KR20120027432A - Health food or pharmaceutical composition comprising chestnut shell extract - Google Patents

Abstract

PURPOSE: Health food or pharmaceutical composition including containing a chestnut skin extract is provided to reduce the pruritus of a user. CONSTITUTION: Health food or pharmaceutical composition contains 0.005-80wt% of chestnut skin extract for reducing or controlling the pruritus of a user. The chestnut skin is selected from the inner skin, the outer skin, or their mixture. The chestnut skin extract is obtained by extracting the chestnut skin using water, low class alcohol having 1-4 carbons, 1,3-butyl glycol, or their mixed solution.

Description

Health food or pharmaceutical composition comprising chestnut extract {HEALTH FOOD OR PHARMACEUTICAL COMPOSITION COMPRISING CHESTNUT SHELL EXTRACT}

The present invention relates to a health food or pharmaceutical composition comprising a chestnut extract.

Itching is defined as an unpleasant skin sensation that causes the desire to scratch, and is a physiological self-defense mechanism such as pain, tactile, cold or hot, that can be perceived when the skin is exposed to harmful stimuli from the outside. It protects you.

Itching is a common symptom common in various skin and systemic diseases.Acute itching due to urticaria or other side effects can be easily cured, but severe chronic itching, such as biliary tract obstruction, kidney disease, or atopic disease Treatment is very difficult.

Itching is caused by a variety of causes, including inflammation, cancer, metabolic diseases, infections, psychiatric illnesses, medications, or stress, and recent studies have shown that organic connections between the skin, peripheral nervous system, and central nervous system cause itching. And deeply involved in response and control.

Recently, it has been found that certain sensory neurons and their receptors specifically respond to itching, and itching is no longer a subtype of pain, but an individual sense of the sensory nervous system, and different itching mediators and their receptors It is believed to be involved in various itching-induced diseases (Steinhoff et al., Journal of Investigative Dermatology, 126, pp 1705-1718, 2006).

Histamines have been used mainly for the study of itching so far, but it has been argued that chronic itching, such as atopy, is due to neurological causes rather than histamine-dependent pathways, which is why antihistamine is atopic disease. It is explained whether or not it is effective in the itch (Stander et al., Experimental Dermatology, 11, pp 12-24, 2002).

In addition, first-generation antihistamines are mainly used for systemic administration, and have anti-sympathetic effects, indicating sedation. First-generation antihistamines, chlorpheniramine, are not known to suppress the itch of atopic dermatitis patients when administered topically (Munday et. al., Dermatology, 205, pp 40-45, 2002), Topical antihistamines are not recommended for atopic dermatitis because of the risk of skin hypersensitivity. Second-generation antihistamines, eastin and terpenadine, which are non-sedative, can cause arrhythmia when taken with drugs that inhibit cytochrome P450 activity (ketoconazole, erythromycin) (Hey et al., Arzneimittelforschung, 46, pp159 -163, 1996), with associated side effects.

Therefore, there is an urgent need for the development of effective and safe side effects for the treatment of itching, in particular for the treatment of chronic itching such as atopy.

Therefore, an object of the present invention is to solve the technical problem that has been requested from the past. Specifically, an object according to an embodiment of the present invention is to provide a health food or pharmaceutical composition for reducing or suppressing itching, including a chestnut extract, a health food or pharmaceutical composition for improving skin barrier, and a health food or pharmaceutical composition for immunosuppression will be.

In accordance with this object, one embodiment according to the present invention, while reducing the side effects of the conventional itch treatments, itching or mitigating health foods or pharmaceuticals containing chestnut extract as an active ingredient having an excellent itch inhibition or alleviation effect It relates to a composition.

One embodiment according to the present invention relates to a health food or pharmaceutical composition for improving skin barrier function comprising chestnut extract as an active ingredient.

One embodiment according to the present invention relates to a health food or pharmaceutical composition for immunosuppression comprising a chestnut extract as an active ingredient.

One embodiment according to the present invention relates to a health food or pharmaceutical composition for improving or treating atopic dermatitis, comprising a chestnut extract as an active ingredient.

The composition according to the present invention includes chestnut extract as an active ingredient, thereby reducing the excellent itch by inhibiting the activity of Proteinase-Activated Receptor-2 (PAR-2), which is a source of stimulation of itching Not only can significantly suppress skin barrier breakdown caused by itching, but can also fundamentally treat immune hypersensitivity reactions that can cause itching through the immunosuppressive activity of chestnut extract have.

