JP2009007328A - Antiallergic composition - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide an antiallergic composition, based on consideration of pharmacological action of Xiangru (an extract of whole grass of Mosla chinensis Maxim. ) which is a crude medicine, particularly Mosla chinensis Maxim. or Elsholtzia ciliata Hylander and having excellent antiallergic action based on chemical mediator isolation-suppressing action which the extract has and degranulation-inhibiting action. <P>SOLUTION: The antiallergic composition comprises an extract of Xiangru with a solvent as an active ingredient, and the Xiangru to be extracted is Mosla chinensis Maxim. or Elsholtzia ciliata Hylander and the solvent for extraction is water, a lower alcohol or a mixture of the lower alcohol with water, preferably ethanol. The antiallergic composition includes a food, especially a food for specified health use, a medicine, a cosmetic, etc. <P>COPYRIGHT: (C)2009,JPO&INPIT

Description

  The present invention relates to an antiallergic composition comprising a plant extract, in particular a solvent extract of Koju, especially Hosoyama Yamajiso or Naginata Koju.

The living body inherently has a homeostatic maintenance function and a natural healing function, and has a function capable of dealing with various internal and external environmental changes. Due to the changes and complexity, the number of people suffering from allergic diseases (various hay fever, atopic dermatitis, food allergies, childhood asthma, etc.) is rapidly increasing.
For the onset of this allergic condition, in terms of molecular biology, chemical mediators such as mast cells, histamine released by degranulation from basophils, or leukotrienes produced by type I allergic reactions, these cells It is said that inflammatory cytokines such as TNF-α produced from the plant are deeply related.

Therefore, antihistamines and the like are most commonly used as therapeutic agents for allergic diseases, but they are not effective for nasal clogging caused by hay fever, and antihistamines are inherently drowsy, dizzy, tired, etc. There is a problem that side effects are likely to occur.
Therefore, it is desired to develop a more effective antiallergic agent that does not have these side effects, but no effective antiallergic agent has yet appeared.

  The inventors of the present invention have been paying attention to the pharmacological activity of natural plants as a raw material, while actively searching for antiallergic agents. That is, although various pharmacological activities are recognized in natural plants, the present inventors have extensively screened for substances having an antiallergic action than natural products. As a result, the inventors have focused on the pharmacological effects of Koju.

Kouju (name of crude drug) refers to the whole grass of Hosobayamajiso, and Hosobayamajiso (scientific name: Mosla chinensis Maxim.) Is an annual plant belonging to the genus Mosla of Labiatae. Although it has been confirmed only slightly in the northern part of Kyushu, it is a plant distributed over a wide range in the Korean peninsula and mainland China.
Xiangru is used in China for symptoms such as summer cold, fever, headache, abdominal pain, and diarrhea (Non-Patent Document 1).

There have been many reports on the volatile components of Hosoyama Yamajiso, which contain many components such as thymol and carvacrol, and their strong antibacterial activity has been reported (Non-patent Document 2, Patent Document 1).
On the other hand, there are few reports on non-volatile components of Hosobayamajiso, and only a few types of flavonoids have been identified as the components (Non-patent Document 3).

  This time, the present inventors have an excellent chemical mediator release inhibitory action and an excellent degranulation inhibitory action from mast cells, basophils, etc., in the extract of Hosoyama Yamajiso or Naginata Kouji known as Kouji As a result, the inventors have newly found that they have antiallergic action and have completed the present invention.

Searching for substances with anti-allergic effects in various plants has been conducted, but among others, Perilla frutescens has been reported to have excellent anti-allergic action and is being used as a functional food (patents) Literature 2, Patent Literature 3). Recently, there is a report on the antiallergic action of Mosla dianthera, a plant belonging to the same Lamiaceae plant (Non-patent Document 4), but the extract of Hosobayamajiso or Naginata Koju has a chemical mediator release inhibitory action and mast cells and The present inventors have been confirmed for the first time to have an antiallergic action based on the degranulation inhibitory action from basophils, and the present invention is specific in that respect.
JP 2001-342106 A Japanese Patent No. 3071669 Japanese Patent No. 3303022 The People's Republic of China, 2005 edition, pages 182-183 Nakajodai Dictionary, 1985, pages 1421-1422, Shogakkan Indian Journal of Chemistry, 35: 392-294, 1996 Toxicology and Applied Pharmacology, 216 (2006), 479-484

  In the Chinese literature, the whole plant of Hosobayamajiso is described as Koju (name of herbal medicine), and the common herbal medicine is Hosobayamajiso. However, in the old book, Naginata Koju (Lamiaceae Naginata Koju, scientific name: Elsholtzia splendens Nakai ex F. Maekawa) similar to Hosobayamajiso was described as Kouji, and in China, depending on the region Naginata Koju may be used as Kouji. Also, in Japan, Naginata Kouji or Inu Kouji (Lamiaceae Inuco genus, scientific name: Mosla punctulata) may be treated as Kouji.

  Accordingly, in the present specification, Kouji generally refers to Mosla chinensis Maxim. Unless otherwise specified, it is broadly referred to as Kouji, including Naginata Kouge and Inu Koju.

