JP6986375B2 - Anti-allergic agents, mediator release inhibitors and oral medications - Google Patents

Description

The present invention relates to an antiallergic agent containing an L-ascorbic acid derivative having an alkyl group or an isomer thereof as an active ingredient, a mediator release inhibitor, and an internal medicine having an antiallergic action.

In recent years, the number of people suffering from type I allergies such as pollinosis has been increasing. Type I allergy develops when mediators such as histamine are released by the degranulation reaction of mast cells.
Therefore, Patent Document 1 describes an antihistamine containing fexofenadine hydrochloride having a histamine H1 receptor antagonistic activity.
Further, Patent Document 2 describes an antihistamine action enhancer containing clemastine fumarate or chlorpheniramine d-maleate as an antihistamine.

Japanese Unexamined Patent Publication No. 2013-75892 Japanese Unexamined Patent Publication No. 2009-185058

Here, the histamine receptor is known to have a wakefulness effect and the like. As a result, taking the above-mentioned antihistamine that antagonizes the histamine receptor may cause drowsiness, malaise, thirst, etc. as side effects, so caution is required when taking it.
Therefore, there has been a demand for a safe anti-allergic agent with fewer side effects.

In view of the above circumstances, an object of the present invention is to provide an antiallergic agent having few side effects and high safety, a mediator release inhibitor, and an internal medicine having an antiallergic action.

In order to achieve the above object, the antiallergic agent according to one embodiment of the present invention contains at least one of compound (A) and compound (B) as an active ingredient.
The above compound (A) is
General formula (1)

（一般式（１）中、Ｒ１は炭素数４以上のアルキル基である）で表されるＬ−アスコルビン酸誘導体、その異性体、又は上記Ｌ−アスコルビン酸誘導体若しくはその異性体の薬学的に許容される塩である。
上記化合物（Ｂ）は、
一般式（２）

(In the general formula (1), R1 is an alkyl group having 4 or more carbon atoms) L-ascorbic acid derivative, an isomer thereof, or the above L-ascorbic acid derivative or an isomer thereof is pharmaceutically acceptable. It is the salt that is made.
The above compound (B) is
General formula (2)

（一般式（２）中、Ｒ２は炭素数１０以上１８以下のアルキル基である）で表されるＬ−アスコルビン酸誘導体又はその異性体である。

(In the general formula (2), R2 is an alkyl group having 10 or more and 18 or less carbon atoms), which is an L-ascorbic acid derivative or an isomer thereof.

Compounds (A) and (B) have an inflammatory mediator release inhibitory action. Therefore, it does not cause side effects such as drowsiness, malaise, and thirst as seen in drugs having an inflammatory mediator receptor antagonistic action. In addition, compounds (A) and (B) are derivatives of L-ascorbic acid, which has an antioxidant effect and is known as vitamin C. Therefore, it is possible to provide an antiallergic agent having few side effects and high safety.

R1 of the above general formula (1) may be an alkyl group having 16 or more carbon atoms.
Alternatively, R2 of the above general formula (2) may be an alkyl group having 12 or more and 16 or less carbon atoms.
This makes it possible to provide an antiallergic agent having a higher inflammatory mediator release inhibitory action.

Moreover, the said antiallergic agent may contain the said compound (A) and the said compound (B) as an active ingredient.
By using both the compounds (A) and (B) as active ingredients, it is possible to provide a highly safe anti-allergic agent having a high inflammatory mediator release inhibitory action and having few side effects.

The mediator release inhibitor according to still another embodiment of the present invention contains at least one of compound (A) and compound (B) as an active ingredient.
The above compound (A) is
General formula (1)

（一般式（１）中、Ｒ１は炭素数４以上のアルキル基である）で表されるＬ−アスコルビン酸誘導体、その異性体、又は上記Ｌ−アスコルビン酸誘導体若しくはその異性体の薬学的に許容される塩である。
上記化合物（Ｂ）は、
一般式（２）

(In the general formula (1), R1 is an alkyl group having 4 or more carbon atoms) L-ascorbic acid derivative, an isomer thereof, or the above L-ascorbic acid derivative or an isomer thereof is pharmaceutically acceptable. It is the salt that is made.
The above compound (B) is
General formula (2)

（一般式（２）中、Ｒ２は炭素数１０以上１８以下のアルキル基である）で表されるＬ−アスコルビン酸誘導体又はその異性体である。

(In the general formula (2), R2 is an alkyl group having 10 or more and 18 or less carbon atoms), which is an L-ascorbic acid derivative or an isomer thereof.

The oral medicine according to still another form of the present invention contains at least one of compound (A) and compound (B) as an active ingredient and has an antiallergic effect.
The above compound (A) is
General formula (1)

（一般式（１）中、Ｒ１は炭素数４以上のアルキル基である）で表されるＬ−アスコルビン酸誘導体、その異性体、又は上記Ｌ−アスコルビン酸誘導体若しくはその異性体の薬学的に許容される塩である。
上記化合物（Ｂ）は、
一般式（２）

(In the general formula (1), R1 is an alkyl group having 4 or more carbon atoms) L-ascorbic acid derivative, an isomer thereof, or the above L-ascorbic acid derivative or an isomer thereof is pharmaceutically acceptable. It is the salt that is made.
The above compound (B) is
General formula (2)

（一般式（２）中、Ｒ２は炭素数１０以上１８以下のアルキル基である）で表されるＬ−アスコルビン酸誘導体又はその異性体である。

(In the general formula (2), R2 is an alkyl group having 10 or more and 18 or less carbon atoms), which is an L-ascorbic acid derivative or an isomer thereof.

Compound (A) and compound (B) also have an antiallergic effect when taken orally. Therefore, it is possible to provide an internal medicine having an antiallergic action with few side effects and high safety.

As described above, according to the present invention, it is possible to provide an antiallergic agent, a mediator release inhibitor, and an internal medicine having an antiallergic action, which have few side effects and are highly safe.

3 is a graph showing the results of 2-O-alkylascorbic acid and L-ascorbic acid (Examples 1 to 5 and Comparative Example 3) in Test Example 1 of the present invention. 3 is a graph showing the results of 3-O-alkylascorbic acid (Examples 6 to 8 and Comparative Examples 1 and 2) in Test Example 1 of the present invention. It is a graph which shows the result of the test example 2 of this invention. It is a graph which shows the result of the test example 3 of this invention. It is a graph which shows the result of the test example 4 of this invention.

Hereinafter, the present invention will be described in detail.

<Outline of the present invention>
The antiallergic agent, mediator release inhibitor, and oral drug of the present invention contain at least one of compound (A) and compound (B), which are derivatives of L-ascorbic acid, as active ingredients. That is, the antiallergic agent, mediator release inhibitor and internal medicine of the present invention may contain compound (A) or compound (B) as an active ingredient, or both compound (A) and compound (B) may be used as active ingredients. good. As described later, the compound (A) and the compound (B) are both alkyl ascorbic acid or the like in which an alkyl chain is bonded to L-ascorbic acid.
L-ascorbic acid derivatives are generally unstable and easily lose their activity. As a result of diligent research, the present inventor has found alkylascorbic acid as an L-ascorbic acid derivative having high stability, activity not lost even when ingested, and high mediator release inhibitory action, and came up with the present invention. did.

