JP2001048799A - Medicine for treating tick allergy - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain the subject medicine which is safe and can be produced in a large amount by including a tea extract. SOLUTION: This medicine for treating tick allergy contains a tea extract, preferably a green tea extract. The tea extract is preferably a tea extract which contains one or more polyphenol compounds selected from (-)-epicatechin, (-)-epigallocatechin, (-)-epicatechin gallate, (-)-epigallocatechin gallate, and so on. The medicine is usually orally administered at a dose of 1 to 1,000 mg, preferably 50 to 500 mg, once to several times for an adult, or is coated on a diseased par in the form of an ointment containing the tea extract in an amount of 0.1 to 20% for once to several times a day for an adult.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to an agent for treating mite allergy.

[0002]

It is known that house dust mites are deeply involved in exogenous asthma, allergic rhinitis and atopic dermatitis [J. Allergy, 39, 325 (19
67) and Ibid., 42, 14 (1968)]. Dermatophagoides pteronisinas, a kind of house dust mite
yssinus), two types of proteases (proteolytic enzymes) have been isolated, and the proteolytic activity of these enzymes is thought to be closely related to mite allergic disease [Lancet, Jul. , 15, 154 (19
89)]. In addition, Dermatophagoides farina
The protease isolated from e), Df-protease, has been experimentally confirmed to have the ability to produce bradykinin, which exacerbates the inflammatory response and enhances vascular permeability [Biochem. Biophys. Act
a, 1074, 62 (1991)].

[0003] Comprehensively judging these facts, it is presumed that mite protease promotes the production of bradykinin, resulting in various mite allergic diseases. Therefore, suppressing the mite protease activity is considered to be useful for the treatment and / or prevention of mite allergic diseases (such as asthma, allergic rhinitis, and atopic dermatitis caused by mites).

[0004] As substances inhibiting tick proteases, Japanese Patent Application Laid-Open No. 6-192085 describes, for example, aprotinin, potato protease inhibitor, soybean trypsin inhibitor, antipain, leupeptin, guanidino fatty acid derivatives, guanidinobenzoic acid derivatives and amidinophenol derivatives. Is disclosed. JP-A-9-40631 discloses a therapeutic agent for mite allergy containing an amidinophenol derivative as an active ingredient.

[0005] However, aprotinin, potato protease inhibitor, soybean trypsin inhibitor,
Antipine and leupeptin are high-molecular-weight polypeptides and are not suitable for mass production because of their difficulty in chemical synthesis. In addition, amidinophenol derivatives use toxic solvents such as methylene chloride, chloroform, and methanol in their production, and are not satisfactory in terms of safety due to by-products generated during compounding and their toxicity.

[0006]

An object of the present invention is to provide a remedy for mite allergy which is safe and can be mass-produced.

[0007]

Means for Solving the Problems The present inventor has conducted intensive studies on a tick protease activity inhibitory substance which causes mite allergic disease. As a result, tea extracts, more specifically polyphenols contained in tea extracts, were found. Have solved the above-mentioned problem, and have completed the present invention. That is, the present invention is a therapeutic agent for mite allergy, which comprises a tea extract.

[0008]

BEST MODE FOR CARRYING OUT THE INVENTION Tea in the present invention is not particularly limited, but includes a camellia plant. The tea extract of the present invention is not particularly limited, and includes, for example, extracts such as green tea which is unfermented tea, oolong tea which is semi-fermented tea, and black tea which is fermented tea. It is preferable to use an extract of green tea, which is unfermented tea, from the viewpoint of a strong inhibitory activity against the green tea. It is more preferable to use polyphenols contained in the green tea extract. In addition, the said polyphenol is (-)-epicatechin, (-)-epigallocatechin, (-)-epicatechin gallate and (-)-epigallocatechin gallate,
Generic term for compounds such as myricetin, quercetin, rutin and kaempferol.

The extract fraction of tea is obtained by using tea leaves or powdered tea leaves with water, hot or hot water, organic solvents such as ethanol, propanol, methyl ethyl ketone, acetone, ethyl acetate, butanol and methyl isobutyl ketone alone or It is a fraction containing polyphenols extracted in combination. More preferably, a fraction containing polyphenols fractionated with an organic solvent is purified by column chromatography (-)-epicatechin, (-)-epigallocatechin, (-)-epicatechin gallate and (-)-. These are polyphenols such as epigallocatechin gallate, myricetin, quercetin, rutin and kaempferol.

