanin, reduces wrinkles and improves skin elasticity by promoting collagen synthesis of fibroblasts in the skin and inhibiting the activity of collagenases, exhibits an effect of anti-glycation that inhibits glycation, exhibits a skin moisturization effect by promoting the synthesis of hyaluronic acid, exhibits an effect of improving skin texture by reducing skin roughness, and exhibits effects of irritation relief and relief of stinging, itching, burning sensations and the like, thereby being usable in the preparation of cosmetic products, pharmaceuticals and foods.

Claims

1. A cosmetic composition for skin whitening, wrinkle improvement, elasticity enhancement, skin soothing, antiglycation, skin moisturization, or skin texture improvement, comprising a Lilium hansonii extract or a fraction thereof as an active ingredient.

2. The cosmetic composition of claim 1, wherein the Lilium hansoni extract is an extract obtained by extracting Lilium hansonii with water, an organic solvent, or a combination thereof.

3. The cosmetic composition of claim 2, wherein the organic solvent is a lower alcohol having 1 to 6 carbon atoms, a polyhydric alcohol, a hydrocarbon-based solvent, or a combination thereof.

4. The cosmetic composition of claim 1, wherein the Lilium hansoni fraction is a fraction obtained from the Lilium hansonii extract using: a polar solvent such as a lower alcohol having 1 to 6 carbon atoms, acetone, or ethyl acetate; a non-polar solvent such as ether, chloroform, benzene, hexane, or dichlorohexane; or a mixed solvent thereof.

5. The cosmetic composition of claim 1, wherein the Lilium hansonii extract or the fraction thereof is included in an amount of 0.0001 parts by weight to 10 parts by weight based on 100 parts by weight of the composition.

6. A pharmaceutical composition for skin whitening, wrinkle improvement, elasticity enhancement, skin soothing, antiglycation, skin moisturization, or skin texture improvement, comprising a Lilium hansonii extract or a fraction thereof as an active ingredient.

7. An external preparation for skin whitening, wrinkle improvement, elasticity enhancement, skin soothing, antiglycation, skin moisturization, or skin texture improvement, comprising a Lilium hansonii extract or a fraction thereof as an active ingredient.

8. A food composition for skin whitening, wrinkle improvement, elasticity enhancement, skin soothing, antiglycation, skin moisturization, or skin texture improvement, comprising a Lilium hansonii extract or a fraction thereof as an active ingredient.

9. A method of achieving skin whitening, wrinkle improvement, elasticity enhancement, skin soothing, antiglycation, skin moisturization, or skin texture improvement, which includes a step of applying composition comprising a Lilium hansonii extract or a fraction thereof as an active ingredient to the skin.

10. The method of claim 9, wherein the Lilium hansonii extract is an extract obtained by extracting Lilium hansonii with water, an organic solvent, or a combination thereof.

11. The method of claim 10, wherein the organic solvent is a lower alcohol having 1 to 6 carbon atoms, a polyhydric alcohol, a hydrocarbon-based solvent, or a combination thereof.

12. The method of claim 9, wherein the Lilium hansonii fraction is a fraction obtained from the Lilium hansonii extract using: a polar solvent such as a lower alcohol having 1 to 6 carbon atoms, acetone, or ethyl acetate; a non-polar solvent such as ether, chloroform, benzene, hexane, or dichlorohexane; or a mixed solvent thereof.

13. The method of claim 9, wherein the Lilium hansonii extract or the fraction thereof is included in an amount of 0.0001 parts by weight to 10 parts by weight based on 100 parts by weight of the composition.

14. A method of achieving skin whitening, wrinkle improvement, elasticity enhancement, skin soothing, antiglycation, skin moisturization, or skin texture improvement, which includes a step of administering composition comprising a Lilium hansonii extract or a fraction thereof as an active ingredient to the subject.

Description

DESCRIPTION OF DRAWINGS

[0088] FIG. 1 shows the results of a questionnaire for evaluating a degree of skin improvement after having 11 women in their 30s to 50s apply an essence containing a Lilium hansonii extract to their entire face twice a day for four weeks.

MODES OF THE INVENTION

[0089] Hereinafter, the present invention will be described in detail through Examples. However, the protection scope of the present invention is not limited by the Examples described below.