<Compound A>
The compound (A) of the present invention is an L-ascorbic acid derivative represented by the general formula (1), an isomer thereof, or a pharmaceutically acceptable salt of the above-mentioned ascorbic acid derivative or an isomer thereof.

一般式（１）中、Ｒ１は炭素数４以上のアルキル基である。すなわち、一般式（１）で表されるＬ−アスコルビン酸誘導体は、２−Ｏ−アルキルアスコルビン酸である。

In the general formula (1), R1 is an alkyl group having 4 or more carbon atoms. That is, the L-ascorbic acid derivative represented by the general formula (1) is 2-O-alkylascorbic acid.

<2-O-alkylascorbic acid>
The 2-O-alkylascorbic acid of the present invention is a compound having an alkyl group R1 at the 2-position of L-ascorbic acid. The alkyl group R1 has 4 or more carbon atoms, preferably 16 or more, and for example, 16 or more and 18 or less. The alkyl group R1 may have a linear structure or a branched structure, but is preferably linear. Specific examples of the 2-O-alkylascorbic acid of the present invention include 2-O-butyl ascorbic acid, 2-O-octyl ascorbic acid, 2-O-dodecyl ascorbic acid, 2-O-hexadecyl ascorbic acid, 2 -O-octadecyl ascorbic acid can be mentioned.

<Isomer of 2-O-alkylascorbic acid>
The isomer of 2-O-alkylascorbic acid of the present invention is a compound in which an alkyl chain is bonded to the 2-position of the isomer of L-ascorbic acid. Examples of the isomer of L-ascorbic acid include D-ascorbic acid, L-alaboascorbic acid, and D-alaboascorbic acid (also referred to as erythorbic acid and isoascorbic acid). Of these, erythorbic acid is preferably used in view of marketability and the like.
These isomers are known to have the same function as L-ascorbic acid (see Suzuki et al., J. Nutr. Sci. Vitaminol., 41, 17-24, (1995), etc.). In the invention, it can be used in the same manner as L-ascorbic acid.

<Salt of 2-O-alkylascorbic acid or its isomer>
The pharmaceutically acceptable salt of 2-O-alkylascorbic acid or an isomer thereof of the present invention typically has a structure in which the 3-position of L-ascorbic acid is an anion.
As the salt of 2-O-alkylascorbic acid or an isomer thereof of the present invention, one having particularly low toxicity is preferable. The inorganic bases that produce these salts include alkali metals (eg, sodium, potassium, etc.), ammonium, alkaline earth metals (eg, calcium, magnesium, strontium, barium, etc.), aluminum salts, and the like, and the organic bases include, for example. Examples thereof include one or a mixture of two or more selected from trimethylamine, triethylamine, pyridine, picoline, N, N-dibenzylethylenediamine, ethanolamine, diethanolamine, trishydroxymethylaminomethane, dicyclohexylamine and the like. Among these, those selected from one or a mixture of two or more selected from sodium, potassium, magnesium, calcium and aluminum are particularly easy to apply.

<Compound B>
The compound (B) of the present invention is an L-ascorbic acid derivative represented by the general formula (2) or an isomer thereof.

一般式（２）中、Ｒ２は炭素数１０以上１８以下のアルキル基である。すなわち、一般式（２）で表されるＬ−アスコルビン酸誘導体は、３−Ｏ−アルキルアスコルビン酸である。
また、３−Ｏ−アルキルアスコルビン酸の異性体は、２−Ｏ−アルキルアスコルビン酸の異性体と同様に、Ｌ−アスコルビン酸の異性体の３位にアルキル鎖が結合されている化合物である。Ｌ−アスコルビン酸の異性体としては、上述のＤ−アスコルビン酸、Ｌ−アラボアスコルビン酸、Ｄ−アラボアスコルビン酸を用いることができる。

In the general formula (2), R2 is an alkyl group having 10 or more and 18 or less carbon atoms. That is, the L-ascorbic acid derivative represented by the general formula (2) is 3-O-alkylascorbic acid.
Further, the isomer of 3-O-alkylascorbic acid is a compound in which an alkyl chain is bonded to the 3-position of the isomer of L-ascorbic acid, similarly to the isomer of 2-O-alkylascorbic acid. As the isomer of L-ascorbic acid, the above-mentioned D-ascorbic acid, L-alaboascorbic acid, and D-alaboascorbic acid can be used.

<3-O-alkylascorbic acid>
The 3-O-alkylascorbic acid of the present invention is a compound having an alkyl group R2 at the 3-position of L-ascorbic acid. The number of carbon atoms of the alkyl group R2 is 10 or more and 18 or less, preferably 12 or more and 16 or less. The alkyl group R2 may have a linear structure or a branched structure, but is preferably linear. Specific examples of the 3-O-alkylascorbic acid of the present invention include 3-O-dodecyl ascorbic acid, 3-O-hexadecyl ascorbic acid, and 3-O-octadecyl ascorbic acid. As described below, 3-O-dodecyl ascorbic acid has a very high inflammatory mediator release inhibitory effect.

<Manufacturing method of alkyl ascorbic acid derivative>
The method for producing alkylascorbic acid of the present invention can be various methods.
The synthesis of 2-O-alkylascorbic acid is described, for example, by Kato et al., "Studies on scavengers of active oxygen species. 1.synthesis and biological activity of 2-O-alkylascorbic acids.", Journal of Medicinal Chemistry, 31, 793. --You can refer to 798 (1988).
As an example, 5,6-isopropyridene-ascorbic acid in which the hydroxyl group at the 5th and 6th positions of L-ascorbic acid is protected with an isopropylidene group is prepared, and the 3-position of 5,6-isopropyridene-ascorbic acid is a protecting group. Protect. Examples of this protecting group include a methoxymethyl group (MOM group) and a benzyl group. Subsequently, an alkylating agent is reacted with the ascorbic acid derivative in which the 5, 6 and 3 positions are protected to perform alkylation. Examples of the alkylating agent include alkyl halides and alkylbenzene sulfonates. Finally, the alkylated L-ascorbic acid derivative can be deprotected to synthesize 2-O-alkylascorbic acid.

For the synthesis of 3-O-alkylascorbic acid, see, for example, Tai et al., "A simple efficient synthesis and biological evaluation of 3-O-ethylascorbic acid." Bioscience Biotechnology and Biochemistry, 12, 1984-1987 (2014). be able to.
As an example, 3-O-alkylascorbic acid can be synthesized by reacting L-ascorbic acid salt with an alkylating agent. Examples of the L-ascorbic acid salt include sodium L-ascorbate. Examples of the alkylating agent include alkyl halides and alkylbenzene sulfonates. As another example, 3-O is obtained by reacting 5,6-isopropyridene-ascorbic acid in which the hydroxyl group at the 5,6 position of L-ascorbic acid is protected with an isopropylidene group with an alkylating agent to deprotect it. -Alkyl ascorbic acid can also be produced.