[0010] The mite allergy of the present invention is an allergy caused by a protease derived from a tick, and is an allergic symptom caused by the mite protease causing an inflammatory reaction and enhancing vascular permeability.

Here, the mite protease is not particularly limited in the kind of the mite, but may be Dermatophagoides falina.
arinae), Dermatophagoides pteronasinas
nyssinus) and Dermatophagoides pteronaisinas (Dermatophagoides p.
Df- such as house dust mites such as teronyssinus)
Protease.

[0012] The therapeutic agent for mite allergy of the present invention is usually
It is used systemically or locally, by oral or transdermal administration. The dosage is age, body weight, symptoms, therapeutic effect, administration method,
Although it varies depending on the treatment time, etc., it is usually 1 mg to 1000 mg, preferably 50 mg to 5 mg per dose per adult.
Oral administration in the range of 00 mg once to several times a day, or an ointment containing 0.1 to 20% per adult is applied to the affected area once to several times a day.

The form for oral administration of the therapeutic agent for mite allergy according to the present invention is not particularly limited, but when the therapeutic agent is solid, it can be provided in the form of tablets, powders, granules and the like. In such a dosage form, although not particularly limited, (-)-epicatechin, (-)-epigallocatechin, (-)-epicatechin gallate (-)
-Epigallocatechin gallate, containing at least one or more polyphenols selected from the group consisting of myricetin, quercetin, rutin and kaempferol, and optionally an inert diluent such as lactose, mannitol,
Examples thereof include one or a mixture of two or more of glucose, hydroxypropyl cellulose, microcrystalline cellulose, starch, polyvinylpyrrolidone, and magnesium aluminate metasilicate. In addition, according to a conventional method, additives other than an inert diluent, for example, a lubricant such as magnesium stearate or a disintegrant such as calcium cellulose glycolate, a stabilizer such as lactose, glutamic acid or aspartic acid Such a solubilizing agent may be contained. If necessary, tablets or pills may be coated with a film of a gastric or enteric substance such as sucrose, gelatin, hydroxypropylcellulose, hydroxypropylmethylcellulose phthalate, or may be coated with two or more layers. . Further, the capsule may be made of an absorbable substance such as gelatin.

When the therapeutic agent is in liquid form, it can be provided as a pharmaceutically acceptable emulsion, solution, suspension, syrup, elixir and the like. Such dosage forms may include, but are not limited to, commonly used inert diluents, such as purified water and ethanol. Also,
In addition to inert diluents, wetting agents, auxiliary agents such as suspending agents,
Sweetening agents, flavoring agents, fragrances and preservatives may be contained. Other dosage forms for oral administration include sprays and the like.

[0015] The dosage form of the skin external preparation of the agent for treating mite allergy according to the present invention includes lotions, emulsions, gels,
It can be provided in creams, ointments, etc., and further, skin cosmetics such as lotions, emulsions, creams, packs, makeup base lotions, makeup base creams,
Liquid or creamy or ointment type foundation,
It can also be provided as makeup cosmetics such as eye color and cheek color, and body cosmetics such as hand cream, leg cream and body lotion.

Further, in addition to the above essential components, if necessary, various components which are usually used in a skin external preparation composition can be added as long as the effects of the present invention are not impaired. For example, oils, water, surfactants and humectants, alcohols, thickeners, antioxidants, sequestering agents, p
H adjusters, preservatives, fragrances, dyes, ultraviolet absorbers, ultraviolet scattering agents, vitamins, amino acids, and the like can be added.

Hereinafter, the present invention will be described in more detail with reference to Examples and Test Examples, but the present invention is not limited to these Examples and Test Examples.

[0018]

Example 1 About 1 liter of water was added to 1 kg of commercially available green tea, and the mixture was stirred.
Extracted at 0 ° C. for 3 hours. The extract obtained by filtration was concentrated to dryness to obtain 350 g of a hot water extract of green tea (38% pure as polyphenols).

Example 2 To 350 g of the hot water extract obtained in Example 1, 8 liters of water was added and dissolved, and the mixture was sequentially partitioned with hexane and chloroform. 10 L of ethyl acetate was added to the aqueous layer after the distribution, and the mixture was vigorously stirred and allowed to stand. The ethyl acetate layer was separated, ethyl acetate was distilled off, and the residue was dried to obtain 70 g of ethyl acetate-soluble fraction (as polyphenols). Purity 74.5%). The proportion of each polyphenol in the ethyl acetate-soluble fraction was (+)-catechin 3.5%, (+)-gallocatechin 14.8%,
(-)-Gallocatechin gallate 11.6% (-)-epicatechin 7%, (-)-epigallocatechin 15%,
4.6% of (-)-epicatechin gallate and (-)-
Epigallocatechin gallate was 18%.