1 is a graph showing the measurement results of the inhibitory effect of PAR-2 activity (trypsin treatment) of chestnut extract according to an embodiment of the present invention;
Figure 2 is a graph showing the measurement results of the inhibitory effect of PAR-2 activity (SLIGKV treatment) of the chestnut extract according to an embodiment of the present invention;
Figure 3 is a graph showing the results of measuring the inhibitory effect of PAR-2 activity (trypsin, SLIGKV treatment) of chestnut 1,3-butylglycol (BG) extract according to an embodiment of the present invention;
Figure 4 is a graph showing the itch (trypsin treatment) inhibitory effect of the chestnut 1,3-butyl glycol (BG) extract according to an embodiment of the present invention;
5 is a graph showing the itch (SLIGRL treatment) inhibitory effect of the chestnut 1,3-butyl glycol (BG) extract according to an embodiment of the present invention;
Figure 6 is a graph showing the effect of inhibiting itching (SLIGRL treatment) of chestnut ethanol extract according to an embodiment of the present invention;
7 is a graph showing the itch (SLIGRL treatment) inhibitory effect of oral administration of chestnut ethanol extract according to an embodiment of the present invention;
8 is a graph showing the effect of reducing the TNF-α secretion of chestnut extract according to an embodiment of the present invention;
9 is a graph showing the IL-6 secretion reduction effect of chestnut extract according to an embodiment of the present invention;
10 is a graph showing the effect of reducing IL-1α secretion of chestnut extract according to an embodiment of the present invention;
Figure 11 is a graph showing the IL-8 secretion reduction effect of the chestnut extract according to an embodiment of the present invention;
12 is a graph showing the GM-CSF secretion reduction effect of the chestnut extract according to an embodiment of the present invention;
Figure 13 is a graph showing the effect of IL-6 secretion by trypsin and active peptide (SLIGKV) of chestnut extract according to an embodiment of the present invention;
Figure 14 is a graph showing the effect of reducing IL-8 secretion by trypsin and active peptide (SLIGKV) of chestnut extract according to an embodiment of the present invention;
Figure 15 is a graph showing the effect of reducing GM-CSF secretion by trypsin and active peptide (SLIGKV) of chestnut extract according to an embodiment of the present invention;
Figure 16 is a graph showing the results of measuring the skin moisturizing effect of the chestnut ethanol extract according to an embodiment of the present invention;
17 is a graph showing the results of measuring the hyperkeratosis improving effect of the chestnut ethanol extract according to an embodiment of the present invention;
18 is a graph showing the results of measuring the itch improvement effect of chestnut ethanol extract in the NC / Nga model according to an embodiment of the present invention;
19 is a graph showing the results of measuring the IgE reduction effect of the chestnut ethanol extract in the NC / Nga model according to an embodiment of the present invention;
Figure 20 is a graph showing the measurement results of the inhibitory effect of PAR-2 activity according to the endothelial (cuticle) and shell (shell) extract of the chestnut in accordance with an embodiment of the present invention.

The present invention relates to a health food or pharmaceutical composition for itching or suppressing itching comprising chestnut extract as an active ingredient. The inventors of the present application, using the proteinase-activated receptor-2 (PAR-2) as a target for the treatment of itching, measured the degree of inhibition of PAR-2 activity by chestnut extract, As demonstrated in the examples, it showed excellent PAR-2 antagonism in vitro, and was also induced by trypsin, SLIGRL, SLIGKV or protease, a peptide that specifically activates PAR-2. In the itch inhibition experiment, it was confirmed that the activity of PAR-2 significantly inhibited to exhibit a very good itch inhibition effect.

The itch is also called pruritus, the cause or form of the itch is not particularly limited, for example, inflammatory dermatitis, atopic dermatitis, dermatitis due to roughness of skin, sweat bands, erosion, frostbite, contact dermatitis, seborrheic dermatitis And dermatitis consisting of psoriasis and psoriasis.

In the case of itching, itching constantly causes the skin to be rubbed, scratched, or pinched. This causes secondary skin damage, including skin erosion, abrasions, scabs, sunburn, hyperpigmentation, or reduction of pigmentation, and the release of various substances that induce inflammatory responses in the skin, which secrete itches again. This increases the circulation of itching-scratching.

However, the chestnut extract contained in the health food or pharmaceutical composition according to the present invention shows an effective skin barrier function improving effect, it can effectively prevent or treat secondary skin damage caused by itching.

The composition has a significant effect on alleviating the skin barrier damage or improving the resilience of the skin barrier to secondary skin damage caused by, for example, itching derived from atopic dermatitis, thereby significantly reducing the skin barrier. Can be improved.

The composition can improve the skin barrier, in particular, by enhancing skin moisturization or preventing skin keratinization, and the inventors of the present application apply oxazolon to hairless mice as demonstrated in the following examples. Experiments were conducted in the treated allergy model, and transepidermal water loss (TEWL) and skin thickness were measured, and chestnut extract showed effective skin moisturization or anti-keratinization effect. It was.

Moreover, the present invention relates to a health food or pharmaceutical composition for immunosuppression comprising chestnut extract as an active ingredient. The composition may be, for example, a health food or pharmaceutical composition for the treatment of atopy, rheumatoid arthritis or Crohn's disease, and may be a composition for the prevention or treatment of atopic disease, which is an immune hypersensitivity reaction. have.

As atopic disease develops from the acute stage to the chronic disease stage due to skin damage, various secretion patterns of immune mediators are observed. The concentration of interleukin in the damaged skin of patients with atopic disease Will increase. In addition, granulocyte macrophage colony stimulating factor (GM-CSF) is known to cause a chronic inflammatory response due to increased secretion in the damaged skin of patients with atopic skin diseases (Matsubara et al.). al., FEBS Letters, 566, pp 195-200, 2004).

In this regard, the inventors of the present application have found that chestnut extract is associated with tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1α (IL- 1α), interleukin-8 (IL-8), or granulocyte-macrophage colony stimulating factor (GM-CSF) were found to be able to significantly reduce the expression, chestnut extract was immunosuppressed Suitable for the active ingredient for health food or pharmaceutical composition.

In particular, the present invention is suitable for the active ingredient of the cosmetic composition for improving or treating atopic dermatitis by including chestnut extract as an active ingredient.

In addition, the interleukin-6 (IL-6) and interleukin-8 (IL-8) are also known as factors causing itching in atopic diseases, chestnut extract is effectively used for the prevention and treatment of itching from atopic diseases. Can be.