The alkylascorbic acid of the present invention obtained by these methods can be purified by an appropriate method in consideration of physical characteristics. Examples of the purification method include salting out, dialysis, filtration, concentration, fractional precipitation, liquid separation extraction, gel chromatography, ion exchange chromatography, high-speed liquid chromatography, gas chromatography, affinity chromatography, gel electrophoresis, isoelectric focusing, etc. Crystallization and the like can be mentioned. These may be used in combination as appropriate.

<Mediator release inhibitor>
The mediator release inhibitor of the present invention contains at least one of the above compound (A) and the above compound (B) as an active ingredient, and can inhibit the release of an inflammatory mediator such as histamine that causes type I allergy. .. As described above, type I allergy is caused by the release of mediators such as histamine by the degranulation reaction of mast cells. As described later, the ascorbic acid derivative can suppress the degranulation reaction of mast cells and can exert a mediator release inhibitory action.

<Anti-allergic agent>
The antiallergic agent and mediator release inhibitor of the present invention contain at least one of the above compound (A) and the above compound (B) as an active ingredient, and can be applied to allergies, particularly type I allergies. Specifically, the antiallergic agent can suppress symptoms such as rhinitis, conjunctivitis, bronchial asthma, food allergy, anaphylaxis, urticaria or atopic dermatitis. The allergen for allergies to which the antiallergic agent is applied is not particularly limited, and may be, for example, pollen, mites, animal fluff, mold, chemical substances, proteins in food, and the like.

<Pharmaceuticals, quasi-drugs>
The antiallergic agent and mediator release inhibitor of the present invention can be used in various dosage forms, for example, as pharmaceuticals or quasi-drugs, depending on the intended use. The administration form may be either oral administration or parenteral administration, and oral administration is preferable. This makes it possible to provide a highly safe anti-allergic agent and the like, which can be administered with low invasiveness. The term "drug" as used herein includes not only human drugs but also veterinary drugs.

The dosage form of the pharmaceutical product or non-pharmaceutical product of the present invention is not particularly limited, but is not limited to liquids (including drinks), syrups, granules, powders, pills, tablets, capsules, troches, chewables, ointments. , Eye drops, nasal drops, inhalants, capsules, patches, injections and the like.
The antiallergic agent and mediator release inhibitor of the present invention can be formulated with an active ingredient such as alkylascorbic acid or a salt thereof according to a known pharmaceutical production method (17th revised Japanese Pharmacopoeia). (See Notification No. 64 of the Ministry of Health, Labor and Welfare on March 7, 2016).
In this case, the antiallergic agent and the mediator release inhibitor include, in addition to the above active ingredients, pharmaceutically acceptable bases, carriers, other medicinal ingredients, additives and the like within the range in which the action of the present invention is not reduced. Can be mixed. The carrier can be selected by referring to the Japanese Pharmacopoeia, "Pharmaceutical Additives Dictionary" (published by Yakuji Nippo Co., Ltd.), or the like. Examples of the additive include stabilizers, dispersants, fluidizers, buffers, lubricants, preservatives, pH adjusters, solvents, solubilizers and the like, which are acceptable as additives for ordinary pharmaceutical products.

<Internal medicine>
At least one of the compound (A) and the compound (B) of the present invention can be an active ingredient of an internal medicine having an antiallergic action. The term "internal medicine" as used herein refers to an internal powder, granules, tablets, capsules, pills, internal liquids, syrups and the like. Thereby, the antiallergic agent and the mediator release inhibitor of the present invention can be orally ingested as pharmaceuticals.
In the oral medicine of the present invention, the dose of the above-mentioned alkyl ascorbic acid can be, for example, 0.1 mg or more and 100 mg or less per day per 1 kg of body weight. The frequency of administration is not particularly limited, but may be about 1 to 6 times a day.

<Beverages and foods>
The antiallergic agent and mediator release inhibitor of the present invention can also be prepared, for example, as beverages and foods. The beverages and foods referred to herein include beverages and foods for humans, as well as animal feeds and pet foods.
Beverages and foods are not particularly limited, but for example, beverages (eg, dairy beverages, lactic acid bacteria beverages, soft beverages containing fruit juice, carbonated beverages, fruit juice beverages, vegetable beverages, vegetable / fruit beverages, alcoholic beverages, sports beverages, sports beverages, etc. , Powdered beverages, tea beverages, etc.), Cold confectionery (eg jelly, bavarois, pudding, etc.), Iced confectionery (eg ice cream, ice milk, lacto ice, sherbet), Confectionery (eg cookies, biscuits, okaki, candy, etc.) Chocolates, gums), breads, noodles (eg Chinese noodles, pasta, udon, soba, plain noodles), soups (including powder or solid soup), seasonings (eg dressing, jelly, sauce, mayonnaise) Source, drink) and the like.
Further, the beverages and foods include functional foods containing supplements (foods for nutritional function, foods for specified health use, etc.), foods for the sick (including foods for people requiring nursing care and foods for people who have difficulty swallowing), and the like. ..
These beverages and foods can be produced by combining known ingredients and materials, or by adding additives and the like that are acceptable in the field of beverages and foods.

<Cosmetics>
The antiallergic agent and mediator release inhibitor of the present invention may be used as a component of cosmetics or quasi-drug medicated cosmetics. Specific examples of such cosmetics include creams, emulsions, lotions, facial masks, and bathing agents for sensitive or allergic skin. The above cosmetics include oily or aqueous bases, vitamins, emollients, whitening agents, moisturizers, antioxidants, buffers, UV absorbers, chelating agents, pH regulators, preservatives, thickeners, alcohols. , Cooling agents, colorants, fragrances and the like can be added as needed.

<Action and effect of the present invention>
The antiallergic agent and mediator release inhibitor of the present invention can effectively inhibit the release of an inflammatory mediator such as histamine. Therefore, side effects such as drowsiness, fatigue, and thirst caused by antihistamines having a mediator receptor antagonistic action can be avoided. Alkyl ascorbic acid is a derivative of L-ascorbic acid, which has an antioxidant effect and is known as vitamin C. Thereby, the present invention can provide a highly safe anti-allergic agent with few side effects.
Furthermore, the antiallergic agent and mediator release inhibitor of the present invention have a high inflammatory mediator release inhibitory effect even by oral administration. This enables less invasive administration and can be provided in a form that is easy for patients to ingest.

Hereinafter, the present invention will be further described based on examples and the like. L-ascorbic acid is abbreviated as "AA", 2-O-alkylascorbic acid is abbreviated as "2-alkylAA", and 3-O-alkylascorbic acid is abbreviated as "3-alkylAA". Other compounds shall be abbreviated as appropriate.

[Example 1: 2-Butyl AA]
As Example 1, an AA derivative having a butyl group having 4 carbon atoms bonded to the 2-position was prepared. This derivative is 2-O-butylascorbic acid, and is hereinafter referred to as 2-butylAA (2-butylAA).
For 2-butyl AA, see Kato et al., "Studies on scavengers of active oxygen species. 1.synthesis and biological activity of 2-O-alkylascorbic acids.", Journal of Medicinal Chemistry, 31, 793-798 (1988). Then, it was manufactured as follows.