<Examples 1 to 3> Preparation of Lilium hansonii Extracts

[0090] The whole plant of Lilium hansonii (Example 1), roots thereof (Example 2), and flowers thereof (Example 3) were provided, and they were extracted at room temperature for three days by adding a 70% aqueous ethanol solution having a mass of 10 times their own mass and then filtered under reduced pressure. The filtered extracts were concentrated and dried using a rotary evaporator (Buchi, Switzerland) to obtain Lilium hansonii extracts.

<Comparative Examples 1 to 3> Preparation of Lilium Genus Plant Extracts

[0091] Whole plants of Lilium candidum (Comparative Example 1), Lilium siberia (Comparative Example 2), and Lilium martagon (Comparative Example 3) were extracted at room temperature for three days by adding a 70% aqueous ethanol solution having a mass of 10 times their own mass and then filtered under reduced pressure. The filtered extracts were concentrated and dried using a rotary evaporator (Buchi, Switzerland) to obtain respective extracts.

<Experimental Example 1> Confirmation of Skin Wrinkle Improvement and Elasticity Enhancement Effects

[0092] 1) Effect of Increasing Total Collagen Amount

[0093] The Lilium hansonii extracts and the three types of Lilium genus plant extracts were added to a culture medium containing human-derived fibroblasts, and then their effects of increasing a total collagen amount at the cellular level were compared.

[0094] The increased total collagen amount was quantified using a Procollagen Type I C-Peptide Enzyme ImmunoAssay kit (PICP EIA kit). Before testing, the cytotoxicity of the Lilium hansonii extracts prepared in Examples 1 to 3 and the Lilium genus plant extracts prepared in Comparative Examples 1 to 3 against human-derived fibroblasts was evaluated at various extract concentrations (μg/ml), and after selecting a concentration (100 μg/ml) not exhibiting cytotoxicity, a degree of increase in a total collagen amount was evaluated.

[0095] After adding each sample to a culture medium containing human-derived fibroblasts and culturing for one day, the culture medium was collected and a degree of increase in a total collagen amount at each concentration was measured at 450 nm using a PICP EIA kit and a spectrophotometer. For the comparison of effects, a degree of increase in a total collagen amount was measured in the same manner for a fibroblast culture medium to which no sample was added (control) and a sample to which vitamin C was added at a final concentration of 52.8 μg/ml. The total collagen amount was measured as UV absorbance, and a rate of increase in the total collagen amount was calculated as a ratio of a total collagen amount relative to that of the control, and the results are summarized in Table 1 below.

TABLE-US-00001 TABLE 1 Sample Collagen increase rate (%) Control (no additive) — Vitamin C (52.8 μg/ml) 26 Example 1 (100 μg/ml) 32 Example 2 (100 μg/ml) 35 Example 3 (100 μg/ml) 34 Comparative Example 1 (100 μg/ml) 19 Comparative Example 2 (100 μg/ml) 11 Comparative Example 3 (100 μg/ml) 13 *Total collagen amount increasing effect (number of repetitions = 3)

[0096] As can be seen from the results of Table 1, Lilium hansonii extracts have an excellent effect of increasing a total collagen amount by promoting collagen synthesis, and exhibited a total collagen amount increasing effect similar or superior to vitamin C and the Lilium genus plant extracts, which are known to be effective for increasing a total collagen amount, at the test concentrations.

[0097] 2) Effect of Inhibiting Collagenase Activity

[0098] The collagenase activity inhibitory effects of the Lilium hansonii extracts or the three types of Lilium genus plant extracts were evaluated as follows.

[0099] Fibroblasts, which are normal human skin cells, were inoculated in a 24-well microplate so that there were 2.5×10.sup.4 cells per well, and cultured in a 10.sup.0% serum DMEM medium under the condition of 37° C. for 24 hours. Subsequently, the 10% serum DMEM medium was removed, and the fibroblasts were washed once with a phosphate buffer solution and then further cultured for 30 minutes in a serum-free DMEM medium supplemented with one of Examples 1 to 3 and Comparative Examples 1 to 3. In this case, as a control, fibroblasts were additionally cultured for 30 minutes in a serum-free DMEM medium without extracts.