  Therefore, in view of the above situation, the present invention is based on the examination of the pharmacological action of Koji, which is a crude drug, in particular Hosobayashi or Naginata, and the extract has an excellent chemical mediator release inhibitory action and mast cells, basophils, etc. An object of the present invention is to provide an antiallergic composition having an antiallergic action based on the degranulation inhibitory action from

The present invention for solving this problem is a chemical mediator release inhibitor characterized by comprising, as a basic aspect, a solvent extract of Koju as an active ingredient.
More specifically, the present invention is a chemical mediator release inhibitor, wherein the chemical mediator is one or more selected from histamine, serotonin, leukotriene, and TNF-α.

  In another basic aspect, the present invention is a degranulation inhibitor characterized by comprising a solvent extract of Koju as an active ingredient.

That is, in a more specific aspect of the present invention, the extract to be extracted is Hosobayamajiso or Naginata Kouji, preferably a whole herb dried product, and the extraction solvent is water, a lower alcohol or a mixed solvent of lower alcohol and water. It is a chemical mediator release inhibitor or degranulation inhibitor.
Among them, a more preferred specific present invention is a chemical mediator release inhibitor or degranulation inhibitor in which the lower alcohol as the extraction solvent is ethanol.

  Therefore, the most preferable present invention is a chemical mediator release inhibitor or degranulation inhibitor characterized by comprising, as an active ingredient, an ethanol extract of a dried whole plant of Hosobayamajiso or Naginata Kouji.

Moreover, this invention is an antiallergic composition containing said chemical mediator release inhibitor or a degranulation inhibitor as another aspect.
That is, in more detail, it is an antiallergic composition characterized by comprising as an active ingredient a solvent extract of Koju, which is a herbal medicine, and in particular, an antiallergic composition in which Koju is Hosobayamajiso or Naginata Koju .

More specifically, the anti-allergic composition in which the Kouji to be extracted is Hosobayamajiso or Naginata Kouji, preferably a whole herb dried product, and the extraction solvent is water, a lower alcohol or a mixed solvent of lower alcohol and water. It is a thing.
Among them, a more preferred specific invention is an antiallergic composition in which the lower alcohol as the extraction solvent is ethanol.

  Therefore, the most preferred present invention is an antiallergic composition characterized in that it comprises, in particular, an ethanol extract of a dried whole plant of Hosobayamajiso or Naginata Kouji as an active ingredient.

  Moreover, this invention is foodstuff containing the above-mentioned antiallergic composition as another aspect, especially food for specified health, and a pharmaceutical containing the above-mentioned antiallergic composition.

  Furthermore, this invention is cosmetics containing the above-mentioned antiallergic composition as another aspect, and is also a feed.

Chemical mediator release inhibitory action and degranulation comprising the solvent extract of Hosoyama Yamajiso or Naginata Koju provided by the present invention (hereinafter, widely referred to as “extract extract” in some cases, including the solvent extract of Koju). The antiallergic composition based on the inhibitory action, as will be apparent from the test examples described below,
(1) It has the effect of suppressing the release of chemical mediators such as histamine produced by degranulation inhibition from mast cells and basophils, or leukotriene produced by type I allergic reaction.
(2) It has the effect of inhibiting the production of TNF-α from mast cells and basophils.
(3) It has an effect of alleviating allergic symptoms of allergic rhinitis model mice.
(4) It has the effect of suppressing weight loss of an allergic rhinitis model mouse (improvement of systemic functional decline).
Therefore, by applying the antiallergic composition containing the extract of the present invention, there is an advantage that it is possible to provide a highly safe antiallergic agent (pharmaceutical, health food, cosmetics) that can be easily consumed in daily life. is doing.

In the present invention, Mosla chinensis Maxim., Which constitutes an extract that is an active ingredient, is an annual plant belonging to the genus Mosla of Labiatae as described above. In Japan, only a few have been confirmed from the Chugoku region to the northern part of Kyushu, but in the Korean peninsula and the mainland of China, the plant is distributed over a wide range.
This dry herbal medicine is called Xiangru, and is one of the Chinese herbal medicine ingredients used for symptoms such as summer cold, fever, headache, abdominal pain, diarrhea in China.
In addition, Naginata Koju (Lamiaceae Naginata Koju genus, scientific name: Elsholtzia ciliata Hylander, Elsholtzia splendens Nakai ex F. Maekawa) has been described as Kouji, and in China, Naginata Koju is used as Kouji in some regions. It may be used. Also, in Japan, Naginata Kouji or Inu Kouji (Lamiaceae Inuco genus, scientific name: Mosla punctulata) may be treated as Kouji.
It has not been known until now that this solvent extract of Hosobayamajiso or Naginata kouju has an antiallergic action, and the present invention is specific in that respect.

  In the present invention, in preparing the extract that is the active ingredient, as the Hosobayamajiso or Naginatakouju, for example, dried parts of stems, leaves, flowers, fruits, etc., or dried whole plants including these It is preferable to use dried whole grass.