(Synthesis of 5,6-isopropyridene AA)
AA was dissolved in acetone (Wako Pure Chemical Industries, Ltd.), acetyl chloride (Wako Pure Chemical Industries, Ltd.) was added, and the mixture was stirred at 40 ° C. for 1 hour. Then, by filtration, washing and drying, 5,6-isopropilideneAA (5,6-isopropilideneAA) was synthesized.

(Synthesis of 5,6-isopropylidene-3-methoxymethyl AA)
Dissolve 5,6-isopropyridene AA in tetrahydrofuran (THF), N, N-dimethylformamide (DMF) (Nacalai Tesque, Inc.), add potassium carbonate (Wako Pure Chemical Industries, Ltd.), and add at 30 ° C for 10 minutes. Stirred. After stirring, chloromethyl methyl ether (Wako Pure Chemical Industries, Ltd.) was added, and the mixture was stirred at 30 ° C. for 3 hours. After stirring, water was added to stop the reaction, the reaction solution was neutralized with 1 M hydrogen chloride (Wako Pure Chemical Industries, Ltd.), and the reaction solution was separated with ethyl acetate (Wako Pure Chemical Industries, Ltd.). The ethyl acetate layer was washed with water, dehydrated and concentrated to obtain 5,6-isopropilidene-3-MOMAA as a reaction product.

(Butylization)
5,6-Isopropylidene-3-methoxymethyl AA was dissolved in THF, DMSO (Dimethyl sulfoxide) (Wako Pure Chemical Industries, Ltd.), potassium carbonate, 1-iodobutane as an alkylating agent. (Tokyo Chemical Industry Co., Ltd.) was added, and the mixture was stirred at 50 ° C. for 2 hours. After stirring, water was added to stop the reaction, the reaction solution was neutralized with 1 M hydrogen chloride, and the solution was separated with ethyl acetate. The ethyl acetate layer was washed, dehydrated and concentrated, and the reaction product was wet-filled in a Wakogel® C-200 column (Wako Pure Chemical Industries, Ltd.) and subjected to hexane (Wako Pure Chemical Industries, Ltd.)-ethyl acetate solvent. Eluted by. As a result, 5,6-isopropilidene-3-MOM-2-butylAA (5,6-isopropilidene-3-MOM-2-butylAA) was obtained.

(Deprotection and purification)
5,6-Isopropylidene-3-methoxymethyl-2-butylAA was dissolved in ethanol, 1 M hydrogen chloride was added, and the mixture was stirred at 80 ° C. for 1 hour. After stirring, the reaction solution was concentrated. The concentrated reaction product was subjected to a DIAION (registered trademark) HP20 column (Mitsubishi Chemical Corporation), and the elution fraction of 2-butyl AA eluted with methanol (Wako Pure Chemical Industries, Ltd.) -water solvent was concentrated and the residue was obtained. Got
The obtained residue was recrystallized from a mixed solution of diisopropyl ether (IPE) (Wako Pure Chemical Industries, Ltd.) and ethyl acetate to obtain 2-butyl AA. The obtained substance was analyzed by nuclear magnetic resonance spectroscopy, high resolution mass spectrometry (HRMS), etc., and confirmed to be 2-butyl AA.

[Example 2: 2-octyl AA]
As Example 2, an AA derivative having an octyl group having 8 carbon atoms bonded to the 2-position was prepared. This AA derivative is 2-O-octyl ascorbic acid, and is hereinafter referred to as 2-octylAA (2-octylAA).
2-octyl AA was produced in the same manner as in Example 1. However, 1-iodooctane (Tokyo Chemical Industry Co., Ltd.) was used as the alkylating agent. A Wakogel® C-200 column and a hexane-ethyl acetate-methanol solvent were used to purify 2-octylAA. The obtained substance was analyzed in the same manner as in Example 1 and confirmed to be 2-octylAA.

[Example 3: 2-Dodecyl AA]
As Example 3, an AA derivative having a dodecyl group having 12 carbon atoms bonded to the 2-position was prepared. This derivative is 2-O-dodecyl ascorbic acid, and is hereinafter referred to as 2-dodecylAA (2-dodecylAA).
2-Dodecyl AA was produced in the same manner as in Example 1. However, as the alkylating agent, 1-iodododecane (Tokyo Chemical Industry Co., Ltd.) was used. The purification of 2-dodecylAA was carried out by separating the solution with ethyl acetate instead of elution by a column, washing the ethyl acetate layer with water, dehydrating and concentrating, and recrystallizing the obtained residue. The obtained substance was analyzed in the same manner as in Example 1 and confirmed to be 2-dodecylAA.

[Example 4: 2-hexadecylAA]
As Example 4, an AA derivative having a hexadecyl group having 16 carbon atoms bonded to the 2-position was prepared. This derivative is 2-O-hexadecyl ascorbic acid, and is hereinafter referred to as 2-hexadecylAA (2-hexadecylAA).
2-Hexadecyl AA was produced in the same manner as in Example 3. However, 1-iodohexadecane (Tokyo Chemical Industry Co., Ltd.) was used as the alkylating agent. The obtained substance was analyzed in the same manner as in Example 1 and confirmed to be 2-hexadecylAA.

[Example 5: 2-octadecylAA]
As Example 5, an AA derivative having an octadecyl group having 18 carbon atoms bonded to the 2-position was prepared. This derivative is 2-O-octadecyl ascorbic acid, and is hereinafter referred to as 2-octadecylAA (2-octadecylAA).
2-Octadecyl AA was produced in the same manner as in Example 3. However, 1-iodooctadecane (Tokyo Chemical Industry Co., Ltd.) was used as the alkylating agent. The obtained substance was analyzed in the same manner as in Example 1 and confirmed to be 2-octadecylAA.

[Comparative Example 1: 3-Butyl AA]
As Comparative Example 1, an AA derivative having a butyl group having 4 carbon atoms bonded to the 3-position was prepared. This derivative is 3-O-butylascorbic acid, and is hereinafter referred to as 3-butylAA (3-butylAA).
3-Butyl AA was produced as follows with reference to Tai et al., "A simple efficient synthesis and biological evaluation of 3-O-ethylascorbic acid." Bioscience Biotechnology and Biochemistry, 12, 1984-1987 (2014). ..
Sodium L-ascorbic acid (Wako Pure Chemical Industries, Ltd.) was dissolved in DMSO and stirred at 50 ° C. for 10 minutes. After stirring, 1-butane iodide as an alkylating agent was added, the mixture was stirred at 50 ° C. for 3 hours, and water was added to stop the reaction. The reaction was directly applied to a DIAION® HP20 column without concentration and eluted with a methanol-aqueous solvent to concentrate the elution fraction of 3-butyl AA. The residue was applied to a wet-filled Wakogel® C-200 column and eluted with a toluene-acetone solvent. The eluted fraction of 3-butyl AA was concentrated to give 3-butyl AA. The obtained substance was analyzed in the same manner as in Example 1 and confirmed to be 3-butyl AA.

