KR20050106194A - Extract and composition for treating atopy-dermatitis - Google Patents

Abstract

The present invention relates to compositions and mixtures containing illite for the treatment of atopic dermatitis, consisting of an illite, an extract of sewage, an extract of a leaf leaf or a mixed composition thereof.

The extract or composition according to the present invention has various functions to reduce the sensitive allergic response of the living body. According to the present invention, it is possible to provide a therapeutic agent for atopic dermatitis which does not have side effects such as skin weakness, systemic hormonal symptoms, addictiveness, insomnia, anxiety, and loss of appetite by conventional prescription for atopic dermatitis.

Description

Extract and Composition for Treating Atopy Dermatitis

The present invention relates to extracts and compositions for the treatment of atopic dermatitis.

Atopy dermatitis is an allergic disease that aggravates the stratum corneum, the outermost skin barrier, due to genetic and environmental immunological causes. Atopic dermatitis is a complex entanglement of a variety of causes, repeated relief and recurrence. Allergic diseases caused by atopy predisposition include allergic dermatitis, allergic rhinitis, asthma, allergic conjunctivitis, and atopic urticaria.

Atopic dermatitis suffers from a significant number of people, with 0.5 to 1 percent of the population and 5 to 10 percent of children suffering from atopic dermatitis. 50% of patients heal within two stones, but 25% go to adolescence, and the remaining 25% will continue without atopic dermatitis even in adults.

The cause of this atopic dermatitis is not well known, but it has been found to be mainly related to genetic factors and immune system deficiency. In addition, dry skin, the characteristics of skin itching easier than normal people, infections caused by bacteria, viruses, fungi, etc., emotional factors, environmental factors seems to be caused by a combination of each other.

The main symptoms of atopic dermatitis are severe itching, dry skin, rashes, rashes, crusts and scaly skin. Above all, itching is characterized by severe itching.

Because atopic dermatitis cannot be fundamentally repaired, atopic dermatitis is a disease that is controlled by proper treatment and avoiding the triggers rather than aiming at cure. Prescriptions for atopic dermatitis are mainly drug therapy such as steroids, antihistamines and antibiotics. Steroids (adrenal corticosteroids) have anti-inflammatory and immunosuppressive effects and have excellent effects, but when applied for a long time, side effects such as skin weakness, systemic hormonal symptoms, and addictive effects may occur. Antihistamines can be used as a temporary measure to relieve itching by preventing histamine from being released from mast cells, which can cause side effects such as insomnia, anxiety and loss of appetite.

Therefore, there is a need for a new concept of atopic dermatitis treatment composition that is effective against atopic dermatitis and has no side effects.

It is an object of the present invention to provide a novel atopic dermatitis treatment material and composition which is excellent in atopic dermatitis and does not have side effects of conventional therapeutic agents.

In order to achieve the object as described above, the inventors conducted a series of two in-vitro experiments, in which a total of 18 single medicinal herb extracts, herbal compositions, and single mineral (ilite) extracts were found to be effective in atopic dermatitis. Materials were selected. The selected extracts, compositions and illite were then applied directly to the animals (NCNga; atopic dermatitis mice) to confirm that the effect was substantially superior to atopic dermatitis.

Accordingly, the present invention relates to a composition for treating atopic dermatitis, which contains illite as an active ingredient. The composition of the present invention may additionally contain sewage extract and / or leaf extract.

The extract according to the present invention may be prepared by first obtaining and drying the sewage or leaf which has been selected according to a conventional method and trimmed to an appropriate size by cold water extraction, hot water extraction or solvent extraction, respectively. For example, it can be manufactured by the following method. First, an appropriate amount of pure water is added to the selected sewage or season, and hot water extraction is performed at 100 ° C. for 3 hours, and then rapidly frozen at minus 40 ° C. for 10-20 hours, vacuum-dried for at least 35 hours at a pressure of 0.23 Torr or less, and then Dry powder can be obtained. In preliminary experiments, extracts or compositions obtained through cold water extraction, hot water extraction or solvent extraction under various conditions showed substantially the same in vitro and in vivo effects. Therefore, the following examples will be described based on hot water extraction.