In the extraction, it is preferable to appropriately cut the dried product and perform solvent extraction. As the solvent used for the extraction, water, a lower alcohol or a mixed solvent of lower alcohol and water is preferable, and it is particularly preferable to extract using ethanol.
Extraction using hot water or extraction using a mixed solvent of lower alcohol and water is possible, but according to the study by the present inventors, the antiallergic action of the extract is particularly The ethanol extract was stronger.

  As a specific method of extraction, commonly used extraction means is adopted, for example, after appropriately cutting the dried whole herb of Hosoyama Yamajiso or Naginata Koju, 1 to 50 times the total weight, Preferably, 10 to 20 times the amount of water, a lower alcohol or a mixed solvent of lower alcohol and water, preferably ethanol, is used for about 1 to 24 hours, and is immersed and heated for extraction in the range of room temperature to the boiling point of the solvent used, The obtained extract is concentrated under reduced pressure, the used extraction solvent is removed, and a target solvent extract of Hosobayamajiso or Naginatakouju can be prepared.

  Thus, it is possible to prepare an extract of Hosoyama Yamajiso or Naginata Kouji, which is an active ingredient of the present invention, and as a result of studies by the present inventors, the obtained extract has an excellent inhibitory effect on the release of chemical mediators and mast cells. It has been found that it has an excellent antiallergic action based on the degranulation inhibitory action from basophils.

  Therefore, the present invention is a chemical mediator release inhibitor or degranulation inhibitor containing such an extract of Hosobayamajiso or Naginata kouju as an active ingredient, and an antiallergic agent containing these chemical mediator release inhibitor or degranulation inhibitor It is a composition.

The chemical mediator release inhibitor or degranulation inhibitor provided by the present invention can be prepared as various forms of preparations by conventional methods using commonly used excipients, additives and the like.
Moreover, the antiallergic composition of this invention can be manufactured by processing into various forms by a normal method using said chemical mediator release inhibitor or a degranulation inhibitor. For example, it can be processed into a solid form, liquid form, emulsified form, paste form, jelly form or the like, and can be produced as a medicine or a food.

  That is, the antiallergic composition provided by the present invention can be effectively applied as a food in addition to a pharmaceutical product. The antiallergic composition of the present invention can be provided as a food that can be taken immediately as it is, a food that can be taken after cooking, or a premixed material for food production.

  The food containing the antiallergic composition of the present invention may be in any form of solid, powder or granule. Specific examples include, but are not limited to, various confectionery such as biscuits, cookies, cakes, snacks, rice crackers, breads, powdered beverages (powdered coffee, powdered cocoa, etc.), rice cakes, caramels and the like. It is not a thing.

  Moreover, various drinks, such as juice, a carbonated drink, a lactic acid bacteria drink, can be mentioned as a liquid, an emulsion form, a paste form, and a jelly form, It is not limited to these. Among them, it is preferably a paste, jelly, or liquid

In particular, when the antiallergic composition of the present invention is used as a pharmaceutical product, it can be made into an ordinary preparation in the form of a tablet, powder, granule, fine granule, liquid or the like. These preparations can be formulated according to a conventional method together with other pharmaceutically acceptable additives, for example, carriers such as excipients, binders and lubricants.
In addition, sweeteners, colorants and the like can be added and taken. These preparations can be carried out by the method described in the Japanese Pharmacopoeia General Rules for Preparations.

  In the antiallergic composition of the present invention, the extract used as an active ingredient should be ingested in an amount of about 1 to 20 g, preferably about 2 to 10 g per day, based on the antiallergic action as a standard. Specifically, the content of the active ingredient to be taken 1 to 3 times a day, preferably 3 times a day immediately before each meal, may be set and designed as a preparation for ingesting the dose.

  When the extract itself is contained, the extract can be diluted with an appropriate solvent to prepare a liquid composition. Alternatively, the extract itself can be mixed with a suitable carrier, preferably a fatty acid triglyceride, and filled in a soft capsule or the like in a liquid state.

  In the above formulation, commonly used excipients, binders, disintegrants, lubricants, stabilizers, surfactants, solubilizers, reducing agents, buffers, adsorbents, fluidizing agents, Select one or more kinds of formulation additives such as antistatic agents, antioxidants, sweeteners, flavoring agents, cooling agents, light-shielding agents, flavoring agents, fragrances, fragrances, coating agents, plasticizers, etc. Needless to say, it may be added.

  Specific examples of such formulation additives include crystalline cellulose, low-substituted hydroxypropylcellulose, hydroxypropylmethylcellulose, croscarmellose sodium, maltodextrin, ethylcellulose, lactose, sorbitol, silicic anhydride, and silicic acid. Magnesium, hydroxypropyl cellulose, stearic acid, oleic acid, liquid paraffin, dicalcium phosphate, dibutyl sebacate, macrogol, propylene glycol, corn starch, starch, pregelatinized starch, gelatin, popidone, crospovidone, glycerin, polysorbate 80, citrus Acid, acesulfame potassium, aspartame, sodium carbonate, talc, magnesium stearate, calcium stearate, etc. That.