[Comparative Example 2: 3-Octil AA]
As Comparative Example 2, an AA derivative having an octyl group having 8 carbon atoms bonded to the 3-position was prepared. This derivative is 3-O-octyl ascorbic acid, and is hereinafter referred to as 3-octylAA (3-octylAA).
3-octyl AA was produced as follows according to Comparative Example 1.
Sodium L-ascorbic acid was dissolved in DMSO and stirred at 50 ° C. for 10 minutes. After stirring, 1-octane iodide as an alkylating agent was added, the mixture was stirred at 50 ° C. for 3 hours, and water was added to stop the reaction. The reaction mixture was separated by ethyl acetate, and the ethyl acetate layer was washed with water, dehydrated and concentrated. The reaction was applied to a wet-filled Wakogel® C-200 column and eluted with a toluene-acetone solvent. The eluted fraction of 3-octylAA was concentrated to give 3-octylAA. The obtained substance was analyzed in the same manner as in Example 1 and confirmed to be 3-octylAA.

[Example 6: 3-Dodecyl AA]
As Example 6, an AA derivative having a dodecyl group having 12 carbon atoms bonded to the 3-position was prepared. This derivative is 3-O-dodecyl ascorbic acid, and is hereinafter referred to as 3-dodecylAA (3-dodecylAA).
3-Dodecyl AA was produced in the same manner as in Comparative Example 2. However, as the alkylating agent, 1-dodecane iodide was used. Further, 3-dodecylAA was obtained by concentrating the eluted fraction of 3-dodecylAA eluted using a Wakogel® C-200 column and then recrystallizing the residue by IPE. The obtained substance was analyzed in the same manner as in Example 1 and confirmed to be 3-dodecylAA.

[Example 7: 3-HexadecylAA]
As Example 7, an AA derivative having a hexadecyl group having 16 carbon atoms bonded to the 3-position was prepared. This derivative is 3-O-hexadecyl ascorbic acid, and is hereinafter referred to as 3-hexadecylAA (3-hexadecylAA).
3-Hexadecyl AA was produced in the same manner as in Example 6. However, 1-hexadecane iodide was used as the alkylating agent. The obtained substance was analyzed in the same manner as in Example 1 and confirmed to be 3-hexadecylAA.

[Example 8: 3-Octadecyl AA]
As Example 8, an AA derivative having an octadecyl group having 18 carbon atoms bonded to the 3-position was prepared. This derivative is 3-O-octadecyl ascorbic acid, and is hereinafter referred to as 3-octadecylAA (3-octadecylAA).
3-Octadecyl AA was produced in the same manner as in Example 6. However, 1-octadecane iodide was used as the alkylating agent. The obtained substance was analyzed in the same manner as in Example 1 and confirmed to be 3-octadecylAA.

[Comparative Example 3: AA]
AA was prepared as Comparative Example 3. AA purchased the product name L-sodium ascorbic acid from Wako Pure Chemical Industries, Ltd.

The following Test Example 1, Test Example 2 and Test Example 3 were carried out using the compounds of Examples 1 to 8 and Comparative Examples 1 to 3.

<Test Example 1: Antigen-stimulated degranulation suppression test (Examples 1 to 8, Comparative Examples 1 to 3)>
First, in order to confirm the action of the compounds according to Examples 1 to 8 and Comparative Examples 1 to 3 on the degranulation of mast cells causing the symptoms of type I allergy, an antigen using rat basophil leukemia cell RBL-2H3 was used. A stimulating degranulation suppression test was performed.

(Cells used)
The rat basophil leukemia cells (RBL-2H3) used in this test example were purchased from Human Science Cell Bank (JCRB). All cells (RBL-2H3) used in the experiment were cultured in DMEM medium containing 10% FBS (inactivated) under the conditions of _{37 ° C and 5% CO 2, and subcultured every 2-4 days.} .. FBS (Fetal bovine serum) (lot.No.42k9155k) was purchased from HyClone. The DMEM medium was purchased from Sigma-Aldrich Japan, and 100 U / ml penicillin and 100 μg / ml streptomycin (both from Nacalai Tesque Co., Ltd.) were added and used.

(Adjustment of reagents)
The antibody solution was prepared by adding mouse monoclonal anti-DNP IgE antibody (Sigma-Aldrich Japan) to Dulbecco's phosphate buffered saline (DPBS) (PBS (-)) (Sigma-Aldrich Japan) to a concentration of 250 μg / ml. rice field. It was then stored at -30 ° C and diluted to 500 ng / ml in DMEM medium containing 10% FBS at the time of use.
MT buffer (Modified Tyrode buffer) is made by adding 137 mM sodium chloride (Wako Pure Chemical Industries, Ltd.), 2.7 mM potassium chloride (Nakarai Tesk Co., Ltd.), 1.8 mM calcium chloride (Wako Pure Chemical Industries, Ltd.) to ultra-pure water. , 1 mM magnesium chloride hexahydrate (Wako Pure Chemical Industries, Ltd.), 5.6 mM (+)-glucose (Wako Pure Chemical Industries, Ltd.), 20 mM 2- [4- (2-hydroxyethyl) -1-piperazinyl ] ethanesulfonic acid (HEPES) (Wako Pure Chemical Industries, Ltd.) and 0.1% bovine serum albumin (BSA) (Nakarai Tesk Co., Ltd.) are added and dissolved, and pH 7.3 with sodium hydroxide (Wako Pure Chemical Industries, Ltd.) Prepared for. The prepared MT buffer was stored at 4 ° C and warmed to 37 ° C at the time of use.
The antigen solution was prepared by adding Dinitrophenyl (DNP) -labeled human serum albumin (HAS) (Sigma-Aldrich Japan) to DPBS (-) to a concentration of 10 mg / ml. The prepared antigen solution was stored at 4 ° C. and diluted with the above MT buffer to 500 ng / ml (final concentration 50 ng / ml) at the time of use.
At the time of use, the 0.1% Triton solution was prepared by diluting Triton X-100 (Sigma-Aldrich Japan) with MT buffer to 0.1%.
When using the substrate solution, add 3.3 mM p-nitrophenyl-2-acetamido-2-deoxy-β-D-glucopyranoside (PNAG) (Wako Pure Chemical Industries, Ltd.) to 100 mM citric acid buffer (pH 4.5). Prepared.
The enzyme reaction terminator (2 M Glycine buffer) was prepared by adding a 2 M glycine solution (Wako Pure Chemical Industries, Ltd.) to sodium hydroxide to a pH of 10.4.
The compounds of Examples 1 to 8 and Comparative Examples 1 and 2 were all prepared to a predetermined concentration by MT buffer containing 0.28% DMSO. Examples 1 to 8 and Comparative Examples 1 and 2 thus prepared are referred to as samples in this test example, respectively.
In addition, as a negative control, an MT buffer containing 0.28% DMSO to which no compound was added was used (indicated as “control” in FIG. 1).
As a positive control, 75 μM oxatomide (Wako Pure Chemical Industries, Ltd.) was used. Oxatomid is known as an H1 receptor blocker and an inflammatory mediator release inhibitor.

