JP2013063953A - Calpain activity inhibitor - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a calpain activity inhibitor with excellent safety, which includes a plant-derived constituent as an active constituent and can be utilized for preventing and treating various diseases caused by the excessive activation of calpain.SOLUTION: As a result of performing zealous research about the calpain activity inhibition action of various plant-derived constituents, one or more kinds of plants selected from plants belonging to Theaceae Camellia or the solvent extracts of the plants, particularly, Camellia sinensis (tea) or the solvent extract thereof is remarkably excellent in calpain activity inhibition action.

Description

  The present invention relates to a calpain activity inhibitor.

  Calpain is an intracellular proteolytic enzyme belonging to the cysteine protease group, and is found in many cells and tissues. Calpain is activated by increasing intracellular calcium concentration, calpain I or μ-calpain activated by μ-molar calcium ions, and calpain II or m activated by m-molar calcium ions. -Calpain is known and further calpain isozymes are envisaged.

  Calpain plays an important role in various physiological processes. Excessive activation of calpain is known to provide a molecular relationship between ischemia or injury induced by increased neuronal calcium and pathological neurodegeneration, such as cardiac ischemia, The onset of renal or central nervous system ischemia, inflammation, muscular dystrophy, cataract, diabetes, HIV disease, damage to the central nervous system, Alzheimer's disease, Huntington's disease, Parkinson's disease, multiple sclerosis, etc. It has been pointed out that it is strongly involved.

  Moreover, since it has been reported that the release of interleukin-1α is inhibited by a calpain activity inhibitor (Non-Patent Document 1), calpain may be involved in the aging phenomenon of the skin. Therefore, the calpain activity inhibitor is effective in delaying skin aging such as skin dryness, pigmentation, and wrinkle generation by inhibiting the production of active oxygen such as superoxide anion by interleukin-1α. I can expect.

  Calpain inhibitors are broadly classified as irreversible and reversible inhibitors and peptide and non-peptide inhibitors. Specifically, specific peptides (Patent Document 1), α-substituted hydrazides (Patent Document 2), indole carboxamides (Patent Document 3), carboxamide compounds (Patent Document 4) and the like have been reported. However, since these calpain inhibitors may cause side effects, there are concerns in terms of safety, and the price is not satisfactory. In addition, no studies have been reported so far on inhibitors of calpain enzymes, which contain plant-derived components with few problems in terms of safety as active ingredients.

Japanese National Patent Publication No. 9-500087 Japanese National Patent Publication No. 11-500124 JP-T-2001-515508 Japanese Translation of PCT International Publication No. 2010-514738

N. Watanabe et al., Cytokine vol. 6 (6), 597-601 (1994)

  The present invention is to provide a calpain activity inhibitor excellent in safety.

  As a result of diligent research on a calpain activity inhibitor comprising a plant-derived component that has few problems in terms of safety as an active ingredient, the present inventors have found that the Uji tea extract and the green tea extract have significantly superior calpain activity inhibitory action. And the present invention has been completed.

That is, the present invention
[1] An object of the present invention is to provide a calpain activity inhibitor comprising as an active ingredient one or more kinds of plants selected from the group consisting of plants belonging to the genus Camellia belonging to Camelliaaceae.
The plant belonging to the genus Camellia belonging to the camellia family is preferably tea leaves (tea), and in the case of a solvent extract, an extract obtained by extraction using ethanol, hydrous methanol or water is used as an active ingredient. It is preferable that
[2] The calpain activity inhibitor may be an external preparation.

  Conventionally, ingredients that have been effective as calpain activity inhibitors have the potential to cause side effects and have been a concern in terms of safety, but according to the present invention, various diseases caused by excessive activation of calpain Therefore, it is possible to provide a calpain activity inhibitor that can be used for prevention and treatment of diseases with few problems in terms of safety.

  Hereinafter, the configuration of the present invention will be described in more detail.

The calpain activity inhibitor according to the present invention comprises a plant belonging to the genus Camellia, or a solvent extract of the plant as an active ingredient.
The camellia genus of the Camellia family according to the present invention is an evergreen, shrub or tree that is known for about 200 species from the eastern part of India to the warm temperate zone of East Asia. Examples of plants belonging to the genus Camellia include tea (Camellia sinensis), camellia (Camellia japonica), sasanqua (Camellia sasanqua) and the like, and one or more of these can be used in combination. The kind of the Camellia family Camellia belonging to the present invention is not particularly limited, but tea (Camellia sinensis) is preferable. The leaves, buds, bark, sap, fruits, seeds, etc. of Camelliaaceae have medicinal effects, and the use site is not particularly limited, but leaves containing catechins are preferred.