The chestnut means a dark brown bark covering the fruit of chestnut, and the chestnut extract used herein refers to one or more extracts selected from the group consisting of chestnut endothelium (hulled skin), skin and mixtures thereof. It may be.

The chestnut is irrelevant to any kind of shell derived from chestnuts, and is not particularly limited, for example, chestnut (Castanea crenata S. et Z., Castanea mollissima Bl., Castanea bulgaris), chestnut (Castanea Bungeana Bl) and It may be one or more chestnut hulls selected from the group consisting of Castanea crenata for multicarpa (Uyeki) Chung. Chestnut hulls can be used as chestnuts, and chestnut hulls (chest husks) can be used, but PAR-2 antagonists are superior in the group treated with chestnut hulls (chest husks) extract. It showed the action.

The extraction method of the chestnut extract is not particularly limited, but may be extracted through at least one solvent selected from the group consisting of water, lower alcohols having 1 to 4 carbon atoms, 1, 3-butylglycol and mixed solvents thereof. For example, the solvent may be at least one selected from the group consisting of water, methanol, ethanol, 1, 3-butylglycol, butanol, and mixtures thereof. For example, the chestnut extract may be 10-100% aqueous alcohol solution. Or it can be extracted from 10 to 100% 1, 3-butylglycol, specifically may be a chestnut 20-90% ethanol extract or 10-70% 1,3-butylglycol extract, more specifically chestnut 40 It may be a ~ 90% aqueous ethanol extract or 10-50% 1,3-butylglycol extract, more specifically the chestnut 60-90% ethanol aqueous extract or 20-40% 1,3-butylglycol extract There.

The content of the chestnut extract included in the composition is not particularly limited, but may be included in an amount of 0.005 to 80% by weight based on the total weight of the composition, and may be preferably included in an amount of 0.01 to 30% by weight. If the content of the chestnut extract is too small, the effect may be insignificant, and if too much, the stability of the formulation may be lowered.

In some cases, it is also possible to combine chestnut extract with one or more of antihistamine, steroids, local anesthesia, immunosuppressants for alleviating or suppressing itching, improving skin barrier and suppressing immunity.

Conventional techniques for relieving atopic dermatitis and itching have been to take antihistamines or apply steroids to external preparations, but these preparations have the disadvantage that they will recur soon, even if they show temporary therapeutic efficacy. In this case, side effects such as central nervous system disorders and digestive disorders have been reported. Corticosteroids, which are corticosteroids, have excellent anti-inflammatory and immunosuppressive effects, but have serious side effects, and the immunosuppressive drug tacrolimus hydrate ointment has been reported to be effective in treating atopic dermatitis. When absorbed into the body in a large amount to the damage site of the skin there is a risk of causing kidney damage is difficult to use in the long term in terms of safety.

Thus, when the composition containing chestnut extract as an active ingredient and one or more of antihistamines, steroids, local anesthesia, and immunosuppressants are appropriately used, it is not accompanied by side effects such as alleviation or suppression of itching, improvement of skin barrier and immune suppression. It can safely produce a big effect.

The pharmaceutical composition may further contain pharmaceutical adjuvants such as preservatives, stabilizers, hydrating or emulsifying accelerators, salts and / or buffers for the control of osmotic pressure, and other therapeutically useful substances, and are commonly used inorganic or organic carriers. In addition to oral administration in the form of a solid, semi-solid or liquid, or parenteral, rectal, topical, transdermal, intravenous, intramuscular, intraperitoneal, subcutaneous, etc., in particular oral administration is preferred.

The oral dosage forms include, for example, tablets, pills, hard and soft capsules, liquids, suspensions, emulsifiers, syrups, and granules, and these formulations may contain diluents (eg, lactose, dextrose, water, etc.) in addition to the active ingredients. Cross, mannitol, sorbitol, cellulose and glycine), lubricants such as silica, talc, stearic acid and its magnesium or calcium salts and polyethylene glycols. Tablets may also contain binders such as magnesium aluminum silicate, starch paste, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose and polyvinylpyrrolidine, optionally starch, agar, alginic acid or its sodium salt Pharmaceutical additives such as disintegrants, absorbents, colorants, flavors, and sweeteners. The tablets can be prepared by conventional mixing, granulating or coating methods.

In addition, the parenteral administration agent may be, for example, an external preparation for skin, and may be a lotion, ointment, gel, cream, patch or spray formulation, but is not limited thereto.

The health food composition is not particularly limited in the formulation, for example, it may be formulated as a tablet, granules, drinks, caramel, diet bar and the like. In addition to the active ingredient, the health food composition of each formulation may be suitably selected by a person skilled in the art according to the formulation or purpose of use, in addition to the active ingredient, and may be synergistic when applied simultaneously with other raw materials. .

Determination of the dosage of the active ingredient is within the level of those skilled in the art, and the daily dosage of the drug depends on various factors such as less progression, onset, age, health condition, complications, etc. of the subject to be administered. Generally, the composition may be administered by dividing the composition 1 to 500 mg / kg, preferably 30 to 200 mg / kg, once or twice a day, and the dosage may be in the range of the present invention by any method. It is not limiting.

Hereinafter, the present invention will be described in more detail with reference to Examples, but the following Examples are provided to illustrate the present invention, and the scope of the present invention is not limited thereto.