(Measurement of degranulation inhibitory activity)
First, the cells used, which had been cultured until semiconfluent, were stripped with 0.05% trypsin-0.02% EDTA and placed on a 96-well flat-bottomed multiplate (NUNC, 167008) at ^{a density of 5.0 x 10 4} cells / well / 200 μl. Sown. After culturing the cells for 24 hours, the culture supernatant was removed, 100 μl of the antibody solution was added to each well, and the cells were incubated for _{2 hours at 37 ° C. and 5% CO 2 to sensitize the cells.} ..
After incubation, the cell layer was washed twice with MT buffer, 90 μl of sample solution and positive control were added to each well, and the cells were incubated for 1 hour at 37 ° C. and 5% CO ₂ .
Subsequently, 10 μl of the antigen solution was added to each well, and after incubating for 1 hour, the reaction was stopped by ice-cooling for 10 minutes, and the supernatant was collected. This supernatant was dispensed into 96-well microplates (“WATSON®”, Fukae Kasei Co., Ltd.) for measurement in 20 μl increments. 100 μl of 0.1% Triton solution was added to the dispensed supernatant, and the cells were disrupted by stirring (500 rpm, 5 minutes). 20 μl of this cell disruption solution was dispensed into a 96-well microplate for measurement.
Subsequently, 40 μl of the substrate solution was added to each well, and the reaction was carried out for 90 minutes under the condition of 37 ° C. After the reaction, the enzyme reaction was stopped with 40 μl of the enzyme reaction stop solution, the color was developed, and the absorbance at 405 nm was measured (product name “MULTISKAN FC”, Thermo Fisher Scientific Co., Ltd.). The β-hexosaminidase release rate (degranulation rate) was calculated by the following formula.
Degranulation rate (%)
= [(SX-SY) / {(SX-SY) + (CLX-CLY)}] x 100-[(BSX-BSY) / {(BSX-BSY) + (BCLX-BCLY)}] x 100
S: Amount of β-hexosaminidase released in the supernatant
CL: β-hexosaminidase release amount of cell disruption solution
X: When the substrate solution and the enzyme reaction stop solution are added in this order
Y: When the enzyme reaction stop solution and the substrate solution are added in this order
B: When using an antibody solution and a cell layer to which no sample was added Y was used to subtract the absorbance of the sample itself.

(Statistical processing)
Quantitative data are shown as mean +/- standard deviation.
The comparison of the mean values between multiple groups was performed by Dunnett's test after one-way ANOVA, and it was judged that there was a significant difference with a risk rate of less than 0.05.

(result)
FIG. 1 is a graph showing the results of 2-alkylAA and AA (Examples 1 to 5 and Comparative Example 3) in this test example, where the horizontal axis shows the sample and the vertical axis shows the degranulation ratio. .. In each sample, the results of concentrations of 50, 75, and 100 μM are shown in order from the left. The numerical values in the bar graph indicate the average value, and each error bar indicates the standard deviation.

As shown in FIG. 1, in all of Examples 1 to 5, a decrease in degranulation was observed in a concentration-dependent manner, and it was confirmed that each compound according to Examples 1 to 5 had a degranulation inhibitory effect. ..
In Examples 1 to 5, the higher the carbon number of the alkyl group, the higher the degranulation inhibitory effect. In particular, the samples of Example 4 (2-hexadecylAA) and Example 5 (2-octadecylAA) showed significant activity as compared with the negative control. As a result, it was confirmed that a high degranulation inhibitory effect can be exhibited by having 16 or more carbon atoms in the alkyl group at the 2-position.
Furthermore, the sample of Example 5 (2-octadecylAA) exhibited the same activity as the positive control oxatomido. From this, it was confirmed that the 2-alkylAA of the present invention has a sufficiently high degranulation inhibitory effect.
On the other hand, in Comparative Example 3, no degranulation inhibitory effect was observed. Therefore, it is considered that the degranulation inhibitory action according to Examples 1 to 5 is not due to the metabolite AA, but to the alkyl ascorbic acid itself of the present invention to which an alkyl group is added.

FIG. 2 is a graph showing the results of 3-alkylAA (Examples 6 to 8 and Comparative Examples 1 and 2) in this test example, in which the horizontal axis shows the sample and the vertical axis shows the degranulation ratio. .. In each sample, the results of concentrations of 50, 75, and 100 μM are shown in order from the left. The numerical values in the bar graph indicate the average value, and each error bar indicates the standard deviation.
As shown in the figure, in all of Examples 6 to 8, a decrease in degranulation was observed as compared with the negative control, and each compound according to Examples 6 to 8 had a degranulation inhibitory effect. Was confirmed.
In particular, in Example 6 (3-dodecylAA), both concentrations showed higher activity than the positive control oxatomido. From this, it was confirmed that the 3-alkylAA of the present invention to which an alkyl group having about 12 carbon atoms was added has a very high degranulation inhibitory effect.
On the other hand, in Comparative Examples 1 and 2, no degranulation inhibitory activity was observed. From this, it was confirmed that 3-alkylAA having 8 or less carbon atoms does not have a degranulation inhibitory effect.

In this test example, rat basophil leukemia cells (RBL-2H3) were used as a model of mast cells, and the β-hexosaminidase release rate was evaluated as the degranulation rate. As a chemical mediator, the cells also contain histamine in addition to β-hexosaminidase. In the same cells, the amount of β-hexosaminidase released is in parallel with the amount of histamine released, so it is considered that the amount of β-hexosaminidase released can be accurately evaluated as an index of degranulation. (See Passante N et al., "The RBP-2H3 cell line: its provenance and suitability as a model for the mast cell.", Inflammation Research, 58, 737-745 (2009)).

<Test Example 2: Calcium ionophore stimulation-induced degranulation suppression test (Examples 4, 5, 6, 7>
Subsequently, in order to investigate the degranulation mechanism of the samples of Examples 4 to 7 in which a particularly high degranulation inhibitory effect was observed in Test Example 1, a calcium ionophore stimulation-induced degranulation inhibition test was conducted.
It is known that degranulation is caused by the activation of signal molecules in the cell membrane by antigen stimulation, which ^{promotes the} influx of Ca 2+ into the cell, resulting in an ^{increase in the intracellular Ca 2+} concentration. ing. Calcium ionophore (A23187) has the ^{effect of enhancing the transport of Ca 2+ in} the cell membrane and allowing extracellular Ca ²⁺ to flow into the cell.
Therefore, in this test example, it was examined whether or not the samples of Examples 4 to 7 suppress calcium ionophore stimulation-induced degranulation.

(Preparation of cells used and reagents)
As the cells used, the same cells as in Test Example 1 were used.
Of the reagents, the calcium ionophore solution was prepared by dissolving calcium ionophore A23187 (Sigma-Aldrich Japan) in DMSO to 50 mM. It was stored at -30 ° C and diluted with MT buffer to 100 μM (final concentration 10 μM) before use.
The other reagents used in this test example were prepared in the same manner as in test example 1.