  When the plant is used as an extract, the extraction solvent is not particularly limited, but the plant is a solvent, for example, a polar solvent such as water, lower alcohol, hydrous lower alcohol, propylene glycol, 1,3-butylene glycol, butanol, or chloroform. The extract obtained by extraction with an organic solvent such as ethyl acetate, various ethers or a mixture thereof is used as it is or after concentration, or the extract is adsorbed, for example, an ion exchange resin is used to remove impurities. It is also possible to use an extract that is adsorbed on a porous polymer column and then eluted with methanol or ethanol and concentrated. Also, a partitioning method, for example, an extract extracted with butanol can be used. Examples of commercial products include Uji tea extract and green tea extract from Maruzen Pharmaceutical Co., Ltd., as the camellia family camellia genus plant components, and these may be used. You can also use what you did.

  The plant or plant solvent extract selected from plants belonging to the genus Camellia obtained in this way has an excellent calpain activity inhibitory action, and prevents and treats various diseases caused by excessive activation of calpain. Useful for.

  It should be noted that extracts from other plants and the like containing catechins, which are the active ingredients of tea leaves (tea), also exhibit a calpain activity inhibitory effect.

  The calpain activity inhibitor of the present invention is highly safe and can be administered by any method of internal use and external use, but is preferably used as an external preparation. The calpain activity inhibitor which is a form of the external preparation of the present invention can be blended with commonly used external base materials, medicinal ingredients and the like. External base materials include hydrocarbons such as liquid paraffin, vegetable oils and fats, waxes, synthetic ester oils, silicone oil phase components, fluorine oil phase components, higher alcohols, fatty acids, thickeners, UV absorption Agent, powder, pigment, color material, anionic surfactant, cationic surfactant, nonionic surfactant, amphoteric surfactant, polyhydric alcohol, sugar, polymer compound, promoting transdermal absorption An agent, a solvent, an antioxidant, a fragrance, a preservative, and the like can be arbitrarily blended. As the medicinal component, an analgesic / anti-inflammatory agent, bactericidal / disinfectant, vitamins, emollient and the like can be used as appropriate. The dosage form of the external preparation of the present invention is arbitrary, and any agent based on an oil-based base, an oil-in-water type, a water-in-oil emulsion base, or a water-based agent Any type can be used. Moreover, as a use, it can use suitably for cosmetics, a skin external preparation, a pharmaceutical ointment, etc. The form may take any form such as lotion, lotion, emulsion, cream, pack, ointment, dispersion, solid, mousse and the like. The dosage of this product is not particularly limited as long as it is within the normal range, but it is usually used in the range of 0.001 to 2000 mg as the active ingredient of calpain activity inhibitor per product per day. The compounding amount in the external preparation varies depending on the use, dosage form, compounding purpose, etc., and is not particularly limited, but is 0.001 to 20.0% by mass in the external preparation as the active ingredient of the calpain activity inhibitor, preferably Is used in the range of 0.01 to 10.0% by mass.

  In addition, the calpain activity inhibitor which uses as an active ingredient the 1 type or 2 or more types of plant selected from the plant which belongs to the camellia family Camellia related to this invention as an external preparation, or the solvent extract of those plants is after ultraviolet irradiation. Has been found to significantly suppress the release of interleukin-1α (IL-1α). This suggests that the calpain activity inhibitor of the present invention inhibits the processing of calpain from Pro-IL-1α to IL-1α, which is a disease caused by an increase in IL-1α level. Expected to be useful for treatment.

  EXAMPLES The present invention will be specifically described below with reference to examples, but the technical scope of the present invention is not limited to these examples.

Evaluation of calpain activity inhibitory action of plant extracts

1. Outline of evaluation To evaluate calpain activity inhibitory action of green tea and Uji tea extract.

2. Evaluation Method Calpain I having a final concentration of 750 ng, green tea or Uji tea extract, and a calcium-containing buffer were added to a 96-well plate. After incubation on ice for 10 minutes, a final concentration of 2.5 μg of D-casein-FITC was added, and the reaction was started at 37 ° C. After reacting for 30 minutes, FITC fluorescence intensity change was measured with a fluorescence plate reader. In addition, each plant extract was evaluated using the solid content obtained by freeze-drying.

3. Evaluation Results In order to compare the calpain activity inhibitory action, the 50% inhibitory concentration (IC50) of each plant extract was evaluated. As a result, both green tea extract and Uji tea extract showed high calpain activity inhibitory action (Table 1).

Human anti-inflammatory action test of Uji tea extract

1. Outline of Evaluation Evaluate the UV-induced erythema inhibitory action (anti-inflammatory action) of Uji tea extract.

2. Evaluation Method 1.5 MED ultraviolet rays were irradiated with a solar simulator onto the site where the preparations containing 1%, 2% and 3% of Uji tea extract and the placebo preparations were applied. After the irradiation, the preparation was applied again, and the state of erythema after 24 hours was evaluated. Evaluation of erythema was carried out by colorimetry of a * value with a color difference meter, which was shown as the difference between the initial value and the a * value of the erythema site 24 hours after irradiation (delta a *) and compared with placebo.

3. Evaluation Results The action of the Uji tea extract combination preparation on erythema induced by ultraviolet irradiation is shown in FIG. In the part where the Uji tea extract combination preparation was applied, a decreasing tendency of delta a * was observed compared with the placebo application part, and the UV-induced erythema inhibitory action of the Uji tea extract was confirmed. It was also confirmed that the erythema inhibitory action of Uji tea extract was dependent on the blending concentration.

(Figure 1)

Next, examples of various preparations using the calpain activity inhibitor of the present invention are shown. About what has no description of a unit, a compounding quantity is the mass%. When Examples 3 to 7 were evaluated according to the evaluation method described in Example 1, a calpain activity inhibitory action was observed.

(tablet)
Uji tea water / ethanol extract 10.0g
Crystalline cellulose 50.0g
Lactose 20.0g
Carboxymethylcellulose 11.0g
Cornstarch 6.0g
Talc 2.0g
Light anhydrous silicic acid 0.5g
Magnesium stearate 0.5g
(Manufacturing method)
The above components were uniformly mixed, and compression-formed with a tableting machine to obtain tablets having a diameter of 8 mm and a mass of 180 mg.

(Granule)
Uji tea water extract 10.0g
Lactose 70.0g
16.5g starch
Hydroxypropylcellulose 3.5g
(Manufacturing method)
The above components were uniformly mixed and kneaded, then granulated by an extrusion granulator, dried, and sieved to obtain granules.

(Hydrophilic cream (hydrophilic ointment / O / W emulsified preparation))
Green tea water / ethanol extract 20.0
White petrolatum 25.0
Stearyl alcohol 20.0
Propylene glycol 12.0
Polyoxyethylene hydrogenated castor oil 60 4.0
Glycerol monooleate 1.0
Preservative appropriate amount 100% with purified water (production method)
A hydrophilic cream was prepared according to a conventional method.

(Water-absorbing cream (water-absorbing ointment / W / O emulsion))
Uji Tea Water / Butylene Glycol Extract 10.0
White petrolatum 40.0
Cetanol 10.0
Sara honey bees 5.0
Sorbitan sesquioleate 5.0
Lauro Macrogol 5.0
Preservative Appropriate amount Make the total amount 100 with purified water.
(Manufacturing method)
A water-absorbing cream was prepared according to a conventional method.

(Lotion preparation)
Uji tea water / ethanol extract 20.0
Concentrated glycerin 10.0
Butylene glycol 4.0
Trehalose 0.03
Sodium hyaluronate 0.01
Chelating agent
pH adjuster appropriate amount preservative appropriate amount make up to 100 with purified water (production method)
A lotion was prepared according to a conventional method.

Claims (4)

A calpain activity inhibitor comprising, as an active ingredient, one or more plants selected from plants belonging to the genus Camellia, or a solvent extract of those plants. The calpain activity inhibitor according to claim 1, wherein the plant belonging to the genus Camellia is tea tree (tea). The calpain activity inhibitor of Claim 1 or 2 which uses the extract obtained by extracting the said solvent using ethanol, water-containing ethanol, or water as an active ingredient. It is an external preparation, The calpain activity inhibitor of any one of Claims 1-3.