[ Example  One] Chestnut  Preparation of Extract

1-1) Chestnut  1,3- Butyl glycol  Preparation of Extract

Chestnut was leached at room temperature for 3 days using 30% 1,3-butylglycol (1,3-buthylene glycol). Subsequently, the filter was sequentially filtered through a filter having a size of 250 mesh, 3 μm, 1 μm, and 0.5 μm. Then, after standing for 3 days at 0 ~ 4 ℃ (Standing), and then filtered sequentially with a filter of 0.5㎛, 0.3㎛, 0.2㎛ size to obtain a chestnut 1,3-butyl glycol extract.

1-2) Chestnut  Preparation of Ethanol Extract

Chestnut was leached for 3 days at room temperature using 70% ethanol. Subsequently, the filter was sequentially filtered through a filter having a size of 250 mesh, 3 μm, 1 μm, 0.5 μm, 0.3 μm, and 0.2 μm. Thereafter, the solution was concentrated at 60 ° C. and then vacuum dried at 30 ° C. for 18 hours to obtain chestnut ethanol extract in powder form.

[ Test Example  One] Chestnut  Of ethanol extract PAR -2 activity inhibitory effect ( in in vitro , Trypsin  process, HEK : Human Epidermal Keratinocyte )

One day before the experiment, keratinocytes (cell name: HaCaT obtained from ATCC) were dispensed to ⁴ × 10 ⁴ cells / well in 96 well plates and incubated for 24 hours in a 37 ° C., 5% CO 2 incubator. . After 24 hours, two 96-well plates were washed with Hanks'Balanced Salt solution (HBSS) buffer, followed by reaction buffer (2 uM Fluo-4-AM, 20% pluronic acid, 2.5 mM probenecid). Put it in. After reacting at 37 ° C., 5% CO ₂ incubator for 30 minutes and at room temperature for 30 minutes, washing twice with HBSS buffer and the chestnut ethanol extract was applied to cells 1ppm, 2ppm, 5ppm, 10ppm, 20ppm, respectively. Were treated at concentrations of 30 ppm and 50 ppm. After reacting for 10 minutes, treated with 2U / ml trypsin (Trypsin) and was measured in Ca ^{2 +} concentration in the cell 80 seconds. In Ca ^{2 +} concentration in the measurement cell presentation flex 3: it was used (FlexStation3 Molecular Device, USA). After the chestnut ethanol extract and trypsin treatment, find the difference between the minimum and maximum values obtained by measuring flex for 80 seconds, and then determine the difference between the minimum and maximum values of Trypsin treatment. The inhibition rate was calculated | required compared with the difference.

Referring to FIG. 1, when PAR-2 is activated by cutting a serine sequence at the N-terminus of PAR-2 by trypsin, calcium ions are introduced into cells, and in the case of the group treated with chestnut extract, PAR It can be seen that the inactivation of -2 is significantly reduced due to the inhibition of -2 activation.

[ Test Example  2] Chestnut  Of ethanol extract PAR -2 activity inhibitory effect ( in in vitro , SLIGKV  process, HEK : Human Epidermal Keratinocyte )

One day prior to the experiment, keratinocytes (cell name: HaCaT obtained from ATCC) were dispensed into ⁴ × 10 ⁴ cells / well in 96 well plates, and then cultured in a 37 ° C., 5% CO ₂ incubator for 24 hours. It was. After 24 hours, two 96-well plates were washed with Hanks' Balanced Salt solution (HBSS) buffer, followed by reaction buffer: 2 uM Fluo-4-AM, 20% pluronic acid, 2.5 mM probenecid ) Into the cells. After reacting at 37 ° C., 5% CO ₂ incubator for 30 minutes and at room temperature for 30 minutes, washing twice with HBSS buffer and the chestnut ethanol extract was applied to cells 1ppm, 2ppm, 5ppm, 10ppm, 20ppm, respectively. Were treated at concentrations of 30 ppm and 50 ppm. After 10 minutes of reaction, the treatment 5uM PAR-AP (SLIGKV) and was measured in Ca ^{2 +} concentration in the cell 80 seconds. In Ca ^{2 +} concentration in the measurement cell presentation flex 3: it was used (FlexStation3 Molecular Device, USA). After the chestnut ethanol extract and 5uM PAR-AP (SLIGKV) treatment, the difference between the minimum value and the maximum value obtained by measuring flex for 80 seconds was calculated, and the value was determined during 5uM PAR-AP (SLIGKV) treatment. The inhibition rate was obtained by comparing the difference between the minimum and maximum values.

Referring to FIG. 2, when SLIGKV (Human), an activating peptide, acts as a direct ligand and PAR-2 is activated, calcium ions are introduced into cells. It can be seen that the influx of calcium ions is significantly reduced.

[ Test Example  3] Chestnut  1,3- Butyl glycol ( BG A) of extract PAR -2 activity inhibitory effect ( in  in vitro, SLIGKV  process, HEK : Human Epidermal Keratinocyte )

One day prior to the experiment, keratinocytes (cell name: HaCaT obtained from ATCC) were dispensed into ⁴ × 10 ⁴ cells / well in 96 well plates, and then cultured in a 37 ° C., 5% CO ₂ incubator for 24 hours. It was. After 24 hours, two 96-well plates were washed with Hanks' Balanced Salt solution (HBSS) buffer, followed by reaction buffer: 2 uM Fluo-4-AM, 20% pluronic acid, 2.5 mM probenecid ) Into the cells. After reacting at 37 ° C., 5% CO ₂ incubator for 30 minutes and at room temperature for 30 minutes, washing twice with HBSS buffer and extracting chestnut 1,3-butylglycol (BG) into the cells, respectively 0.1 Treatment was carried out at concentrations of w / v%, 0.5w / v% and 1w / v%. After reacting for 10 minutes, treated with 2U / ml trypsin (Trypsin) or 5uM PAR-AP (SLIGKV) and was measured in Ca ^{2 +} concentration in the cell 80 seconds. In Ca ^{2 +} concentration in the measurement cell presentation flex 3: it was used (FlexStation3 Molecular Device, USA). After the chestnut 1,3-butylglycol (BG) extract and 2U / ml Trypsin or 5uM PAR-AP (SLIGKV) treatment, the difference between the minimum and maximum values obtained by measuring the flex for 80 seconds was measured. After the determination, the value was compared with the difference between the minimum value and the maximum value at 2 U / ml Trypsin or 5 uM PAR-AP (SLIGKV) treatment to determine the inhibition rate.

Referring to Figure 3, it can be seen that the influx of intracellular calcium ions by trypsin or PAR-2 active peptide (SLIGKV) is significantly reduced by increasing the concentration of chestnut extract.

[ Test Example  4] Chestnut  1,3- Butyl glycol ( BG Inhibitory effect of itching induced by trypsin

Scratching behavior of hairless mice in 200 Pg / site injection of trypsin (in PBS buffer) was determined by extracting chestnut 1,3-butylglycol (BG) extract (in PBS buffer). It was observed to be suppressed for each concentration by simultaneous injection at concentrations of 1w / v%, 10w / v% and 20w / v%, respectively. Each measurement was expressed as% change relative to the average value of two Baseline measurements, and the results are shown in FIG. 4.

Referring to FIG. 4, the 1,3-butylglycol (BG) treatment group and the 1% chestnut 1,3-butylglycol (BG) extract treatment group have no effect of suppressing the itch caused by trypsin. Although observed, 10%, 20% chestnut 1,3-butylglycol (BG) extract treatment group was observed to have a very large inhibitory effect.

[ Test Example  5] Chestnut  1,3- Butyl glycol ( BG A) of extract SLIGRL Itching Inhibition Effect Induced by

Chestnut 1,3-butylglycol (BG) extract with scratching behavior following injection of 50 µg / site intradermal (ID) injection of Murine PAR-2 active peptide in PBS buffer (SLIGRL) in hairless mice (in PBS buffer) was injected simultaneously at concentrations of 1%, 10% and 20%, respectively, and observed to be inhibited by concentration. Each measurement was expressed as% change relative to the average of two Baseline measurements, and the results are shown in FIG. 5.

Referring to FIG. 5, the concentration of chestnut extract increased from 1% chestnut 1,3-butylglycol (BG) extract treatment group to 10% and 20% chestnut 1,3-butylglycol (BG) extract treatment group. As a result, it was observed that the degree of suppression of the itch caused by SLIGRL was increased.

[ Test Example  6] Chestnut  Ethanol Extracts According to Skin Application SLIGRL Itching Inhibition Effect Induced by

The skin-coated chestnut extract in each test group was dissolved in a solution of water, ethanol, and 1,3 butyl glycol in a ratio of 5: 3: 2, and the test concentration was a powdered chestnut ethanol extract. Used at concentrations of 1, 3 and 10 w / v%. The vehicle group used a preparation made without adding the extract of Example 1, and as an itch-inducing substance, PAR-2 Activating Peptide (SLIGRL; rodent-derived peptide) was dissolved in PBS buffer and 50 ㎍ / Intradermal injection at site concentration.

200 μl of each sample of chestnut ethanol extract was treated by spreading it on the back of hairless mice. The treatment was performed twice daily for four days, divided into morning and afternoon, and last treated 30 minutes prior to the morning injection of itching on the fifth day.

Number of scrapes for 40 minutes per measurement for 2 days prior to skin application of chestnut ethanol extract to compare itching with itch-inducing substance (PAR-2 active peptide; SLIGRL) and reduction of itching by chestnut ethanol extract The value calculated by calculating the average value over two days was compared with the number of scratches for 40 minutes after the stimulus source injection, and expressed as a% change value.

The itch experiment began with a 20-minute adaptation of a hairless mouse into a transparent cage for observation 20 minutes before the itch-injecting material was injected. At the end of the acclimatization time, an itch-inducing substance (PAR-2 active peptide; SLIGRL) was intradermal injected into the dorsal skin of the adapted hairless mice. Immediately after the injection, the hairless mice were placed in the cage again for 40 minutes, and the number of times of scratching the itch-injected site was measured with the hind legs. Only scratching of the injection site with the hind paw was considered as an indication of itching behavior, except for the forefoot or biting by mouth.

Referring to FIG. 6, after applying chestnut ethanol extract (Chestnut Inner Bark Extract) to the skin, the number of scratches was reduced even in a normal state without itch irritation at a concentration of 10%, and when itching was irritated 1% It can be seen that the number of scratches decreases from the concentration.

[ Test Example  7] Chestnut  Ethanol Extracts According to Oral Administration SLIGRL Itching Inhibition Effect Induced by

The extract obtained in Example 1 was orally administered to 8 week old female hairless mice to observe the effect of inhibiting itching.

All the chestnut extracts treated orally in each test group were used as a suspension by mixing with distilled water, and the test concentration was orally administered with powder and chestnut ethanol extract at the concentrations of 200 and 500 mg / Kg, respectively. In the vehicle group, pure distilled water was used without adding the extract of Example 1, and as an itch-inducing substance, PAR-2 Activating Peptide (SLIGRL; rodent-derived peptide) was dissolved in PBS buffer and 50 ㎍ / Intradermal injection at site concentration.

For oral administration, the appropriate samples were taken and administered once a day for 5 days, and finally administered 30 minutes before the morning of the itch-inducing substance on the 5th day.

To compare the increase in the itch caused by the itch-inducing substance (PAR-2 active peptide; SLIGRL) and the reduction in the itch by oral administration of chestnut ethanol extract, 40 minutes per measurement for 2 days prior to oral administration of chestnut ethanol extract The number of scratches was measured, and the value calculated for the average value over two days was compared with the number of scratches for 40 minutes after injecting the stimulus source and expressed as a% change value.

The itch experiment began with a 20-minute adaptation of a hairless mouse into a transparent cage for observation 20 minutes before the itch-injecting material was injected. At the end of the adaptation time, an itch-inducing substance (PAR-2 active peptide; SLIGRL) was intradermal injected into the dorsal skin of the adapted hairless mice. Immediately after the injection, the hairless mice were placed in the cage again for 40 minutes, and the number of times of scratching the itch-injected site was measured with the hind legs. Only scratching of the injection site with the hind paw was considered as an indication of itching behavior, except for the forefoot or biting by mouth.

Referring to Figure 7, after oral administration of the chestnut ethanol extract, it can be seen that the inhibitory effect of the chestnut on the itching stimulation decreases with concentration.

[ Test Example  8] Chestnut  Extract TNF -α secretion reduction

One day before the experiment, normal human skin keratinocytes (NHEK, obtained from Lonza) were dispensed in 96 well plates at 5 × 10 ⁴ cells / well, and then in a 37 ° C., 5% CO ₂ incubator. Incubated for 24 hours. After 24 hours, the cells were washed twice with PBS and replaced with serum free KBM (keratinocyte basement media). Chestnuts were treated in each well by concentration (10, 25, 50 ppm) and reacted for 30 minutes, followed by PGSA (10, 50 ppm) and LPS (1 ppm), respectively. After incubation for 24 hours at 37 ℃, 5% CO ₂ incubator (incubator), the culture was taken and subjected to ELISA for TNF-α. ELISA used the experimental method of the manufacturer (BD science).

Referring to Figure 8, it can be observed that chestnut significantly reduces the secretion of TNF-α increased by PGSA and LPS.

[ Test Example  9] Chestnut  Extract IL -6 secretion reduction

Substantially the same method as in Test Example 8 was used except that ELISA was performed on IL-6.

Referring to Figure 9, it can be observed that chestnut significantly inhibits the secretion of IL-6 increased by PGSA and LPS.

[ Test Example  10] Chestnut  Extract IL -1α secretion decreased

Substantially the same method as in Test Example 8 was used except that ELISA for IL-1α was performed.

Referring to FIG. 10, it can be observed that chestnut reduces the amount of IL-1α secreted by PGSA and LPS according to the concentration.

[ Test Example  11] Chestnut  Extract IL -8 secretion reduction

Substantially the same method as in Test Example 8 was used except that ELISA was performed on IL-8.

Referring to Figure 11, it can be seen that chestnut significantly reduces the secretion of IL-8 increased by PGSA and LPS.

[ Test Example  12] Chestnut  Extract GM - CSF  Decreased secretion

Except that the ELISA for GM-CSF was carried out substantially the same method as in Test Example 8.

Referring to FIG. 12, it can be observed that as the concentration of chestnut increases, the amount of GM-CSF secreted by PGSA and LPS decreases.

[ Test Example  13] Chestnut  Trypsin and Activity of Extracts Peptide (SLIGKV)  by IL -6 secretion inhibitory effect

One day before the experiment, normal human skin keratinocytes (NHEK, obtained from Lonza) were dispensed in 96 well plates at 5 × 10 ⁴ cells / well, and then in a 37 ° C., 5% CO ₂ incubator. Incubate for 24 hours. After 24 hours, the cells were washed twice with PBS and replaced with serum free KBM (keratinocyte basement media). Each well was treated with chestnut extract by concentration (10, 50ppm) and reacted for 30 minutes, followed by trypsin (10nM) or PAR-2 activating peptide (SLIGKV, 50 uM), respectively. After incubation for 24 hours at 37 ℃, 5% CO ₂ incubator (incubator), the culture was taken and subjected to ELISA for IL-6. ELISA used the experimental method of the manufacturer (BD science).

Referring to Figure 13, it can be observed that chestnut extract inhibits the secretion of IL-6 by trypsin and active peptide (SLIGKV) in a concentration-dependent manner.

[ Test Example  14] Chestnut  Trypsin and Activity of Extracts Peptide (SLIGKV)  by IL -8 secretion inhibitory effect

Substantially the same method as in Test Example 13 was used except that ELISA was performed on IL-8.

Referring to Figure 14, it can be observed that chestnut extract inhibits the secretion of IL-8 by trypsin and active peptide (SLIGKV) in a concentration-dependent manner.

[ Test Example  15] Chestnut  Trypsin and Activity of Extracts Peptide (SLIGKV)  by GM -CSF secretion inhibitory effect

Substantially the same method as Test Example 13 was used except that ELISA on GM-CSF was performed.

Referring to Figure 15, it can be observed that chestnut extract inhibits the secretion of GM-CSF by trypsin and active peptide (SLIGKV) concentration-dependently.

[ Test Example  16] Chestnut  Skin Moisturizing Effect of Ethanol Extracts

The recovery ability of skin barrier function due to long-term skin damage of chestnut ethanol extract of Example 1 was evaluated. First, oxazolone was periodically applied to the back of young hairless mice (Orient, Korea) 7 to 8 weeks after birth for 6 days to damage the skin barrier of experimental animals. Subsequently, transepidermal water loss (TEWL) was measured using an evaporator (Delpin, Finland) to group experimental animals with skin that exhibited transdermal moisture loss of 50 g / m ² or more per hour. Each bark extract was applied twice a day for 10 days at a dose of 100 μl per 5 cm ² of skin area, and the transdermal moisture loss (TEWL) of the test animals was measured, and the results are shown in FIG. 16.

As shown in FIG. 16, the increased transdermal moisture loss due to atopic dermatitis induced by oxazolone in the allergy model treated with oxazolone in hairless mice is a barrier damaged by applying chestnut extract. It was found that there is a function to restore the function to normal skin.

[ Test Example  17] Chestnut  Of ethanol extract Hyperkeratosis  Improvement effect measurement

In Example 1, the skin keratinization improvement effect of the chestnut ethanol extract was evaluated. First, oxazolone was periodically applied to the back of young hairless mice (Orient, Korea) 7 to 8 weeks after birth for 6 days to damage the skin barrier of experimental animals. Then, jeungbalgye (Delphin, Finland) by transepidermal water loss (Transepidermal Water Loss: TEWL) measurements by the test substance, to separate the group of animals with skin showing a 50g / m ² or more percutaneous water loss per skin area of 5cm ² each After the continuous application for 10 days twice a day at 100 μl volume, the skin thickness was measured using a two-folding measurement micrometer (Mitsubishi, Japan). Shown in

Referring to FIG. 17, it can be seen that the increased skin thickness due to dermatitis induced by oxazolon has a skin hyperkeratosis inhibitory effect by using chestnut extract powder.

[ Test Example  18] Chestnut  Of ethanol extract NC Of Nga In the model IgE Measures to reduce and improve itching

In Example 1, the effect of reducing IgE and itching in serum of chestnut ethanol extract was evaluated by NC / Nga mouse model. First, the back part of the young NC / Nga mice 7 to 8 weeks after birth was depilated using a clipper, and the remaining hair was completely removed using a depilatory cream. Df (dermatophagoides farinae) 100mg was uniformly applied twice a week for three weeks and four times on the exposed dorsal and auricle areas, causing atopic disease. After three times a week, the chestnut extract was divided into two groups, 100 mg / kg and 250 mg / kg, and NC / Nga mice were placed in the observation cage for itch measurement. The number of times was measured. Except for scratching with forefoot or biting by mouth, only scratching with hind foot was considered as an indication of itching. Serum IgE was measured by separating serum from blood samples collected through the orbital vein and carrying out ELISA using an Opt IgE kit from BD Science. It was shown in Fig. 18 that the itch was reduced in the 250 mg / kg administration group compared to the untreated group. Referring to FIG. 19, it can be seen that the decrease in IgE is significantly shown in the 250 mg / kg administration group.

[ Test Example  19] of Chestnut Crust Ethanol Extract and Chestnut Crust Ethanol Extract PAR -2 activity inhibitory effect comparison in in vitro , SLIGKV  process, HEK : Human Epidermal Keratinocyte )

Chestnut husks and cuticles were separated and dried, chestnut husk extract was prepared in the same manner as in Example 1-2), chestnut husk extract was also prepared in the same manner as in Example 1-2). Using the same method as in Test Example 2, the effect of inhibiting the PAR-2 activity of the chestnut husk ethanol extract and chestnut husk ethanol extract was measured and shown in Table 2 and FIG.

TABLE 2

[ Test Example  20] of Chestnut Crust Ethanol Extract and Chestnut Crust Ethanol Extract PAR -2 activity inhibitory effect comparison in in vitro , SLIGKV  process, HEK : Human Epidermal Keratinocyte )

The effect of inhibiting PAR-2 activity was measured in the same manner as in Test Example 19, except that the concentrations of the husks and skins of the chestnut extract were treated as shown in Table 3 below.

[Table 3]

Although a formulation example of the composition according to the present invention is described below, the pharmaceutical composition or health food composition is applicable to various formulations, which are intended to explain in detail only, not intended to limit the present invention.

[Formulation Example 1] Ointment in the external preparation for skin

The ointment was prepared in a conventional manner according to the composition described in Table 1 below.

[Table 1]

Formulation Example 2 Preparation of Powder

Example 1 ............... 100 mg

Lactose ... .......... 100 mg

Talc .................. .......... 10 mg

maintain................................................. ........... 5 mg

The above ingredients are mixed and filled in an airtight cloth to prepare a powder.

Formulation Example 3 Preparation of Tablet

Example 1 ........................ 50 mg

Corn Starch .100 mg

Lactose ... ...... 100 mg

Magnesium Stearate ......................................... 2 mg

Vitamin C ... .... 50 mg

After mixing the above components, tablets are prepared by tableting according to a conventional method for preparing tablets.

Formulation Example 4 Preparation of Capsule

Example 1 ............... 50 mg

Corn Starch ... mg

Lactose ... ..... 100 mg

Magnesium Stearate ..................... 2 mg

Vitamin C ... ... 50 mg

Serine ... ...... 50 mg

According to a conventional capsule preparation method, the above ingredients are mixed and filled into gelatin capsules to prepare capsules.

Formulation Example 5 Preparation of Liquid

Example 1 ............. 100 mg

Isomerized sugar ... ... 10 g

Mannitol ... .... 5 g

Vitamin C ... ... 50 mg

Serine ... ....... 50 mg

maintain................................................. ....... suitable

Purified water................................................. ...... suitable

According to the conventional method for preparing a liquid, each component is added and dissolved in purified water, lemon flavor is added, the above components are mixed, purified water is added, the whole is adjusted to 100 ml by adding purified water, and then filled in a brown bottle. The solution is prepared by sterilization.

Formulation Example 6 Preparation of Health Food

Example 1 ................... 1000 mg

Vitamin mixtures

Vitamin A Acetate ............... 70 μg

Vitamin E ......................................... 1.0 mg

Vitamin B1 .................................. 0.13 mg

Vitamin B2 ................................... 0.15 mg

Vitamin B6 ......................................... 0.5 mg

Vitamin B12 ......................... 0.2 μg

Vitamin C ......................................... 10 mg

Biotin ......................................... 10 μg

Nicotinic Acid Amide ... 1.7 mg

Folic acid ......................................... 50 ㎍

Calcium Pantothenate ......................................... 0.5 mg

Mineral mixture

Ferrous Sulfate ............... 1.75 mg

Zinc Oxide ............... 0.82 mg

Magnesium Carbonate ... 25.3 mg

Potassium monophosphate ......................................... 15 mg

Dibasic Calcium Phosphate ......................................... 55 mg

Potassium Citrate ... 90 mg

Calcium Carbonate ... 100 mg

Magnesium Chloride ......................................... 24.8 mg

Although the composition ratio of the above-mentioned vitamin and mineral mixture is comparatively mixed with a composition suitable for health food as a preferred embodiment, the compounding ratio may be arbitrarily modified, and the above ingredients are mixed according to a conventional method for producing healthy foods , Granules can be prepared and used in the manufacture of health food compositions according to conventional methods.

Formulation Example 7 Preparation of Healthy Drink

Example 1 .................. 1000 mg

Citric Acid ... ......... 1000 mg

oligosaccharide................................................. ........ 100 g

Plum concentrate ..................... ......... 2 g

Taurine ... ............. 1 g

Purified water is added to the whole ........... 900 ㎖

After mixing the above components according to the conventional healthy beverage manufacturing method, and stirred and heated at 85 ℃ for about 1 hour, the resulting solution is filtered and obtained by sterilization in a sterilized 2 ℃ container, sealed sterilized and stored in the refrigerator Used to prepare the healthy beverage composition of the invention.

Although the composition ratio is composed of the ingredients suitable for the preferred beverage in a preferred embodiment, the composition ratio may be arbitrarily modified according to regional and ethnic preferences such as demand hierarchy, demand country, and use purpose.

Claims (15)

Health food or pharmaceutical composition for itching or suppressing itching comprising chestnut extract as an active ingredient. The method of claim 1,
The composition is an itching that relieves or suppresses itching caused by one or more of inflammatory dermatitis, atopic dermatitis, dermatitis due to rough skin, sweat bands, erosion, frostbite, contact dermatitis, seborrheic dermatitis, dermatitis consisting of psoriasis and psoriasis Health food or pharmaceutical composition for alleviation or inhibition. The method of claim 1,
The composition is a health food or pharmaceutical composition for itching relief or inhibition to alleviate or suppress the itching caused by atopic dermatitis. Health food or pharmaceutical composition for improving skin barrier function comprising chestnut extract as an active ingredient. The method of claim 4, wherein
The composition is for improving the resilience of skin barrier damage or skin barrier damage derived from atopic dermatitis, health food or pharmaceutical composition for skin barrier improvement. The method of claim 4, wherein
The composition is a health food or pharmaceutical composition for improving the skin barrier is to improve skin moisturizing or prevent skin hyperkeratinization. Health food or pharmaceutical composition for immune suppression comprising chestnut extract as an active ingredient. The method of claim 7, wherein
The composition is a health food or pharmaceutical composition for immune suppression for the prevention or treatment of atopic diseases. The method of claim 7, wherein
The composition is a health food or pharmaceutical composition for immunosuppression for the treatment of atopy. Health food or pharmaceutical composition for improving or treating atopic dermatitis, comprising chestnut extract as an active ingredient. The method according to any one of claims 1 to 10,
The chestnut extract is a health food or pharmaceutical composition containing 0.005 to 80% by weight based on the total weight of the composition. The method according to any one of claims 1 to 10,
The chestnut is at least one health food or pharmaceutical composition selected from the group consisting of chestnut endothelium, the outer shell and mixtures thereof. The method of claim 12,
The chestnut is a skin or shell of the chestnut (health) or a pharmaceutical composition. The method according to any one of claims 1 to 10,
The chestnut extract is a health food or pharmaceutical composition extracted through at least one solvent selected from water, lower alcohols having 1 to 4 carbon atoms, 1, 3-butylglycol and mixed solvents thereof. The method of claim 14,
The chestnut extract is a health food or pharmaceutical composition is extracted through a solvent selected from the group consisting of water, methanol, ethanol, butanol, 1, 3-butylglycol and mixtures thereof.

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