[0100] 30 minutes after sample treatment, fibroblasts were stimulated with 10 ng/ml of tumor necrosis factor-α (TNF-α), which is a material known to produce MMP-1, a collagenase, and cultured for 24 hours. In this case, among the controls not including the Lilium hansonii extract of the present invention, a TNF-α untreated control and a TNF-α treated control were used as a TNF-α untreated group and a TNF-α treated group, respectively.

[0101] After collecting supernatant from each well, the amount (ng/ml) of newly synthesized MMP-1 was measured using an MMP-1 assay kit (Amersham, USA), and a collagenase activity inhibition rate was determined by calculating an MMP-1 production inhibition rate (%) according to Mathematical Formula 1 below, and the results are shown in Table 2 below.

[0102] In the following Mathematical Formula 1, the amount of MMP-1 in the negative control refers to the amount of MMP-1 in the TNF-α untreated group, and the amount of MMP-1 in the positive control refers to the amount of MMP-1 in the TNF-α treated group, and the amount of MMP-1 in an experimental group refers to the amount of MMP-1 in a group in which one of Examples 1 to 3 and Comparative Examples 1 to 3 was added at each concentration.

[00001]$\begin{array}{cc}\mathrm{MMP}-1\mathrm{production}\mathrm{inhibition}\mathrm{rate}\left(\%\right)=\left(1-\frac{X-Z}{Y-Z}\right)\times 100& \left[\mathrm{Mathematical}\mathrm{Formula}1\right]\end{array}$$\left(X=\mathrm{MMP}-1\mathrm{amount}\mathrm{in}\mathrm{experimental}\mathrm{group},y=\mathrm{MMP}-1\mathrm{in}\mathrm{positive}\mathrm{control},Z=\mathrm{MMP}-1\mathrm{amount}\mathrm{in}\mathrm{negative}\mathrm{control}\right)$

TABLE-US-00002 TABLE 2 Sample Inhibition rate (%) Negative control (TNF-α untreated) — Positive control (TNF-α treated) — Example 1 (100 μg/ml) 75.5 Example 2 (100 μg/ml) 80.4 Example 3 (100 μg/ml) 82.1 Comparative Example 1 (100 μg/ml) 44.3 Comparative Example 2 (100 μg/ml) 36.1 Comparative Example 3 (100 μg/ml) 26.0 *Collagenase activity inhibitory effect (number of repetitions = 3)

[0103] Referring to the results of Table 2, when Examples 1 to 3 and Comparative Examples 1 to 3 were evaluated, it can be seen that the Lilium hansonii extracts were about two to three times more effective in inhibiting MMP-1, which is a collagenase, than the Lilium genus plant extracts of the same concentration.

<Experimental Example 2> Confirmation of Skin Whitening Effect

[0104] 1) Effect of Inhibiting Tyrosinase

[0105] A tyrosinase enzyme extracted from mushrooms and commercially available from SIGMA was used. First, tyrosine, a substrate, was dissolved in distilled water to make a solution having a concentration of 0.3 mg/ml, 1.0 ml of the solution was input into each test tube, and 1.0 ml of a potassium-phosphate buffer solution (concentration: 0.1 mol, pH: 6.8) and 0.7 ml of distilled water were added thereto. 0.2 ml of a sample solution, which had been prepared by adding one of Examples 1 to 3 of the present invention and Comparative Examples 1 to 3 to an ethanol solution at a final concentration of 100 μg/ml, was added to a reaction solution and then reacted in a 37° C. thermostat for 10 minutes. In this case, a control was prepared by adding only 0.2 ml of an ethanol solution instead of the sample solution, and vitamin C was used as a positive control. After adding 0.1 ml of a 2,500 units/ml tyrosinase solution, the reaction solution was again reacted in a 37° C. thermostat for 10 minutes. The test tube containing the reaction solution was quenched by placing in ice water to stop the reaction, and absorbance was measured at a wavelength of 475 nm using a photoelectric spectrometer, and the results are shown in Table 3 below. The tyrosinase inhibitory effect of each sample was calculated according to the following Mathematical Formula 2.

[00002]$\begin{array}{cc}\mathrm{Tyrosinase}\mathrm{inhibition}\mathrm{rate}\left(\%\right)=100-\left(\frac{\mathrm{Reaction}\mathrm{absorbance}\mathrm{of}\mathrm{sample}}{\mathrm{Reaction}\mathrm{absorbance}\mathrm{of}\mathrm{control}}\times 100\right)& \left[\mathrm{Mathematical}\mathrm{Formula}2\right]\end{array}$

TABLE-US-00003 TABLE 3 Sample Tyrosinase inhibition rate (%) Control (no additive) 0 Vitamin C (52.8 μg/ml) 50 Example 1 (100 μg/ml) 59 Example 2 (100 μg/ml) 60 Example 3 (100 μg/ml) 58 Comparative Example 1 (100 μg/ml) 25 Comparative Example 2 (100 μg/ml) 34 Comparative Example 3 (100 μg/ml) 33 *Tyrosinase inhibitory effect (number of repetitions = 3)

[0106] Referring to Table 3, it can be confirmed that the Lilium hansonii extracts had a tyrosinase inhibitory effect, and in particular, at a concentration of 100 sg/ml, Examples 1 to 3 had a higher inhibitory effect than ascorbic acid (vitamin C) known as a tyrosinase inhibitor, so an excellent whitening effect was exhibited.

[0107] 2) Confirmation of Effect of Reducing Total Melanin Amount

[0108] Each of Examples 1 to 3 and Comparative Examples 1 to 3 was added to a mouse melanoma cell (B-16 mouse melanoma cell) culture medium, and a whitening effect thereof at the cellular level was tested.

[0109] Each extract was tested after being added to the culture medium at a final concentration of 100 μg/ml, and arbutin, which was a control, was added to the medium at a concentration of 100 μg/ml. B-16 melanoma cells were treated with the experimental group or the control and then cultured for three days.

[0110] Subsequently, the cultured cells were treated with trypsin, removed from the culture vessel, and then centrifuged to extract melanin. The removed cells were treated with 1 ml of a sodium hydroxide solution (1 N) and then boiled for 10 minutes to dissolve melanin, and absorbance was measured at 400 nm using a spectrophotometer to measure the amount of produced melanin.

[0111] The amount of melanin was measured as absorbance per unit cell number (10.sup.6 cells), and a ratio thereof relative to the total melanin amount of the control was calculated as an inhibition rate (%), and the results are summarized in Table 4.

TABLE-US-00004 TABLE 4 Total melanin Inhibition Sample amount (Abs) rate (%) Control (no additive) 0.254 — Arbutin (100 μg/ml) 0.126 50.4 Example 1 (100 μg/ml) 0.113 55.5 Example 2 (100 μg/ml) 0.112 55.9 Example 3 (100 μg/ml) 0.125 50.8 Comparative Example 1 (100 μg/ml) 0.199 21.7 Comparative Example 2 (100 μg/ml) 0.177 30.3 Comparative Example 3 (100 μg/ml) 0.217 14.6 *Total melanin amount reduction effect (number of repetitions = 3)

[0112] Referring to the results of the above Table 4, it can be confirmed that all Examples related to a Lilium hansonii extract had a whitening effect. In particular, it can be seen that Examples 1 to 3 had a superior melanin inhibitory effect as compared to arbutin, which is known to be effective for whitening, of the same concentration and were also twice or more effective than the Lilium genus plant extracts of Comparative Examples.

<Experimental Example 3> Confirmation of Skin Moisturizing Effect (1)

[0113] 1) Confirmation of Effect of Promoting Hyaluronic Acid Synthesis

[0114] Human dermal fibroblasts were cultured at a concentration of 1×10.sup.5 cells/mL in a DMEM medium containing 10% fetal bovine serum and, after dispensing 500 μl thereof into each well of a 24-well plate, cultured for 18 hours. Subsequently, each sample was diluted in a DMEM medium not containing fetal bovine serum, and each sample was added at various concentrations. As a positive control, an epidermal growth factor (EGF) known to promote hyaluronic acid production was treated at a concentration of 10 ppb. After culturing for 24 hours, a cell culture medium was recovered, and the concentration of hyaluronic acid was measured using a Hyaluronan ELISA kit (R&D Systems).

[0115] In addition, the hyaluronic acid concentration of each experimental group treated with each sample was quantified based on the hyaluronic acid concentration (100%) of a control not containing an additive, and the results are shown in the following Table 5.

TABLE-US-00005 TABLE 5 Sample Hyaluronic acid (%) No additive (control) 100 EGF 10 ng/ml (10 ppb) (positive control) 150.9 Example 1 (100 μg/ml) 170.3 Example 2 (100 μg/ml) 190.8 Example 3 (100 μg/ml) 160.5 Comparative Example 1 (100 μg/ml) 108.9 Comparative Example 2 (100 μg/ml) 116.7 Comparative Example 3 (100 μg/ml) 114.2 *Hyaluronic acid synthesis promoting effect (number of repetitions = 3)

[0116] Referring to the above Table 5, it can be confirmed that the synthesis of hyaluronic acid was increased in all Examples related to a Lilium hansonii extract. In particular, in the case of Examples 1 to 3, higher concentrations and superior efficacy as compared to the case of the positive control were exhibited, and it can be seen that the effect was excellent as compared to the case of Comparative Examples, which are extracts of other Lilium genus plants.

<Preparation Example 1> Manufacture of Essence Including Lilium hansonii Extract

[0117] To confirm the skin improvement effect of the Lilium hansonii extracts prepared in the above Examples 1 to 3, essences were manufactured according to compositions of the following Table 6.

TABLE-US-00006 TABLE 6 Ingredient (parts by weight) A B C D Lilium hansonii extract 2 (Example 1) 2 (Example 2) 2 (Example 3) 0 Purified water To 100 To 100 To 100 To 100 Ethanol 10 10 10 10 Hexanediol 2 2 2 2 Glycerin 1 1 1 1 PEG-150 1 1 1 1 Dipropylene glycol 1 1 1 1 Glycereth-26 1 1 1 1 Sodium hyaluronate 4 4 4 4 PEG-8 4 4 4 4 Sodium citrate 1 1 1 1 Citric acid 0.5 0.5 0.5 0.5 Fragrance 0.5 0.5 0.5 0.5 Total 100 100 100 100

<Experimental Example 4> Confirmation of Skin Moisturizing Effect (2)

[0118] 1) Change in Skin Moisture Content Before and After Using Essence

[0119] To test a skin moisturizing effect, skin evaluation was performed while using the essences manufactured in the above Preparation Example 1. As the evaluation subjects, 20 women in their 30s and 40s were randomly divided into five people per group, and each group was asked to apply one of essences A to D to their entire face, and two weeks later, the moisture content of the skin was measured. Specifically, the 20 subjects were asked to wash their faces with soap, and then their skin was acclimatized to constant temperature and humidity conditions (temperature: 20±2° C., relative humidity: 40±2%). Using a corneometer (CM825), the capacitance of the skin surface was repeatedly measured three times. The average results are shown in Table 7.

TABLE-US-00007 TABLE 7 Rate of change in moisture content after two weeks (%) A 23 B 30 C 28 D 7 *Change in skin moisture content

[0120] Referring to the above Table 7, when essences (A to C) containing a Lilium hansonii extract were used for two weeks, the moisture content of the skin surface was significantly higher than in the case of the control (D). Therefore, it can be seen that a Lilium hansonii extract has an excellent moisturizing effect.

<Experimental Example 5> Confirmation of Skin Texture Improvement Effect

[0121] 1) Confirmation of Skin Texture Improvement Effect on Human Skin

[0122] A sensory evaluation was performed to confirm the skin texture improvement effect of the essences manufactured in Preparation Example 1. After recruiting 40 women in their 30s to 50s, 10 subjects were randomly selected and asked to apply one of the essences A to D to their entire face twice a day for four weeks. After four weeks of use, a degree of improvement in skin roughness was scored on a scale of 1 to 10 to compare skin texture improvement effects. Here, scores close to 1 represent a low skin texture improvement effect, and scores close to 10 represent an excellent skin texture improvement effect. An average score for each essence is shown in the following Table 8.

TABLE-US-00008 TABLE 8 Skin texture improvement effect (out of 10) A 7.8 B 8.0 C 8.2 D 6.5

[0123] Referring to Table 8, it can be seen that the skin texture improvement effect is greater when an essence (A, B, or C) including a Lilium hansonii extract is used rather than when an essence (D) not including a Lilium hansonii extract is used.

[0124] 2) Questionnaire Evaluation of Degree of Skin Improvement

[0125] A questionnaire evaluation was performed to confirm a skin improvement effect of essence A of the above-described Preparation Example 1. 11 women in their 30s to 50s were asked to apply A to their entire face twice a day for four weeks. After four weeks of use, a questionnaire was filled out to evaluate a degree of skin improvement, and the results are shown in FIG. 1. FIG. 1 shows the percentage of subjects who responded “yes” or “strongly yes.”

[0126] Referring to FIG. 1, it can be seen that 70% or more of users who used essence A of Preparation Example 1 including a Lilium hansonii extract experienced an excellent skin texture improvement effect.

<Experimental Example 6> Confirmation of Antiglycation Effect

[0127] To confirm an antiglycation efficacy, glycation inhibitory activity was measured using L-arginine and glucose.

[0128] A solution of 1 M L-arginine and 1 M glucose in 1 M phosphate buffer (pH 7.4) was first prepared and diluted with 1 M phosphate buffer so that the concentration of the sample was 100 μg/ml. 1 M L-arginine and 1 M phosphate buffer were mixed in a ratio of 1:4, and 80 μl thereof was dispensed into each well of a 96-well plate. To this, each sample, which was diluted to 100 μg/ml, and 100 μl of 0.01 M aminoguanidine, which was used as a positive control, were added. After sufficiently mixing the sample, finally, glucose which was diluted with 1 M phosphate buffer so that the final concentration of glucose was 0.1 M was added and then reacted at 70° C. for four hours. In the 96-well plate, absorbance was measured at 420 nm using a spectrophotometer to measure a degree of glycation.

[0129] The glycation experimental group of Mathematical Formula 3 is an experimental group in which glycation was induced by adding 1 M L-arginine and 1 M glucose, and to measure the absorbance of a sample itself, absorbance was measured at 420 nm after adding only 1 M L-arginine and the sample and no glucose.

[0130] The glycation inhibitory activity can be calculated according to the following Mathematical Formula 3. Each group was tested three times, and an average value was obtained.

[00003]$\begin{array}{cc}\mathrm{Glycation}\mathrm{inhibition}\mathrm{rate}\left(\%\right)=\frac{\left(\begin{array}{c}\mathrm{Absorbance}\mathrm{of}\mathrm{sample}\mathrm{after}\mathrm{reaction}-\\ \mathrm{Absorbance}\mathrm{of}\mathrm{sample}\mathrm{before}\mathrm{reaction}\end{array}\right)}{\mathrm{Glycation}\mathrm{experimental}\mathrm{group}}*100& \left[\mathrm{Mathematical}\mathrm{Formula}3\right]\end{array}$

TABLE-US-00009 TABLE 9 Sample Glycation inhibition rate (%) Control (DMSO, 50 μg/ml) — Positive control (Aminoguanidine, 55 μg/ml) 54.89 Example 1 (50 μg/ml) 48.24 Example 2 (50 μg/ml) 40.55 Example 3 (50 μg/ml) 50.18 Comparative Example 1 (50 μg/ml) 21.35 Comparative Example 2 (50 μg/ml) 11.83 Comparative Example 3 (50 μg/ml) 17.22 *Glycation inhibitor effect (number of repetitions = 3)

[0131] Referring to the above Table 9, it can be confirmed that all Examples related to a Lilium hansonii extract had an antiglycation effect, and when compared to aminoguanidine known as an antiglycation material, the antiglycation effect was similar to that of aminoguanidine. Therefore, it can be expected that skin troubles caused by a final glycation product will be improved by the excellent antiglycation effect of a Lilium hansonii extract.

<Experimental Example 7> Confirmation of Skin Soothing Effect

[0132] 1) Confirmation of Irritation Relieving Effect

[0133] To confirm an irritation relieving effect of a Lilium hansonii extract, an effect of inhibiting prostaglandin E2 (PGE2), which is a material related to irritation, was evaluated.

[0134] First, to quantify PGE2, a RAW 264.7 mouse macrophage cell line (KCLB no 40071, Korean Cell Line Bank, Seoul, Korea) was purchased, and 100 μl was dispensed into each well of a 96-well microplate at a cell inoculation concentration of 2.5×10.sup.6 cells/ml. As a medium, DMEM (GibcoBRL, Grand Island, N.Y., USA) supplemented with 10% FBS (GibcoBRL, Grand Island, N.Y., USA) and 1% penicillin/streptomycin (Thermo Scientific Hyclone, Waltham, Mass., USA) was used, and culture was carried out under the conditions of 5% CO.sub.2 and 37° C. In the cell culture medium in which the above-described RAW 264.7 cells had been dispensed, 1 μg/ml of lipopolysaccharide (LPS) (Sigma, St. Louis, Mo., US) was treated to induce irritation, and one of Examples 1 to 3 and Comparative Examples 1 to 3 was treated. Subsequently, the cells were cultured for 24 hours, centrifuged at 900 rpm and 4° C. for five minutes, and then the cell culture medium was recovered to measure the amount of PGE2 produced in the culture medium. The PGE2 was quantified using a commercially available PGE2 ELISA kit (MyBioSource Co., Ltd., San Diego, Calif., USA), according to the manufacturer's instructions.

TABLE-US-00010 TABLE 10 PGE2 production PGE2 inhibition Sample amount (pg/mL) rate (%) Untreated 323 — Example 1 (50 μg/ml) 165 48.9 Example 2 (50 μg/ml) 157 51.4 Example 3 (50 μg/ml) 161 50.2 Comparative Example 1 (50 μg/ml) 205 36.5 Comparative Example 2 (50 μg/ml) 211 34.7 Comparative Example 3 (50 μg/ml) 228 29.4 *Skin irritation relief effect (number of repetitions = 3)

[0135] As a result, it can be seen that all of Examples 1 to 3 related to a Lilium hansonii extract had an effect of reducing PGE2 production. That is, it was confirmed that a Lilium hansonii extract had the ability to alleviate skin irritation through having the effect of inhibiting PGE2 production.

[0136] 2) Confirmation of Stinging, Itchiness, and Hotness Soothing Effect

[0137] To confirm the skin soothing effect of a Lilium hansonii extract, irritation was induced, and then whether stinging, itchiness, or hotness was relieved was evaluated.

[0138] A chamber containing a 0.2% aqueous sodium laureth sulfate (SLES) solution was first attached to the arms of 15 healthy adult males and 15 healthy adult females, and after eight hours had elapsed, the Finn chamber was removed, and the subjects were asked to evaluate the stinging, itchiness, and hotness they experienced based on the criteria shown in the following table. The evaluation criteria are shown in the following Table 11.

[0139] <Skin Soothing Effect Evaluation Criteria>

TABLE-US-00011 TABLE 11 Score Criteria 1 No irritation 2 Very little irritation 3 Distinct irritation 4 Strong irritation 5 Very strong irritation to the extent of causing pain

[0140] Subsequently, the subjects were asked to apply a randomly chosen one of the previously produced essences A to D once a day (at 9 am) for three days to the skin area where SLES had been applied and then evaluate the stinging, itchiness, and hotness they were experiencing. The results are as follows.

TABLE-US-00012 TABLE 12 Evaluation item: Stinging Day 0 Day 1 Day 2 Day 3 A 4.2 2.0 1.4 1.2 B 2.0 1.8 1.6 C 2.0 1.6 1.6 D 3.6 3.6 3.0

TABLE-US-00013 TABLE 13 Evaluation item: Itchiness Day 0 Day 1 Day 2 Day 3 A 4.6 2.2 1.6 1.2 B 2.0 1.4 1.4 C 2.0 1.8 1.6 D 4.0 3.8 3.6

TABLE-US-00014 TABLE 14 Evaluation item: Hotness Day 0 Day 1 Day 2 Day 3 A 3.0 1.2 1.0 1.0 B 1.6 1.4 1.0 C 1.4 1.4 1.2 D 2.8 2.4 2.0

[0141] Referring to the above Tables 12 to 14, it can be seen that after using essences A to C including a Lilium hansonii extract, the degree of stinging, itchiness, and hotness was greatly reduced. On the other hand, in the case of essence D not including a Lilium hansonii extract, it can be seen that the difference between day 0 and day 3 was insignificant.