  In preparation of the anti-allergic composition of the present invention, other ingredients such as vitamins and minerals are added within a range of quality and quantity that does not impair the anti-allergic action of the extract as an active ingredient. It can also be made.

  Furthermore, although this invention is an antiallergic composition which contains the extracted extract prepared above as an active ingredient, this composition can be used as a foodstuff. That is, the antiallergic composition prepared as described above can be directly mixed with food and other ingredients to form a food for specified health use (special insurance) or animal feed.

  Hereinafter, the present invention will be described in more detail by describing specific examples and test examples. However, the scope of the present invention is not limited to these examples.

Example 1 Preparation of Ethanol Extract of Hosoyama Yamajiso 1 L of ethanol was added to 100 g of dried whole material of Hosobayamajiso, heated and extracted at 70 ° C. for 2 hours, filtered, concentrated to dryness under reduced pressure, and 4.5 g An extract was obtained.

Example 2: Preparation of a water extract of Hosoyama Yamajiso 1 L of water was added to 100 g of the dried material of Hosobayamajiso, heated and extracted at 70 ° C for 2 hours, filtered, concentrated to dryness under reduced pressure, and 10.1 g An extract was obtained.

Example 3: Preparation of ethanol extract of Naginata ethanol extract of ethanol extract of Naginata was prepared in the same manner as in Example 1 above. That is, using 100 g of dried whole plant of Naginata, 1 L of ethanol was added, heated and extracted at 70 ° C. for 2 hours, filtered, concentrated to dryness under reduced pressure, and 4.4 g of extract was obtained. It was.

Example 3: Preparation of water extract of Naginata kouju Water extract of Naginata kouju was prepared in the same manner as in Example 2 above. That is, 1 L of water was added to 100 g of dried whole herb of Naginata, extracted by heating at 70 ° C. for 2 hours, filtered, and concentrated to dryness under reduced pressure to obtain 18.5 g of extract.

  The anti-allergic action effect based on the chemical mediator release inhibitory action and degranulation inhibitory action of the extract of Hosobayamajiso or Naginata kouju obtained above will be demonstrated by the following test examples.

Test Example 1: Effect test on degranulation The degranulation inhibition test is released together with histamine and the like by degranulation from rat basophil leukemia cells (RBL-2H3: obtained from Tohoku University Institute of Aging Medicine and Medical Cell Resource Center). The method of measuring the activity of β-hexosaminidase was used.

[Method]
That is, RBL-2H3 cells grown by subculture were released into a MEM culture solution containing 10% fetal bovine serum, and 400 μL of cells at a concentration of 3.5 × 10 ⁵ cells / mL were added to a 48-well plate. After 60 minutes, 100 μL of anti-dinitrophenyl (DNP) -IgE antibody was added to a final concentration of 0.45 μg / mL, and the cells were cultured at 37 ° C. for 24 hours with 5% carbon dioxide gas. The cultured RBL-2H3 cells were washed with 500 μL of siraganian buffer (119 mM sodium chloride, 5 mM potassium chloride, 0.4 mM magnesium chloride, 25 mM PIPES, 40 mM sodium hydroxide, pH 7.2), and then siraganian containing bovine serum albumin (BSA). 160 μL of buffer (containing 5.6 mM glucose, 1 mM calcium chloride, 0.1% BSA) was added, and the mixture was cultured at 37 ° C. for 10 minutes with 5% carbon dioxide gas.

To this was added 20 μL of DMSO or a sample diluted to an appropriate amount with DMSO (DMSO final concentration 0.1%), and cultured at 37 ° C. for 10 minutes under 5% carbon dioxide gas. Next, 20 μL of DNP-BSA serving as an antigen was added to a final concentration of 10 μg / mL, followed by culturing at 37 ° C. with 5% carbon dioxide for 30 minutes to induce degranulation. Thereafter, the reaction was stopped on ice for 10 minutes and centrifuged (1000 rpm, 4 ° C., 5 minutes), and then 50 μL of the supernatant was transferred to a 96-well plate, to which 1 mM PNAG (p-nitrophenyl-N-acetyl-β-D-glucosamide was added). 50 μL of 3.42 g prepared in 100 mL of 0.1 M citrate buffer pH 4.5 was added, and reacted for 60 minutes at 37 ° C. with 5% carbon dioxide gas. After the reaction, 200 μL of stop buffer (0.1 M NaHCO ₃ / Na ₂ CO ₃ , pH 10) was added, the absorbance at 405 nm was measured, and the β-hexosaminidase release inhibition rate (%) was determined by the following formula.

β-hexosaminidase release inhibition rate (%) = [1− (C−B−D) / (A−B)] × 100
In the above,
A: DNP-BSA stimulation, absorbance value of DMSO only B: DNP-BSA non-stimulation, absorbance value of DMSO only C: DNP-BSA stimulation, absorbance value of sample (+) D: Absorption value of sample (+) only

  As test samples, the ethanol extract of Hosobayamajiso obtained in Example 1 was 10, 50, 100, and 200 μg / mL, and the water extract of Hosobayamajiso obtained in Example 2 was 10, 50, 100, and 200 μg / mL. 100 and 200 μg / mL of the extract of Naginata ethanol obtained in Example 3 and 100 μg / mL of the aqueous extract of Naginata Koju obtained in Example 4 were used.