(Measurement and statistical processing of degranulation inhibitory activity by calcium ionophore stimulation)
Cells detached with 0.05% trypsin-0.02% EDTA solution were seeded on a 96-well flat-bottomed multiplate (NUNC, 167008) at ^{a density of 5.0 x 10 4} cells / well / 200 μl. After culturing the cells for 24 hours, the cell layer was washed twice with MT buffer, 90 μl of sample solution and positive control were added to each well, and then incubated for _{20 minutes under the conditions of 37 ° C. and 5% CO 2.} ..
To each well, 10 μl of calcium ionophore solution was added and incubated for 1 hour under the conditions of _{37 ° C. and 5% CO 2 to induce degranulation.} The plate was then ice-cooled for 10 minutes to stop the reaction. The supernatant was collected and dispensed into a 96-well microplate (“WATSON®”, Fukae Kasei Co., Ltd.) for measurement in 20 μl increments.
On the other hand, to the cell layer, 100 μl of 0.1% Triton solution was added, and the mixture was lysed by stirring (500 rpm, 5 minutes). 20 μl of this cell disruption solution was dispensed into a 96-well microplate for measurement.
Subsequently, 40 μl of the substrate solution was added to each well, and the reaction was carried out for 90 minutes under the condition of 37 ° C. After the reaction, the enzyme reaction was stopped with 40 μl of an enzyme reaction terminator (glycine buffer), the color was developed, and the absorbance at 405 nm was measured (product name “MULTISKAN FC”, Thermo Fisher Scientific Co., Ltd.). The β-hexosaminidase release rate (degranulation rate) was calculated by the same formula as in Test Example 1.
Statistical processing was performed in the same manner as in Test Example 1.

(result)
FIG. 3 is a graph showing the results in this test example, in which the horizontal axis shows the sample and the vertical axis shows the degranulation ratio. In each sample, the results of concentrations of 25, 50, 75, and 100 μM are shown in order from the left. The numerical values in the bar graph indicate the average value, and each error bar indicates the standard deviation.

As shown in the figure, all of the samples of Examples 4 to 7 suppressed calcium ionophore stimulation-induced degranulation. In particular, in the samples of Examples 4 to 6, that is, 2-hexadecyl AA, 2-octadecyl AA, and 3-dodecyl AA, a high degranulation inhibitory effect equal to or higher than that of the positive control oxatomid was observed. Of these, 3-dodecylAA of Example 6 showed a higher degranulation inhibitory effect than oxatomid, and was confirmed to have extremely high activity.

<Test Example 3: Degranulation suppression test by co-addition of Example 5 (2-octadecylAA) and Example 6 (3-dodecylAA)>
In order to further investigate the mechanism of suppressing degranulation of alkyl AA, the present inventor ^{measured the intracellular Ca 2+} dynamics at the time of antigen stimulation when alkyl AA was added over time. ^{As a result, an increase in Ca 2+} concentration was observed when 2-alkyl AA was added, ^{but an increase in Ca 2+} concentration was inhibited when 3-alkyl AA was added (not shown).
^{That is, 2-alkylAA} increases the intracellular Ca 2+ concentration at the time of antigen stimulation, but can suppress degranulation. On the other hand, 3-alkylAA suppresses degranulation without increasing ^{the intracellular Ca 2+ concentration during antigen stimulation.} This suggests that 2-alkylAA and 3-alkylAA have different mechanisms for suppressing degranulation. In that case, when these are co-added, it is presumed that an additive effect is observed.
Therefore, the present inventor co-added 2-octadecylAA of Example 5 and 3-dodecylAA of Example 6 and evaluated whether or not an additive effect was observed in suppressing degranulation by antigen stimulation.

As the cells used, the same cells as in Test Example 1 were used.
The reagents were also prepared in the same manner as in Test Example 1.
The measurement and statistical processing of the degranulation inhibitory activity were also carried out in the same manner as in Test Example 1.

(result)
FIG. 4 is a graph showing the results in this test example, in which the horizontal axis shows the sample and the vertical axis shows the degranulation ratio. In each sample, the results of the concentration of 12.5, 25, 50, 75, 100, 150 μM are shown in order from the left.
As shown in the figure, when 25 μM of Example 5 (2-octadecylAA) and Example 6 (3-dodecylAA) were added alone, about 10 in Example 5 (2-octadecylAA). %, Example 6 (3-dodecylAA) showed an inhibition rate of about 25%, whereas co-addition showed an inhibition rate of about 40%. Furthermore, when 50 μM was added, Example 5 (2-octadecylAA) showed a degranulation inhibition rate of about 24%, and Example 6 (3-dodecylAA) showed a degranulation inhibition rate of about 60%, whereas when co-added, about It showed an inhibition rate of 81%.
From this result, it was confirmed that the co-addition of Example 5 (2-octadecylAA) and Example 6 (3-dodecylAA) added the degranulation inhibitory effect.
In the co-addition, no concentration-dependent decrease was observed at concentrations of 100 and 150 μM, but a significant decrease in the degranulation rate was observed in both cases.

<Test Example 4: Passive cutaneous anaphylaxis test (PCA) (Examples 5 and 6)>
From the results of Test Examples 1 to 3, it was confirmed that the alkyl AA of the present invention has an antiallergic effect in vitro.
Therefore, in order to evaluate whether the alkyl AA of the present invention has an anti-allergic effect in vivo and maintains its activity even when ingested orally, Example 5 (2-octadecyl AA) and Example 6 Mouse PCA using (3-dodecylAA) was performed.
The PCA reaction is a reaction in which a blue dye is exposed at a site where an antibody is intradermally administered when an antibody is intradermally administered and then a corresponding antigen and a dye are intravenously administered, which is a kind of local anaphylaxis. The PCA reaction is considered to be a reaction that occurs when mast cells bound to an antibody are stimulated by an antigen and release mediators such as histamine to increase vascular permeability, and is used as a type I allergy model. There is.

(Adjustment of reagents)
The 0.9% sodium chloride aqueous solution was prepared by dissolving sodium chloride in ultrapure water so as to be 0.9%, and filtering and sterilizing it with a 0.22 μm filter.
For the antibody solution, mouse monoclonal anti-DNP IgE antibody was added to DPBS (-) and dissolved to 250 μg / ml. Then, it was stored at -30 ° C and diluted with 0.9% aqueous sodium chloride solution to 5 μg / ml at the time of use.
The DNP-HAS solution was diluted with a 0.9% aqueous sodium chloride solution (light-shielded after dilution) and mixed with the Evans blue solution to prepare a final concentration of 0.4 mg / ml.
The Evans blue solution was prepared by diluting Evans blue with a 0.9% aqueous sodium chloride solution and mixing with the DNP-HAS solution to a final concentration of 1%.
In the 1 M potassium hydroxide aqueous solution, potassium hydroxide was dissolved in ultrapure water so as to be 1 M.
In the 0.4 M phosphoric acid aqueous solution, phosphoric acid was dissolved in ultrapure water so as to be 0.4 M.
Acetone-0.4 M phosphoric acid mixed solution was prepared by mixing acetone and 0.4 M phosphoric acid aqueous solution at a ratio of 13: 5 (v / v).
The compounds of Examples 5 and 6 were both suspended in 5% ethanol / DPBS so as to have an oral dose of 100,200 μmol / kg and mixed so as to be uniform. Examples 5 and 6 thus prepared are referred to as samples in this test example, respectively.
As a positive control, oxatomide was suspended in ethanol / DPBS to an oral dose of 100 μmol / kg and mixed uniformly.