The illite is a fine mica group belonging to a monoclinic system having a chemical composition of (K, H ₃ O) Al ₂ (Si, Al) ₄ O ₁₀ (H ₂ O, OH) ₂ . I) As a mineral, it exhibits characteristics such as heavy metal adsorption, toxic gas adsorption, far infrared radiation and anion generation. Ilite has a very favorable molecular structure for absorption and adsorption of organic substances, and it is effective for joint pain, muscle pain, arthritis, ulcers, insect bites and burns as well as purification and detoxification effects. It may also be added.

Hasao (何) is a vine perennial herb with a dicotyledonous plant, the herbaceous perennial plant, and its reddish brown tuber is called sewage in one shot. It is used as a tonic, tonic and laxative. Hasuo said, "Inflammation, removing phlegm and phlegm, curing various boils, hemorrhoids, chronic fatigue, postpartum sickness, treatment of women's postpartum, and treatment of kidneys and blood, strengthening the muscles, The bone marrow is faithful (consensus), and "protects the liver and kidneys and clears the blood (herbaceous, synonymical dictionary)." Recent pharmacological experiments show that there are tonic action, hematopoietic function strengthening, fatigue recovery promoting action, and sedative action.

Persimmon leaves contain vitamin C, flavonoid glycosides, resins, tannins, carotene, organic acids, sugars, essential oils, chlorophyll and many other ingredients. Vitamin C is known to be 20 times higher than lemons. From ancient times, the leaflet is used to lower the blood pressure and improve the symptom of feeling up.

The composition according to the present invention, as can be seen in the following examples and applications, exhibits a superior and safe therapeutic effect compared to conventional atopic dermatitis therapeutic drugs. The composition may be used as a therapeutic agent for atopic dermatitis and as a dietary supplement for treatment and prevention, in which case various supplemental herbal medicines, vitamins, minerals, dietary fiber and other medicinal-food supplements (eg, excipients) may be used in the composition. It will be apparent to those skilled in the art that these can be mixed with, enhancers, coatings, etc.).

Hereinafter, the present invention will be described in more detail in Examples and Application Examples. The following examples are merely illustrative of the present invention in detail, and thus, the nature of the technical idea of the present invention is not changed or reduced in scope. It will be apparent to those skilled in the art that various embodiments and applications not shown in the following examples are possible. The following application examples were performed only with respect to the composition of the illite, sewage extract, leaflet extract alone and (illite + sewage extract + leaflet extract), but all of them showed a significant effect, which was not tested in the application ( It can be expected that the illite + sewage extract) mixed composition and the (ilite + leaf extract) mixed composition also show significant results.

EXAMPLES Preparation of Extracts or Compositions According to the Present Invention

(1) Preparation of the light

The illite collected from Jeungsan Kaolin Mine in Mudu 2-ri, Nam-myeon, Jeongseon-gun, Gangwon-do, Korea was purchased and used. The particle diameter was 2000-3000 mesh.

(2) Preparation of sewage or leaf extract

10 L of water was mixed with 0.3 Kg of dry sewage or leaf which had been cut to an appropriate size in advance and then hot water extracted at 100 ° C. for 3 hours. The hot water extract was then filtered and the filtrate was left for 1 hour at 4 ° C. refrigeration and concentrated for 10-15 hours on a 60 m water bath at 700 mHg pressure. The concentrated solution was rapidly frozen at -40 ° C for 15 hours, vacuum-dried for more than 35 hours at a pressure of 0.23 Torr or less, and then rapidly frozen at -40 ° C for 1 hour. When the water was completely evaporated, the residue was triturated to obtain dried sewage or dry leaf extract powder, respectively.

(3) Preparation of mixed composition of sewage and leaf extract

The sewage and the leaf obtained in (1) are mixed in the same weight ratio, or the dried sewage and the leaf which have been previously selected and cut to an appropriate size are mixed in the same weight ratio, and then extracted and dried by the same method as in (1). A composition powder was obtained.

In the present embodiment, sewage and leaf were made to the same weight ratio, but according to a preliminary experiment separately, even if the weight ratio of sewage and leaf was 1: (0.3 to 3), the object of the present invention was satisfied.

(4) Preparation of Sewage, Leaf and Illite Mixture Composition

900 ml of pure water was added to 1 parts by weight of the mixture of sewage and leaf obtained in (1) or (2), respectively, and 25 parts by weight of illite was added little by little while heating and stirring at 100 ° C to prepare a mixture of sewage, leaf and illite. Prepared.

In the present embodiment, for the sake of convenience, the sewage: leaflet: illite is 1: 1: 25 weight ratio, but according to a preliminary experiment, the sewage: leaflet: illite weight ratio is 1: (0.3-3) :( 1-50) even if The effect intended by the present invention was shown.

Experimental Method for Application Example 1

(1) Cell culture

HMC monoculture, HMC and anti-human IgE (10ug / ml) co-culture, HMC and anti-human IgE (10ug / ml) and the above Example to examine the reaction mechanism for the extract after pre-culture of HMC cells Substances obtained in (Ilite and illite and sewage, a mixture of leaf extract) and the like and ointment (1.0, 10, 100ug / ml) were co-cultured. Cultivation was carried out for 48 hours at 37 ℃ while maintaining a constant concentration of 5% CO ₂ in 10% FBS, DMEM medium.

Treatment of the extract or commercial ointment according to the present invention was specifically carried out as follows.

HMCs at 5 × 10 ⁴ cells were co-cultured with various concentrations of candidate medicinal herbs and anti-human IgE (10ug / ml) in DMEM medium supplemented with 10% FBS (37 ° C, 5% CO ₂ concentration, 48h). ). Radiolabeled Methyl- [ ³ H] thymidine (1 Ci per well) was added to the cell populations cultured for 48 hours. After 44 hours, 96-well plates cultured cells were labeled with 3H-thymidine (Amersham Pharmacia Biotech, UK, specific radioactivity 5.0 Ci / mmol) for 4 hours (final concentration 0.5 Ci) to be introduced into the DNA of proliferated HMCs. The cells of each sample were collected with Skatron Instruments Cell Harvester, washed with distilled water and collected on glass fiber filter papers (Skatron, Lier, Norway). Filter discs containing high molecular weight material including DNA were transferred to scintillation minivials and then Ready Safe scintillation fluid (Beckman Instruments, Fullerton, CA) was added. The amount of intracellular radioactive material was measured using a Beckman LS5000CE scintillation spectrometer (Duclos et al.).

(2) RT-PCR Performance and SDS Gel Electrophoresis

Total cellular RNA was isolated from the cultured HMCs in a conventional manner, and the purity of RNA was determined based on the A260: A280 ratio, followed by agarose gel electrophoresis to confirm the integrity of the RNA.

Using the isolated and purified RNA as a template, RT-PCR (Quantitative reverse transcriptase-PCR) was performed under the following conditions: 94 ° C, 5 min, and then [94 ° C, 1 min; 57 ° C. for the optimal annealing temperature for cytokines as described elsewhere, 1 min; 72 cycles, 1 min] at 30 cycles, and the final reaction was 72 캜 for 10 min. The same amount of PCR result was confirmed by ethidium bromide staining.

According to the purpose of PCR, each of the following praigo sets was applied.

① Human β-actin oligonucleotide sequence

5'-gTggggCgCCCCAggCACCA-3 '

5'-CTCCTTAATgTCACgCACgATTTC-3

② Human IL-4 oligonucleotide sequence

5'-GGGTCTCACCTCCCAACTGCT-3 '

5'-CGAACACTTTGAATATTTCTCTCTC-3 ';

③ Human IL-6

5'-ATGAACTCCTTCTCCACAAGCGC-3 ';

5'-GAAGAGCCCTCAGGCTGGACTG-3 '.

④ Human TNF-α

5'-AGCGGCTGACTGAACTCAGATTGTTAG-3 '

5'-GTCACAGTTTTCAGCTGTATAGGG-3 '

RT-PCR products were electrophoresed and stained in a conventional manner to observe bands, and the relative intensity of each lane band was measured on an electrophoretic gel.

(3) Determination of TNF-α and histamine concentrations in the supernatant

The supernatant was separated by centrifuging the cell culture at 1500 g for 20 minutes. TNF-α concentration was measured in the supernatant separated by ELISA (Endogen, Woburn, MA) method. The histamine RIA kit was used to analyze the histamine concentration in the supernatant.

Experimental Method 2 for Application Example

(1) total RNA isolation

The facial area of the experimental animal (NCNga mouse) was collected, and skin tissue was removed, and skin tissue (0.1 g) and RNAzol ^B 500 μl were mixed and ground until dissolved. 50 µl of chloroform was added to the mixed suspension, followed by mixing for 15 seconds. The mixture was left on ice for 15 minutes and then centrifuged at 13,000 rpm to recover about 200 μl of the supernatant. After 200 μl of 2-propanol was added to the supernatant, the mixture was slowly shaken and allowed to stand on ice for 15 minutes. This was again centrifuged at 13,000 rpm, washed with 80% EtOH and dried for 3 minutes in a vaccum pump to extract RNA. The extracted RNA was dissolved in 20 μl distilled water treated with diethyl pyrocarbonate (DEPC), inactivated in 75 ℃ heating block, and used for first strand cDNA synthesis.

(2) reverse transcriptase-polymerase chain reaction

3 μg of the total RNA prepared as described above was denatured at 75 ° C. for 5 minutes, followed by 2.5 μl of 10 mM dNTPs mix, 1 μl of random sequence hexanucleotides (25 pmole / 25 μl), and 1 μl of RNase inhibitor as RNA inhibitor. (20 U / μl), 1 μl of 100 mM DTT, 4.5 μl of 5 × RT buffer (250 mM Tris-HCl, pH 8.3, 375 mM KCl, 15 mM MgCl ₂ ), then 1 μl of M-MLV RT (200 U / μl) was added again and the final volume was 20 μl as DEPC treated distilled water. After mixing the reaction mixture well, centrifugation was performed at 2,000 rpm for 5 seconds to react for 60 minutes in a 37 ° C constant temperature water bath to synthesize first-strand cDNA, and then left at 95 ° C for 5 minutes to inactivate M-MLV RT. Synthesized cDNA was used for polymerase chain reaction (PCR).

(3) cDNA PCR

PCR was performed using a Primus 96 Legal PCR system (with high pressure lid, MWG in germany). The reaction was performed using 3 μl of the synthesized cDNA as a template, and primers for the template were sense primer (20 pmole) to amplify β-actin, IL-6, TNF-α, cyclooxygenase-2, and NOS-II genes. / Μl) and antisense primer (20 pmole / μl) were added, and 1 μl was added, followed by 3 μl of 2.5 mM dNTPs, 3 μl of 10 × PCR buffer (100 mM Tris-HCl, pH 8.3, 500 mM KCl, 15). mM MgCl ₂ ), and 0.18 μl of Taq polymerase (5 U / μl) were added, followed by addition of sterile distilled water to a final volume of 30 μl and pre-denaturation; 95 ° C., 5 minutes, denaturation; 95 ° C., 5 minutes, annealing; 55 ° C., 1 min, elongation; After 25 cycles of 72 ° C and 1 minute, PCR was performed under post-elongation conditions at 72 ° C for 3 minutes.

Each PCR product was loaded onto 1.2% agarose gel by 20 ㎕ and analyzed by electrophoresis for 20 min at 120 V.

Application Example 1 of the extract or composition according to the present invention in vitro  Test

Performed indirectly a therapeutic effect on atopic dermatitis in illite and extract obtained in Example (in vitro) treatment effect by applying the substance to the anti-human various concentrations of IgE with the human HMC-I cells allergy is induced in order to measure Was assayed. In each of the tables below, "normal" refers to the group not treated with anti-human IgE, "active" refers to the group treated with anti-human IgE, and "ilite" refers to the group treated with the illite mentioned in the above examples. "Sewage" refers to the group treated with the sewage extract obtained in the Example, "single leaf" means the group treated with the leaflet extract obtained in the Examples, 100, 10, 1 is the treated concentration of the extract (㎍ / Ml).

(1) RT-PCR analysis of IL and TNF-α

Atopic dermatitis is a type of allergic disease, and one of the allergic reactions is an increase in the expression of interleukin (IL) and tumor necrosis factor (TNF). Therefore, the degree of inhibition of the production of interleukin and tumor necrosis factor in the treatment of the substance according to the present invention was analyzed.

In order to analyze the expression level of IL4, IL6, IL13 and TNF-α of each treatment group through RT-PCR analysis, RT-PCR was performed by separating the total RNA of each treatment group according to the above-described RT-PCR analysis method. Then, electrophoresis was performed and the relative intensity of the band brightness was measured.

Results of electrophoresis after RT-PCR of IL4, IL6, IL13 and TNF-α are shown in FIGS. 1 to 4, respectively, and the relative intensities of these band brightnesses were measured and summarized in Table 1 below.

normal activation Illite Shouao Leaf 100 10 One 100 10 One 100 10 One IL4 81 134 41 88 128 35 78 144 21 144 167 IL6 75 196 61 151 165 140 149 199 39 175 164 IL13 58 150 81 107 - 63 99 - - - - TNF-α 142 163 101 125 124 20 41 172 29 106 104

As can be seen from the diagram, the treatment of the illite according to the present invention, the sewage extract and the leaflet extract mixed with the illite, respectively, significantly reduced the production of interleukin, and each 100 μg / ml treatment caused an allergic reaction. The production of interleukins was reduced to levels that did not occur. TNF-α also reduced production in the same manner.

Therefore, it can be seen that the mixed composition of illite, iliat and sewage extract, and leaf extract according to the present invention can reduce (treat) allergic reactions such as atopic dermatitis.

(2) TNF-α concentration measurement

In the above (1), it was confirmed that the illite, sewage extract, and leaf extract according to the present invention significantly reduced the amount of RNA transcription of TNF-α. Subsequently, direct protein analysis was performed to determine whether the production of TNF-α was substantially reduced. It was. The substance dose was 100 μg / ml. The unit of expression amount is µg / ml.

The analysis results are shown in Table 2 and FIG. 5 below.

Expression average Deviation Confidence interval (95%) Statistical significance (95%) Statistical significance (99%) normal 26 4.5 (17.18, 34.82) Note Note activation 720 96 (531.84, 908.16) Illite 356 49 (259.96, 452.04) Note Note Shouao 45 7.8 (29.712, 60.288) Note Leaf 78 13.7 (51.148, 104.852) Note Note

As a result of measuring the concentration of TNF-α secreted into the culture by ELISA (Endogen, Woburn, MA), as shown in the table and FIG. 5, the results were almost identical to the RNA gene expression experiment (RT-PCR). In other words, it was found that the amount of TNF-α secretion was significantly decreased in the illite treatment group, sewage extract treatment group and the leaflet extract treatment group compared to the active group (statistically significant under 99% significance level).

Therefore, it was reconfirmed that the mixed composition such as the illite, illite and sewage extract, and leaf extract according to the present invention can reduce (treat) allergic reactions such as atopic dermatitis.

(3) histamine protein concentration measurement

In fact, histamine poisoning and allergic diseases have similar symptoms, and histamine is known to be effective in treating allergic diseases. Therefore, it was analyzed how much inhibition of the production and secretion of histamine during the treatment of the present invention. The extract dose was 100 μg / ml. The unit of expression amount is µg / ml.

The analysis results are shown in Table 3 below and FIG. 6.

Expression Average Deviation Confidence interval (95%) Statistical significance (95%) Statistical significance (99%) normal 7.4 2.4 (2.696, 12.104) Note Note activation 48 12.3 (23.892, 72.108) Illite 21 3.4 (14.336, 27.664) Note Note Shouao 10.5 1.6 (7.364, 13.664) Note Leaf 15.7 2.5 (10.8, 20.6) Note Note

As can be seen from the diagram, the same as in the case of TNF-α, the illite treatment group, sewage extract treatment group and the leaflet extract treatment group showed that the histamine secretion significantly decreased compared to the active group (statistically 99% Significant under the significance level).

Therefore, it was confirmed that the mixed composition such as illite, illite and sewage extract, and leaf extract according to the present invention can reduce (treat) allergic reactions such as atopic dermatitis.

Application Example 2 of the extract or composition according to the present invention in vivo  Test

EXAMPLES The therapeutic effect of the obtained material for atopic dermatitis was directly confirmed ( in vivo ). In order to compare the effect, the coating ointment (Dormaiko-chi ointment) used for atopic dermatitis in Japan was used as a control. The therapeutic effects were assayed by applying the substances and ointments obtained in the above examples to various concentrations of human HMC-I cells induced by allergy with anti-human IgE.

Six-week-old females of NCNga mice (Japan SLC, Shizuoka, Japan), atopic dermatitis mice, were housed at 23 ± 2 ° C (55 ± 15%) for 12 hours to 12 hours (light-dark cycle). Experiments were performed after two weeks of adaptation in the environment.

EXAMPLES The obtained material and ointment were orally administered (75 mg / kg) three times a week (month, Wednesday, Friday) for 6 weeks to 8-week-old NCNga mice, and at the same time, the skin was applied to the neck (10 mg / day). For oral administration, 75 mg / kg of the drug, such as extract, was dissolved in 0.1 cc of pure water, and 10 mg of the drug, such as extract, was mixed with 200 mu l of castor oil.

In the following tables, "control" means no treatment group, "illight" means the group treated with the illite mentioned in the above examples, and "sewage" means the group treated with the sewage extract obtained in the examples. , "Leaf" means the group treated with the leaf extract obtained in Example, "composition" means the group treated with the composition obtained in Example (4), and "ointment" means the group treated with the Japanese atopy treatment.

(1) RT-PCR analysis of IL6

In order to confirm the expression level of IL6 in the experimental animal tissues according to the method mentioned in "Experimental Method 2 for Application" described above, RT-PCR was performed, followed by electrophoresis, and the relative intensities of the bands were measured. After electrophoresis of IL6 after RT-PCR, photographs of the results are shown in FIG.

Expression Control 114 Illite 53 Ointment 124 Shouao 31 Leaf 72 Composition 15

As can be seen in the chart, the illite, sewage extract, leaf extract, and the culture medium of the experimental animals treated with the composition, the expression level of IL6 was reduced to almost 1/2 level compared to the control group. In contrast, the "ointment" treatment group showed more expression than the control group.

Therefore, it can be seen that the illite, sewage extract, leaf extract and mixed compositions thereof according to the present invention can reduce (treat) allergic reactions such as atopic dermatitis.

(2) IgE concentration measurement

Atopic dermatitis is a hypersensitivity reaction of the immune system that occurs when harmful substances such as allergens enter. When harmful chemicals enter the body, B lymphocytes in the blood make antibody protein E (IgE) and chemical transporters such as histamine and prostaglandin. Releases.

Therefore, the concentration of IgE in experimental animal serum was measured to confirm whether the substance according to the present invention can reduce the production of antibody protein.

Experimental animals 8, 10, 12, and 15 weeks of blood using a capilery tube from the eye of the mouse was collected about 100 μl of blood and serum was isolated to measure the amount of IgE (Fig. 8 and Table 5). The unit is μg / ml.

Week 8 Week 10 Week 12 Week 15 Average Standard Deviation Average Standard Deviation Average Standard Deviation Average Standard Deviation 95% level of significance Control 621.6 60.2 948.8 78.6 1169 163.2 1778 178 Illite 381.9 87.7 617.2 71.9 688.7 183.2 1361 306 Ointment 363.2 62.2 643.6 75.8 938.1 121 1027 93 Note Shouao 457.6 131.3 811.6 142.3 1320.7 161.7 1509 281 Leaf 318.2 55.7 932.7 273 882.5 178 1165 300 Composition 296.2 64.3 347.1 151.8 309.9 42.9 867 260 Note

As can be seen in the chart, the illite, extract and composition treatment group according to the present invention showed a significantly lower IgE concentration than the control group in the entire experiment. In particular, the composition treatment group according to the present invention showed a significantly superior IgE production inhibitory effect compared to the "ointment" treatment group, which is a conventional treatment for atopic dermatitis. Ointment treatment group and composition treatment group showed significance at 95% significance level.

Therefore, it was confirmed that the illite, sewage extract, leaf extract and mixed compositions thereof according to the present invention are effective in reducing (treating) allergic reactions such as atopic dermatitis.

(3) IL-6 concentration measurement

At 15 weeks, NCNga mice were anesthetized with ethyl ether, blood was collected by cardiac puncture, and serum was separated. IL-6 was quantified in serum by ELISA kit method (FIG. 9 and Table 6). The unit is μg / ml.

Average Standard Deviation 95% significance level Control 19.2 1.6 Illite 3.5 1.1 Note Ointment 8.5 2.4 Note Shouao 7.8 1.6 Note Leaf 5.2 1.1 Note Composition 7.6 2.7 Note

As can be seen in the figures and tables, the substance treatment group according to the present invention showed significantly lower blood IL-6 concentrations than the control group, and all showed significance at the 95% significance level.

Therefore, it was reaffirmed that the elite, sewage extract, leaf extract, and mixed compositions thereof according to the present invention were effective in reducing (treating) atopic dermatitis.

(4) IgG2a concentration measurement

At 15 weeks, NCNga mice were anesthetized with ethyl ether, blood was collected by cardiac puncture, and serum was separated. Mab-Based Mouse Ig isotyping kit (PharMingen, San Diego, Calif) was used to measure the concentration of immunoglobulins IgG2a in serum. (Figure 10 and Table 7). The unit is μg / ml.

Average Standard Deviation 95% significance level Control 4.25 0.51 Illite 2.04 0.23 Note Ointment 3.21 0.72 Shouao 2.44 0.31 Note Leaf 1.87 0.29 Note Composition 2.68 0.26 Note

As can be seen in the table and table, the illite, extract and composition treatment group according to the present invention showed a significantly lower blood IgG2a concentration than the control group. In particular, "ointment" was not significant at the 95% significance level, all of the illite, extract and composition treatment group according to the present invention showed a significance.

Therefore, it could be seen that the illite, sewage extract, leaf extract and mixed compositions thereof according to the present invention are effective in reducing (treating) atopic dermatitis.

(5) Tissue culture analysis of biological samples

After 15 weeks, the facial and neck areas of NCNga mice were collected, 1g of skin tissue was removed, washed with DMEM medium, mixed with chopping, and cultured in Petri-dish with 10% FBS-DMEM. After 7 days the culture supernatant was removed and replaced with fresh 10% FBS-DMEM culture. After culturing for 7 days, the culture supernatant was separated and IL-13 concentration in the culture was measured by ELISA kit (FIG. 11 and Table 8). The unit is μg / ml.

Average Standard Deviation Control 268.7 41.1 Illite 102.7 4.1 Ointment 108.3 4.3 Shouao 150.1 2.9 Leaf 95.5 32.3 Composition 134 31

As can be seen in Figures and Tables, the tissue culture solution of the substance treatment group according to the present invention showed an IL-13 concentration in the culture medium is significantly lower than the tissue culture solution of the control group.

Therefore, it could be confirmed that the illite, sewage extract, leaf extract and mixed composition thereof according to the present invention are effective in reducing (treating) atopic dermatitis.

According to the present invention, a new concept of atopic dermatitis treatment which is effective for atopic dermatitis from natural products that have been recognized for safety and stability becomes possible. Therefore, it is possible to provide atopic dermatitis treatment without side effects such as skin weakness, systemic hormonal symptoms, addictiveness, insomnia, anorexia, and loss of appetite by conventional prescription for atopic dermatitis.

1 to 4 are photographs showing the results of electrophoresis after RT-PCR of IL4, IL6, IL13 and TNF-α, respectively, from total RNA of human HMC-1 cells treated with a therapeutic agent according to the present invention.

5 is a chart showing the concentration of TNF-α in human HMC-1 cell culture treated with a therapeutic agent according to the present invention.

6 is a chart showing the concentration of histamine in human HMC-1 cell culture treated with a therapeutic agent according to the present invention.

Figure 7 is a photograph of the results of electrophoresis after performing RT-PCR of IL6 in a treatment animal tissue treated with the present invention.

Figure 8 is a chart showing the change in the concentration of IgE in serum with time in the experimental animals treated with the present invention.

9 is a chart showing the concentration of IL-6 in serum in experimental animals treated with a therapeutic agent according to the present invention.

10 is a chart showing the concentration of IgG2a in serum in experimental animals treated with a therapeutic agent according to the present invention.

11 is a chart showing the concentration of IL-13 in the tissue separation culture solution of experimental animals treated with a therapeutic agent according to the present invention.

Claims (4)

Atopic dermatitis treatment composition containing illite as an active ingredient. The method of claim 1, A composition for treating atopic dermatitis, further comprising sewage extract as an active ingredient. The method according to claim 1 or 2, A composition for treating atopic dermatitis, further comprising a leaf extract as an active ingredient. The method of claim 2 or 3, The extract is a composition for treating atopic dermatitis, characterized in that obtained by cold water extraction, hot water extraction or solvent extraction.