As a control sample, 100 μg / mL each of an extract of ethanol and an extract of water from a leaf prepared in the same manner as in Examples 1 and 2 from perilla (soba) was used.
Further, azelastine (generic name) and tranilast (generic name) at 10 and 100 μg / mL were used as activity controls.
[result]
The obtained inhibitory effect (n = 3) on β-hexosaminidase release from RBL-2H3 cells of each test sample is shown in Table 1 below.
The inhibition rate was expressed as an average value ± standard deviation.

  As can be seen from the results shown in the table, an inhibitory effect was observed in the ethanol and water extract of Hosobayamajiso and the ethanol extract of Naginata of the present invention. In particular, it is found that the ethanol extract of Hosobayamajiso is stronger than soba and shows an excellent inhibition rate.

Test Example 2: effect test for TNF-alpha production [Method]
As in Test Example 1, RBL-2H3 cells grown by subculture were suspended in a MEM culture solution containing 10% fetal bovine serum, and the concentration was 7.5 × 10 ⁵ cells / mL in a 24-well plate. The cells were seeded at 400 μL, and after 60 minutes, 100 μL of anti-DNP-IgE antibody was added to a final concentration of 0.45 μg / mL, followed by culturing at 37 ° C. with 5% carbon dioxide for 24 hours. The cultured RBL-2H3 cells were washed with a culture solution, 160 μL of the culture solution was added, and the cells were cultured for 10 minutes at 37 ° C. with 5% carbon dioxide gas. To this was added 20 μL of DMSO or a sample diluted to an appropriate amount with DMSO (DMSO final concentration 0.1%), and cultured at 37 ° C. for 10 minutes under 5% carbon dioxide gas. Next, 20 μL of DNP-BSA serving as an antigen was added to a final concentration of 10 μg / mL, and the cells were cultured for 4 hours at 37 ° C. with 5% carbon dioxide gas.
The concentration of TNF-α produced in the culture solution was measured by sandwich ELISA (Rat TNF OptEIA ELISA Set. Manufactured by Nippon Becton Dexon), and the inhibition rate (%) of TNF-α production was determined by the following formula. It was.

TNF-α production inhibition rate (%) = [1- (C−B) / (A−B)] × 100
In the above,
A: DNP-BSA stimulation, absorbance value of DMSO only B: DNP-BSA non-stimulation, absorbance value of DMSO only C: DNP-BSA stimulation, absorbance value of sample (+)

  As test samples, the ethanol extract of Hosoyama Yamajiso obtained in Example 1 was 10, 50, 100, and 200 μg / mL, and the water extract of Hosoyamajiso obtained in Example 2 was 100 and 200 μg / mL, Example 3. 100 and 200 μg / mL of the ethanol extract of Naginata obtained in the above, and 100 μg / mL of the water extract of Naginata obtained in Example 4 were used.

In addition, as a control sample, 100 μg / mL each of an extract of ethanol and an extract of water from a leaf prepared in the same manner as in Examples 1 and 2 from perilla (soba) was used.
Furthermore, azelastine (generic name) and tranilast (generic name) at 10 and 100 μg / mL were used as activity controls.

[result]
The obtained inhibitory effect (n = 3) on TNF-α production from RBL-2H3 cells of each material is shown in Table 2 below.
The inhibition rate was expressed as an average value ± standard deviation.

  As can be seen from the results shown in the table, an inhibitory effect was observed in the ethanol and water extract of Hosobayamajiso and the ethanol extract of Naginata of the present invention. In particular, in the ethanol extract of Hosoyama Yamajiso, an excellent dose-dependent inhibitory effect on TNF-α production was observed.

Test Example 3: Effect test on leukotriene production [Method]
As in Test Example 1, RBL-2H3 cells grown by subculture were released into a MEM culture solution containing 10% fetal bovine serum, and the concentration was 7.5 × 10 ⁵ cells / mL in a 24-well plate. The cells were seeded at 400 μL, and after 60 minutes, 100 μL of anti-DNP-IgE antibody was added to a final concentration of 0.45 μg / mL, followed by culturing at 37 ° C. for 5 hours with 5% carbon dioxide. The cultured RBL-2H3 cells were washed with a culture solution, 160 μL of the culture solution was added, and the cells were cultured for 10 minutes at 37 ° C. with 5% carbon dioxide gas.

To this was added 20 μL of DMSO or a sample diluted to an appropriate amount with DMSO (DMSO final concentration 0.1%), and cultured at 37 ° C. for 10 minutes under 5% carbon dioxide gas. Next, 20 μL of DNP-BSA serving as an antigen was added to a final concentration of 10 μg / mL, and the cells were cultured for 1 hour at 37 ° C. with 5% carbon dioxide gas.
The concentration of leukotriene produced in the culture was measured by the competitive EIA method (Cysteinyl-Leukotriene EIA KIT, manufactured by Cayman Chemical Company), and the leukotriene production inhibition rate (%) was determined by the following formula.

Leukotriene production inhibition rate (%) = [1− (C−B) / (A−B)] × 100
In the above,
A: DNP-BSA stimulation, absorbance value of DMSO only B: DNP-BSA non-stimulation, absorbance value of DMSO only C: DNP-BSA stimulation, absorbance value of sample (+)

  As test samples, 2, 10, 50, and 100 μg / mL of the extract of Hosoyama Yamajiso obtained in Example 1 and 50, 100, and 200 μg / mL of the water extract of Hosoyama Yamajiso obtained in Example 2 were used. The ethanol extract of Naginata obtained in Example 3 was used at 50 and 100 μg / mL, and the water extract of Naginata obtained in Example 4 was used at 100 μg / mL.

In addition, as a control sample, 100 μg / mL each of an extract of ethanol and an extract of water from a leaf prepared in the same manner as in Examples 1 and 2 from perilla (soba) was used.
Furthermore, 10 and 100 μg / mL azelastine (generic name) and 50 and 100 μg / mL tranilast (generic name) were used as activity controls.

[result]
The obtained inhibitory effect (n = 3) on leukotriene production from RBL-2H3 cells in each material is shown in Table 3 below.
The inhibition rate was expressed as an average value ± standard deviation.

  As can be seen from the results shown in the table, the leukotriene production inhibitory effect was observed in the ethanol and water extract of Hosobayamajiso and the ethanol extract of Naginata of the present invention. In particular, the ethanol extract of Hosoyama Yamajiso was shown to have excellent inhibitory activity.

Test Example 4: Toxicity test for cells [Method]
As in Test Example 1, RBL-2H3 cells grown by subculture were released into a MEM culture solution containing 10% fetal bovine serum, and the concentration was 7.5 × 10 ⁵ cells / mL in a 96-well plate. 100 μL of each cell was seeded and cultured at 37 ° C. for 24 hours with 5% carbon dioxide gas. The cultured RBL-2H3 cells were washed with a culture solution, 90 μL of the culture solution was added, and the cells were cultured for 10 minutes at 37 ° C. with 5% carbon dioxide gas.

To this was added 10 μL of DMSO or a sample diluted to an appropriate amount with DMSO (DMSO final concentration 0.1%), and cultured at 37 ° C. for 24 hours under 5% carbon dioxide gas. After the cultivation, the survival rate was measured using a cell proliferation assay kit (Cell counting Kit-8, manufactured by Dojindo Laboratories).
The survival rate (%) is the survival rate when each sample was added with the control (absorbance when DMSO alone was added) being 100%.

  As test samples, 50 and 100 μg / mL of the extract of Hosoyamajiso obtained in Example 1 and 50 and 100 μg / mL of the water extract of Hosoyamajizo obtained in Example 2 were obtained in Example 3. 50 and 100 μg / mL of Naginata ethanol extract was used, and 50 and 100 μg / mL of Naginata water extract obtained in Example 4 were used.

Further, as a control sample, 50 and 100 μg / mL of an ethanol extract and an aqueous extract of soba prepared from perilla (Soba) in the same manner as in Examples 1 and 2 were used.
Furthermore, 2, 10 and 50 μg / mL of azelastine (generic name) and 10 and 50 μg / mL of tranilast (generic name) were used as activity controls.

[result]
The survival rate (n = 3) of each material obtained is shown in Table 4 below.

  As can be seen from the results shown in the table, the extract of Hosobayamajiso or Naginatakouju of the present invention has a high survival rate of 90% or more even after 24 hours from the addition, and it was judged that there is no toxicity to cells. . From this, it was suggested that the degranulation inhibitory action and each chemical mediator production inhibitory action of the extract of Hosobayamajiso or Naginata Koju of the present invention are not due to cytotoxicity.

Test Example 5: Effect Test against allergic rhinitis mouse model [Method]
10 μg of OVA (ovalbumin) and 1.3 mg of aluminum hydroxide gel were dissolved in 0.2 mL of physiological saline, and peritoneally into the abdominal cavity of balb / c mice (6 weeks old / female) once a week for a total of 3 times. Made. Furthermore, from one week after the final sensitization, sneezing and nasal scratching behaviors were induced by nasal nasal injection of 10 μL of OVA 25 mg / mL solution into both nasal cavities for 7 days.
The ethanol extract of Hosoyama Yamajiso obtained in Example 1 and the active control azelastine were suspended in a 0.5% carboxymethylcellulose (CMC) solution and used for administration. The test group consisted of a control group (10 mice) administered with 0.5% CMC only, 3 groups (10 mice each) administered with Hosoyama Yamajiso extract at doses of 50, 200, and 800 mg / kg, and 10 mg / kg of azelastine. An administered group (7 animals) was provided. The test substance is administered orally once every hour during the OVA nasal nasal period, and the number of sneezes and nasal scratches for 10 minutes immediately after nasal nasal on the last day of the OVA nasal nose is measured. The effect was evaluated.
For tests between groups, Bonferroni's multiple comparison test was used, and those with a risk rate of 5% or less were judged to be significantly different.

[result]
Table 5 below shows the effect of Hosoyama Yamajiso extract and azelastine as an activity control on the sneezing and nasal scratching behavior of allergic rhinitis mice.
Moreover, those results were shown as a graph in FIG.1 and FIG.2.

FIG. 1 is a diagram showing the action on sneezing in allergic rhinitis model mice. The control and Hosobayamajiso-administered groups each represented 10 animals, and the azelastine-administered group represented 7 data with mean ± standard deviation.
In the figure, *: p <0.05, **: p <0.01, statistically different from control (Bonferroni's multiple comparison test)

  FIG. 2 is a diagram showing an effect on nasal scratching behavior of allergic rhinitis model mice. The control and Hosobayamajiso-administered groups each represented 10 animals, and the azelastine-administered group represented 7 data.

＊：危険率５％以下で無投与対照群と比較し、有意差あり。
＊＊：危険率１％以下で無投与対照群と比較し、有意差あり。

*: Significant difference compared to untreated control group with a risk rate of 5% or less.
**: Significant difference compared to untreated control group with a risk rate of 1% or less.

A dose-dependent sneezing and suppression of nasal scratching were observed in the group to which the extract of Hosoyama Yamajiso of the present invention was orally administered relative to the control group (0.5% CMC administration group). In particular, with regard to the number of sneezes, a statistically significant inhibitory effect was observed from the doses of 200 mg / kg and 800 mg / kg extract of Hosoyama Yamajiso extract.
Also in the active control azelastine, the administration of 10 mg / kg showed the effect of suppressing sneezing and nasal scratching.

FIG. 3 shows the effects of the extract of Hosobayamajiso and azelastine of the present invention on the change in body weight during the antigen nasal period for 7 days.
As can be seen from the results in the figure, in the control group (0.5% CMC administration group), a gradual decrease in body weight was observed from the start of the antigen nasal drop to the last day. Then, the effect of suppressing weight loss from a dose of 200 mg / kg was observed.
In the azelastine administration group, which is an activity control, the effect of suppressing weight loss was not observed.

From the above test results, the extract of Hosoyama Yamajiso of the present invention has an effect of suppressing the release of chemical mediators such as histamine by inhibiting degranulation from mast cells and basophils. From this, it was confirmed to have an effect of inhibiting the production of TNF-α.
Furthermore, it has an effect of alleviating allergic symptoms of allergic rhinitis model mice, and in addition, it has been confirmed to have an effect of suppressing weight loss of allergic rhinitis model mice (improvement of systemic functional decline).

Test Example 4: Toxicity test [Method]
Toxicity tests were conducted using ddy mice (7 weeks old, male) by single large dose administration of the extract of Hosobayamajiso.
The ethanol extract of Hosoyama Yamajiso obtained in Example 1 was suspended in a 0.5% CMC solution and used for oral administration. The test group was provided with a control group to which only 0.5% CMC was administered, and a group to which Hosobayamajiso extract was administered at a dose of 2500 mg / kg (3 mice for each group), and the state of the mice for 14 days after the administration was observed. .

[result]
General symptoms and body weight course for 14 days after administration of Hosoyama Yamajiso extract were maintained in the same state as the control group. On day 14 after administration, the abdomen was opened and each organ (thymus, heart, lung, liver, spleen, kidney, testis) was removed, and the organ weight was measured and macroscopically observed. There were no abnormalities.

Example 3: Pharmaceutical containing the extract of Koji (Hosobayashi or Naginata) Extract 55.0 g of the extract of Hosobayamajiso obtained in Example 1 or the extract of Naginata Koji obtained in Example 2 and 249.5 g of lactose and stearic acid Tablets having a diameter of 10 mm and a weight of 300 mg were produced by tableting with 0.5 g of magnesium using a single-punch tableting machine.

(2) Powder:
Magnesium stearate 0.5 g is added to 100.0 g ethanol extract of Hosobayamajiso obtained in Example 1 or Naginata kouju obtained in Example 2 and compressed, crushed, granulated, sieved, and 20-50 mesh granules An agent was obtained.

Example 4: Various foods and drinks containing an ethanol extract of Koji (Hosobayashi or Naginata) In the composition shown below, Hosobayashi moji obtained in Example 1 or Naginata Koji ethanol extract (Example 2) Hereinafter, various foods and drinks containing simply extract) were produced.
(1) 飴:
(Composition) (Parts by weight)
Powdered sorbitol 89.70
Fragrance 0.25
Extracted extract 10.00
Sorbitol seed 0.05
Total amount 100.00

(2) Gum:
(Composition) (Parts by weight)
Gum base 20.00
Calcium carbonate 2.00
Stevioside 0.10
Extracted extract 10.00
Lactose 66.90
Perfume 1.00
Total amount 100.00