(Animal used)
JcI: ICR male mice (7 weeks old) (purchased from Japan SLC) were used. They were bred in a breeding room with a constant temperature and humidity, and solid feed (CE-2) was given at 5 g / mouse per day, and tap water was allowed to be freely ingested. After preliminary breeding for about one week, it was used for the experiment.

(Measurement of Evans blue amount in PCA reaction)
20 μl of 0.9% sodium chloride aqueous solution or anti-DNP-IgE antibody was subcutaneously administered to the auricles of ICR male mice under anesthesia. After 24 hours, under anesthesia, 5% ethanol / DPBS was applied to the negative control group, sample solution was applied to the 2-octadecyl AA (Example 5) group and 3-dodecyl AA (Example 6) group, and positive control (Example 6). The oxatomide group was orally administered with an oxatomide solution at 100 or 200 μmol / kg bw, respectively. The number of mice in each group was 5 or 6. Two hours later, 0.25 ml of DNP-HAS solution was administered to the tail vein. Thirty minutes later, the patient was euthanized by dislocation of the cervical spine, and the amount of Evans blue leaking into the skin of both ears was measured. The auricle after euthanasia was cut and immersed in 500 μl of 1 M potassium hydroxide aqueous solution and dissolved overnight (37 ° C). Then, 3.75 ml of a mixed solution of acetone-0.4 M phosphoric acid was added, and the mixture was stirred and then centrifuged (1000 xg, 20 min), and the absorbance of the supernatant at 620 nm was measured.

(result)
FIG. 5 is a graph showing the results of this test example, in which the horizontal axis represents a sample and the vertical axis represents an Evans blue amount [%]. The amount of Evans blue indicates the ratio of the absorbance of each group when the absorbance of Evans blue in the negative control is 100%. In each sample, the results of 100, 200 μmol / kg bw are shown in order from the left. For oxatomide, the results are shown at 100 μmol / kg bw. The numerical values in the bar graph indicate the average value, and each error bar indicates the standard deviation.
As shown in the figure, both Examples 5 and 6 showed significant inhibitory activity in a concentration-dependent manner. Further, even when compared with oxatomido, which is an antiallergic agent used as a positive control, the above-mentioned alkyl AA was confirmed to have the same level of inhibitory activity at a concentration about twice that of oxatomid.
From the above results, it was confirmed that the alkyl AA of the present invention has an anti-allergic effect even by oral administration.

<Summary>
From the results of Test Examples 1 to 4, it was confirmed that the alkyl AA according to Examples 1 to 8 of the present invention has a mediator release inhibitory action as an antiallergic action.

From the results of Test Example 1, it was confirmed that 2-alkylAA had a mediator release inhibitory action in all the samples, but it was confirmed that the longer the length of the alkyl group, the higher the mediator release inhibitory action. Was done. In particular, it was confirmed that when the alkyl group has 16 or more carbon atoms, it has a high mediator release inhibitory effect.
Further, 3-alkylAA was found to have a high mediator release inhibitory effect on 3-dodecyl AA having 12 carbon atoms and hexadecyl AA having 16 carbon atoms, and the same effect was also observed on octadecyl AA having 18 carbon atoms. From this, it was confirmed that 3-alkylAA has high activity in a configuration having an alkyl group having 12 or more and 16 or less carbon atoms, and if it has an alkyl group having 10 or more and 18 or less carbon atoms, it has a mediator release inhibitory action. Have.

From the results of Test Example 3, it was confirmed that the additive action was generated by using both 2-alkylAA and 3-alkylAA. In the same test example, 2-octadecylAA was used as 2-alkylAA and 3-dodecylAA was used as 3-alkylAA. In view of the results of Test Example 1, not limited to this example, both 2-alkylAA having an alkyl group having 4 or more carbon atoms and 3-alkylAA having an alkyl group having 10 or more and 18 or less carbon atoms are effective. When used as an ingredient, an antiallergic agent having a mediator release inhibitory action can be provided.

From the results of Test Example 4, it was confirmed that 2-octadecylAA and 3-dodecylAA suppress the PCA reaction by oral administration. In view of the results of Test Example 1, 2-alkylAA having an alkyl group having 4 or more carbon atoms and 3-alkylAA having an alkyl group having 10 or more and 18 or less carbon atoms are both type I by oral administration. It is presumed to have the effect of suppressing allergies. Thereby, the alkyl AA of the present invention can be used as an active ingredient of an internal medicine having an anti-allergic action and a mediator release inhibitory action.

Claims (6)

An antiallergic agent containing at least one of compound (A) and compound (B) as an active ingredient.
The compound (A) is
General formula (1)

(In the general formula (1), R ₁ is an alkyl group having 4 or more carbon atoms) L-ascorbic acid derivative, an isomer thereof, or the L-ascorbic acid derivative or an isomer thereof pharmaceutically It is an acceptable salt, and the isomer in the description of the general formula (1) is 2-O-alkyl-D-ascorbic acid, 2-O-alkyl-L-alaboascorbic acid or 2-O-alkyl. -D-means alaboscorbic acid,
The compound (B) is
General formula (2)

(In the general formula (2), R ₂ is an alkyl group having 10 to 18 carbon atoms) said in explanation of Ri L- ascorbic acid derivative or an isomer der represented by the general formula (2) isomer, 3-O-alkyl -D- ascorbic acid, 3-O-alkyl -L- araboascorbic acid or 3-O-alkyl -D- antiallergic agent you mean araboascorbic acid. The antiallergic agent according to claim 1.
_{R 1} of the general formula (1) is an antiallergic agent which is an alkyl group having 16 or more carbon atoms. The antiallergic agent according to claim 1 or 2.
_{R 2} of the general formula (2) is an antiallergic agent which is an alkyl group having 12 or more and 16 or less carbon atoms. The antiallergic agent according to any one of claims 1 to 3.
An antiallergic agent containing the compound (A) and the compound (B) as an active ingredient. A mediator release inhibitor containing at least one of compound (A) and compound (B) as an active ingredient.
The compound (A) is
General formula (1)

(In the general formula (1), R ₁ is an alkyl group having 4 or more carbon atoms) L-ascorbic acid derivative, an isomer thereof, or the L-ascorbic acid derivative or an isomer thereof pharmaceutically It is an acceptable salt, and the isomer in the description of the general formula (1) is 2-O-alkyl-D-ascorbic acid, 2-O-alkyl-L-alaboascorbic acid or 2-O-alkyl. -D-means alaboscorbic acid,
The compound (B) is
General formula (2)

(In the general formula (2), R ₂ is an alkyl group having 10 to 18 carbon atoms) said in explanation of Ri L- ascorbic acid derivative or an isomer der represented by the general formula (2) isomer, 3-O-alkyl -D- ascorbic acid, 3-O-alkyl -L- araboascorbic acid or 3-O-alkyl -D- mediator release inhibitors you mean araboascorbic acid. The antiallergic agent according to any one of claims 1 to 4, which is an internal medicine.

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