Example 3 10 g of the ethyl acetate-soluble fraction obtained in Example 2 was subjected to silica gel column chromatography (solvent, chloroform:
Methyl alcohol, 20: 1, 10: 1, v / v), Sephadex LH-20 column chromatography (solvent, methyl alcohol), recycling HPLC (Nippon Bunka Kogyo LC-908, GS-320 column, solvent) By sequentially using (methyl alcohol), 0.3 g of (+)-catechin, 1.22 g of (+)-gallocatetekin, 0.9 g of (−)-gallocatechin gallate,
0.5 g of (-)-epicatechin, 0.38 g of (-)-epicatechin gallate, 1.2 of (-)-epigallocatechin 1.2
g and (-) epigallocatechin gallate 1.5 g of polyphenols were obtained.

Example 4 3 kg of hexane was added to 1 kg of commercially available dry green tea to perform defatting. The defatted tea leaves were extracted with 5 liters of acetone, and the obtained acetone layer was concentrated to dryness, and a fraction containing myricetin, quercetin, rutin, and kaempferol 2.
5 g were obtained.

Example 5 The fraction containing myricetin, quercetin, rutin and kaempferol obtained in Example 4 was subjected to recycling HPLC.
(LC-908, GS-320 column manufactured by Nippon Bunka Kogyo,
Using solvent methyl alcohol), 0.2 g of myricetin, 0.5 g of quercetin, 0.1 g of rutin, and 0.15 g of kaempferol were obtained.

Test Example 1 Inhibitory effect on house dust mite protease House dust mite protease (Df-protease) was replaced with house dust mite (Dermatophagoides farina).
e) from the culture extract of Int. Arch. Aller
gy Appl. Immunol. , 91, 80 (19
90) Purified according to the method described. The purified enzyme [150 units / mg (37 ° C, pH 8)] was stored at -80 ° C in a phosphate buffer solution (PBS) (1 mg / ml). The stored enzyme solution was diluted with cold 1 mM hydrochloric acid before use. The activity of the enzyme was determined using a synthetic substrate (Boc-G
ln-Gly-Arg-MCA) at 37 ° C. using the method of Kawabata et al. [Eur. J. Biochem. , 17
2, 17 (1988)]. A substrate (0 to 50 μM, dissolved in dimethyl sulfoxide) in 50 mM Tris-HCl buffer (pH 8.0, 1880 μl) containing various compounds (0 to 0.2 mM)
100 μl) was added. After addition of an enzyme (1.1 nM, 20 μl) dissolved in 1 mM hydrochloric acid, the initial value of the hydrolysis was measured with a fluorescence spectrophotometer, excited at 380 nm, emitted at 460 nm. The inhibitory effect of each compound on Df-protease was evaluated by changing the molar ratio of each compound to Df-protease. That is, a mixture of Df-protease and each compound was cultured at 37 ° C. for 1 to 3 minutes (the culture time was varied depending on each compound), and the remaining enzyme activity was measured under the same conditions as above. Ki
The value was determined from the Lineweaver-Burk plot.

The results are shown in Table 1.

[0025]

[Table 1]

As is clear from Table 1, the hot water extract of green tea obtained in Example 1, the ethyl acetate fraction obtained in Example 2, the (+)-catechin obtained in Example 3, +)-Gallocatechin, (-)-gallocatechin gallate, (-)
-Epicatechin, (-)-epicatechin gallate,
(-)-Epigallocatechin, and (-)-epicatechin, (-)-of (-) epigallocatechin gallate
Epigallocatechin, (−)-epicatechin gallate and (−)-epigallocatechin gallate, the fraction containing myricetin, quercetin, rutin and kaempferol obtained in Example 4, and the myricetin obtained in Example 5 , Quercetin, rutin and kaempferol are Df-
It had an inhibitory effect on protease.