(3) Caramel:
(Composition) (Parts by weight)
Granulated sugar 32.00
Minamata 20.00
Milk powder 30.00
Hardened oil 4.00
Salt 0.60
Perfume 0.03
Water 3.37
Extracted extract 10.00
Total amount 100.00

(4) Carbonated drink:
(Composition) (Parts by weight)
Granulated sugar 8.00
Concentrated lemon juice 1.00
L-ascorbic acid 0.10
Citric acid 0.09
Sodium citrate 0.05
Coloring agent 0.05
Carbonated water 80.71
Extracted extract 10.00
Total amount 100.00

(5) Juice:
(Composition) (Parts by weight)
Frozen concentrated orange juice 5.00
Fructose glucose liquid sugar 1.00
Citric acid 0.10
L-ascorbic acid 0.09
Extracted extract 10.00
Fragrance 0.20
Dye 0.10
Water 83.51
Total amount 100.00

(6) Lactic acid bacteria beverage:
(Composition) (Parts by weight)
Milk solid 21% fermented milk 14.76
Fructose dextrose liquid sugar 13.31
Pectin 0.50
Citric acid 0.08
Fragrance 0.15
Water 61.20
Extracted extract 10.00
Total amount 100.00

(7) Alcoholic beverages:
(Composition) (Parts by weight)
50% ethanol 32.00
Sugar 8.60
Fruit juice 2.40
Extracted extract 10.00
Water 47.00
Total amount 100.00

Example 5: Various cosmetics containing an ethanol extract of Koji (Hosobayashi or Naginata) In the composition shown below, the extract of Hosobayashi obtained in Example 1 or the ethanol extract of Naginata obtained in Example 2 (below) And various cosmetics containing the extract).
(1) Emollient cream:
(Composition) (Parts by weight)
Beeswax 2.0
Stearyl alcohol 5.0
Stearic acid 8.0
Squalane 10.0
Self-emulsifying propylene glycol 3.0
Monostearate polyoxyethylene cetyl ether (20EO) 1.0
Fragrance 0.5
Antioxidant Trace amount of preservative Trace amount of propylene glycol 4.8
Glycerin 3.0
Sodium hyaluronate 0.1
Extract extract 0.1
Triethanolamine 1.0
Purified water 61.5
Total amount 100.0

(2) Emulsion foundation:
(Composition) (Parts by weight)
Stearic acid 2.4
Propylene glycol monostearate 2.0
Cetostearyl alcohol 0.2
Liquid lanolin 2.0
Liquid paraffin 3.0
Isopropyl myristate 8.5
Propyl paraoxybenzoate Trace purified water 64.1
Sodium carboxymethylcellulose 0.2
Bentonite 0.5
Propylene glycol 3.8
Sodium hyaluronate 0.1
Extract extract 0.1
Triethanolamine 1.1
Methyl parahydroxybenzoate Trace amount of titanium oxide 8.0
Talc 4.0
Coloring content Trace fragrance Trace amount
Squalane 10.0
Total amount 100.0

  As described above, the present invention provides an antiallergic composition having an excellent antiallergic action, which the extract thereof has, based on the study of the pharmacological action of natural plants, specifically Koji, Hosobayashi and Naginata. Because it is provided, it is possible to provide a highly safe antiallergic agent (pharmaceuticals, health food, cosmetics, etc.) that can be easily consumed in daily life by applying the extract of Hosobayamajiso or Naginata Koju of the present invention. The industrial applicability is tremendous.

It is a figure which shows the effect | action with respect to the sneezing of the allergic rhinitis model mouse in Experiment 3. It is a figure which shows the effect | action with respect to the nasal scratching behavior of the allergic rhinitis model mouse in Test Example 3. It is a figure which shows the body weight change of the model mouse in Test Example 3.

Claims (13)

  A chemical mediator release inhibitor characterized by comprising a solvent extract of Koju as an active ingredient.   The chemical mediator release inhibitor according to claim 1, wherein the chemical mediator is one or more selected from histamine, serotonin, leukotriene, and TNF-α.   A degranulation inhibitor comprising a solvent extract of Koju as an active ingredient.   The chemical mediator release inhibitor or the degranulation inhibitor according to any one of claims 1, 2, and 3, wherein Koju is Hosobayamajiso or Naginata Koju.   4. The chemical mediator release inhibitor or degranulation inhibitor according to any one of claims 1, 2 and 3, wherein the solvent extract of Koju is a solvent extract of Hosobayamajiso or Naginata Koju.   6. The chemical mediator release inhibitor or degranulation inhibitor according to claim 5, wherein the extraction solvent is water, a lower alcohol or a mixed solvent of lower alcohol and water.   The chemical mediator release inhibitor or degranulation inhibitor according to claim 6, wherein the lower alcohol is ethanol.   The antiallergic composition containing the chemical mediator release inhibitor or degranulation inhibitor according to any one of claims 1 to 7.   A food containing the antiallergic composition according to claim 8.   The food according to claim 9, wherein the food is a food for specified health use.   A medicament comprising the antiallergic composition according to claim 8.   Cosmetics containing the antiallergic composition of Claim 8.   A feed containing the antiallergic composition according to claim 8.

Images