Test Example 2 Effect of Atopic Dermatitis Model Mouse Male atopic dermatitis model mouse due to mites (NC / N
ga mouse: Japanese SLC) was tested for its inhibitory effect on the lesion of atopic dermatitis due to tick allergy. This mouse is an atopic dermatitis model mouse in which the symptoms of atopic dermatitis develop when a tick infests the host. Example 1 in a model mouse (3 weeks old, n = 10)
The substances obtained in 2, 3, 4, and 5 were dissolved in drinking water to a concentration of 0.05% (Examples 4 and 5 were pre-dissolved in ethanol) and given as drinking water, and bred up to 15 weeks. Was done. At 16 weeks of age, the average of pruritus scratching behavior (number of times / 10 minutes), the number of injured sites, and changes in blood IgE concentration were observed for 5 days at the age of 16 weeks.

The results are shown in Table 2.

[0029]

[Table 2]

As is clear from Table 2, the green tea hot water extract obtained in Example 1, the ethyl acetate fraction obtained in Example 2, the (+)-catechin obtained in Example 3, +)-Gallocatechin, (-)-gallocatechin gallate, (-)
-Epicatechin, (-)-epicatechin gallate,
(-)-Epigallocatechin, and (-)-epicatechin, (-)-of (-) epigallocatechin gallate
Epigallocatechin, (−)-epicatechin gallate and (−)-epigallocatechin gallate, the fraction containing myricetin, quercetin, rutin and kaempferol obtained in Example 4, and the myricetin obtained in Example 5 , Quercetin, rutin and kaempferol were found to have an effect of suppressing allergic symptoms in atopic dermatitis model mice.

The embodiments of the present invention and the target products are as follows. (1) A therapeutic agent for mite allergy, comprising a tea extract. (2) The therapeutic agent for mite allergy according to (1), wherein the tea is green tea. (3) The therapeutic agent for mite allergy according to (1), wherein the tea extract is at least one selected from the group consisting of polyphenols. (4) Polyphenol is (+)-catechin, (+)-gallocatechin, (−)-gallocatechin gallate, (−)-
(3) which is at least one selected from the group consisting of epicatechin, (−)-epigallocatechin, (−)-epicatechin gallate (−)-epigallocatechin gallate, myricetin, quercetin, rutin and kaempferol The therapeutic agent for mite allergy according to the above. (5) Polyphenol is (-)-epicatechin, (-)
-The therapeutic agent for mite allergy according to (3), which is at least one selected from the group consisting of -epigallocatechin, (-)-epicatechin gallate (-)-epigallocatechin gallate, myricetin, quercetin, rutin, and kaempferol. . (6) A method for producing a therapeutic agent for mite allergy containing a mite protease inhibitor, comprising a tea extract. (7) A method for producing a therapeutic agent for mite allergy containing the tick protease inhibitor according to (1), wherein the tea is green tea. (8) An oral preparation containing the tick protease inhibitor according to any one of (1) to (5). (9) A skin external preparation containing the tick protease inhibitor according to any one of (1) to (5). (10) A therapeutic agent wherein the disease of mite allergy according to any of (1) to (5) is asthma. (11) A therapeutic agent, wherein the mite allergic disease according to any of (1) to (5) is allergic rhinitis. (12) A therapeutic agent, wherein the mite allergic disease according to any of (1) to (5) is atopic dermatitis.

[0032]

INDUSTRIAL APPLICABILITY The present invention has a strong inhibitory activity against mite proteases and an inhibitory effect on inflammation caused by mite proteases, and has very little toxicity. It is useful for the treatment and / or prevention of human mite allergic diseases, for example, asthma, allergic rhinitis, atopic dermatitis, etc. caused by mites.

 ────────────────────────────────────────────────── ─── Continuing on the front page (72) Inventor Mizuko Futami F-term (reference) 2-20-31, Urawa-shi, Saitama 4C086 AA01 AA02 BA08 GA17 MA01 MA02 MA03 MA04 NA14 ZB13 ZB39 ZC20 4C088 AB45 AC05 BA08 BA09 BA10 BA11 BA31 MA01 NA14 ZB13 ZB39 ZC20

Claims (4)

[Claims] 1. An agent for treating mite allergy, which comprises a tea extract. 2. The therapeutic agent for mite allergy according to claim 1, wherein the tea extract is a green tea extract. 3. The therapeutic agent for mite allergy according to claim 1, wherein the tea extract is a tea extract fraction containing polyphenols. 4. The group of polyphenols consisting of (-)-epicatechin, (-)-epigallocatechin, (-)-epicatechin gallate (-)-epigallocatechin gallate, myricetin, quercetin, rutin and kaempferol. 4. The therapeutic agent for mite allergy according to claim 3, which is at least one or more selected from the group consisting of: