KR101702897B1 - Composition for preventing or treating gout comprising caffeic acid conjugated with biotin - Google Patents

Abstract

The present invention relates to a pharmaceutical composition and a health food for preventing or treating gout containing an active ingredient of catechin phosphate (BTCA) compound having biotin binding thereto. More particularly, the present invention relates to a pharmaceutical composition for prevention and treatment of gout comprising catechophosphoric acid (BTCA) Significantly reduced the level of increase of cytokines induced by urate (MSU) crystals, and in particular inhibited the production of IL-1 [beta] and inhibited NLRP3 inflammasome activity, leading to induction by urate crystals And inhibited the inflammatory response. Accordingly, the BTCA compound of the present invention can be used as a pharmaceutical composition and a health food for the prevention or treatment of gout caused by inflammation induced by MSU crystals.

Description

TECHNICAL FIELD [0001] The present invention relates to a composition for preventing or treating gout, which contains, as an active ingredient, a caffeic acid compound having a biotin-

The present invention relates to a pharmaceutical composition for preventing or treating gout and a health food containing a compound (BTCA) in which caffeic acid is conjugated with biotin as an active ingredient.

Gout is a metabolic disorder in which the crystals of uric acid formed by persistent high uric acid in the blood can not be discharged out of the body and accumulate in various tissues to cause various symptoms. Increased serum uric acid may occur due to excessive production of uric acid and failure to properly discharge due to abnormal excretion through the kidneys.

High uric acid in the blood does not cause gout immediately, but uric acid crystals are easily formed by high uric acid, and when the state of being deposited in various tissues persists for 10 to 20 years, it is caused by induction factors. It is commonly found in men aged 40 to 50, but the age at onset is gradually decreasing with changes in diet and environment. Women are women who have had a long-term use of immunosuppressants after menopause or after organ transplantation, The possibility of developing the disease is high.

The cause of this gout is that most of the cells that make up our body have nuclei, which are made up of nucleic acids that contain genetic information. Nucleic acid is composed of purine or pyrimidine. When the cell reaches its end of life, it breaks down and the purine body in the nucleus is decomposed to produce a large amount of uric acid. The crystals of sodium urate precipitate in the tissues and cause inflammation. Hyperuricemia, gouty, Stomatitis and the like.

Currently, these gout treatments mainly use colchicine, non-steroidal anti-inflammatory drugs or steroids, but no medicines have been developed to radically treat gout. However, allopurinol, an inhibitor of uric acid uric acid production, and probenecid, a uric acid kidney excretion promoter, are known. However, allopurinol has serious adverse effects such as skin rash, gastrointestinal disorder, bone marrow suppression, itching and muscle pain Therefore, there is a demand for the development of a therapeutic agent capable of preventing or treating gout while being safe to the human body and reducing side effects.

Korean Patent Publication No. 10-2010-0064519 (published on June 15, 2010)

As described above, the present invention relates to a composition for treating gout induced by uric acid crystals deposited on the body, and a therapeutic drug which has been used in the past has a problem of excessive side effects, To prevent or treat gout, and to provide a pharmaceutical composition or a health food.

The present invention provides a pharmaceutical composition for prevention or treatment of gout, which comprises a compound (BTCA) represented by the following formula (1), which is bound to caffeic acid and biotin, as an active ingredient.

[Chemical Formula 1]

The present invention also provides a health food for prevention or improvement of gout comprising caffeic acid and biotin combined and containing the compound represented by the formula (1) as an active ingredient.

According to the present invention, the biotin-conjugated caffeic acid compound (BTCA) is a cytokine derived from MSU crystals in a gout animal model in which the release of inflammatory mediators by monosodium urate monohydrate (hereinafter "MSU" Cine increased significantly, and in particular inhibited the inflammatory response induced by MSU crystals by inhibiting the production of IL-1β induced by NLRP3 inflammasome.

Accordingly, the biotin-conjugated caffeic acid compound (BTCA) of the present invention can be used as a pharmaceutical composition and health food for preventing or treating gout induced by MSU crystals.

FIG. 1 shows results of ELISA analysis in which an inhibitory effect of each compound on IL-1β secretion amount induced by MSU crystals after oral administration of each compound bound with biotin to a gout model induced by MSU crystal was confirmed.

The present invention provides a pharmaceutical composition for prevention or treatment of gout comprising, as an active ingredient, a compound (BTCA) represented by the following formula (1), wherein caffeic acid and biotin are combined:

[Chemical Formula 1]

In Formula 1,

R ¹ or R ² may be the same or different and may be any one of OH, NH _2, and SH, and the solid line with the dotted line represents a single bond or a double bond.

The ventilation of the present invention may be caused by monosodium urate monohydrate (MSU) crystals, and more particularly, it may be induced by deposition of MSU crystals on the body, but is not limited thereto.

The compound may inhibit the production of IL-1? Activated by urate (MSU) crystals.

According to one embodiment of the present invention, C57BL / 6 mice local inflammation animal model (Murine air-pouch model) biotin coupled to urate (MSU) making the ventilation animal model induced by injecting crystal suspension, and caffeic acid in (BTCA) was orally administered to animal models, it was confirmed that IL-1 beta secretion, which is an inflammatory reaction substance induced by uric acid (MSU) crystals, is suppressed. Accordingly, the composition containing BTCA, which is a biotin-conjugated compound of caffeic acid according to the present invention, as an active ingredient is useful as a pharmaceutical composition for preventing or treating gout by inhibiting the inflammatory reaction induced by urate (MSU) .

The pharmaceutical composition of the present invention may be contained in an amount of 0.01 to 90 parts by weight based on 100 parts by weight of the total amount of the pharmaceutical composition, but is not limited thereto.

In one embodiment of the present invention, the pharmaceutical composition for prevention or treatment of gout containing the BTCA compound as an active ingredient may be administered orally or parenterally in the form of injections, granules, powders, tablets, pills, capsules, suppositories, Any one of the formulations selected from the group consisting of emulsions, drops, and liquids can be used.

In another embodiment of the present invention, the pharmaceutical composition for prevention or treatment of gout comprising the BTCA compound as an active ingredient can be administered orally or parenterally in association with a suitable carrier, excipient, disintegrant, sweetener, coating agent, swelling agent, The lubricant may further comprise at least one additive selected from the group consisting of a lubricant, a flavoring agent, an antioxidant, a buffer, a bacteriostatic agent, a diluent, a dispersant, a surfactant, a binder and a lubricant.

Specific examples of carriers, excipients and diluents include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, Cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. Solid formulations for oral administration may be in the form of tablets, pills, powders, granules, capsules These solid preparations can be prepared by mixing at least one excipient, for example, starch, calcium carbonate, sucrose or lactose, gelatin, etc., into the composition. In addition to simple excipients, lubricants such as magnesium stearate and talc may also be used. Examples of the liquid preparation for oral use include suspensions, solutions, emulsions, syrups and the like, and various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included in addition to water and liquid paraffin which are commonly used simple diluents. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, suppositories, and the like. Examples of the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. As the suppository base, witepsol, macrogol, tween 61, cacao paper, laurin, glycerogelatin and the like can be used.

According to one embodiment of the present invention, the pharmaceutical composition may be administered orally, intraarterally, intraperitoneally, intramuscularly, intraarterally, intraperitoneally, intrasternally, transdermally, nasally, inhaled, topically, rectally, ≪ / RTI > can be administered to the subject in a conventional manner.

The preferred dose of the BTCA compound may vary depending on the condition and body weight of the subject, the type and degree of the disease, the drug form, the administration route and the period, and may be appropriately selected by those skilled in the art. According to one embodiment of the present invention, the daily dose may be 0.01 to 200 mg / kg, specifically 0.1 to 200 mg / kg, more specifically 0.1 to 100 mg / kg, though it is not limited thereto. The administration may be performed once a day or divided into several times, and thus the scope of the present invention is not limited thereto.

In the present invention, the 'subject' may be a mammal including a human, but is not limited thereto.

The present invention also provides a health food for prevention or improvement of gout comprising a compound represented by the following formula (1) as an active ingredient, wherein caffeic acid and biotin are combined:

[Chemical Formula 1]

In Formula 1,

R ¹ or R ² may be the same or different and may be any one of OH, NH _2, and SH, and the solid line with the dotted line represents a single bond or a double bond.

The health food may be provided in the form of powder, granules, tablets, capsules, syrups or beverages. The health food may contain other food or food additives (such as food or drink) in addition to the biotin-conjugated caffeic acid (BTCA) And can be suitably used according to a conventional method. The amount of the active ingredient to be mixed can be suitably determined according to its use purpose, for example, prevention, health or therapeutic treatment.

The effective dose of the BTCA compound contained in the health food may be used in accordance with the effective dose of the pharmaceutical composition, but may be less than the above range for health and hygiene purposes or long-term intake for health control purposes , It is clear that the active ingredient can be used in an amount exceeding the above range since there is no problem in terms of safety.

There is no particular limitation on the type of the health food, and examples thereof include meat, sausage, bread, chocolate, candy, snack, confectionery, pizza, ramen, other noodles, gums, dairy products including ice cream, Drinks, alcoholic beverages and vitamin complexes.

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, the present invention will be described in detail with reference to the following examples. However, the following examples are intended to illustrate the contents of the present invention, but the scope of the present invention is not limited to the following examples. Embodiments of the present invention are provided to more fully describe the present invention to those skilled in the art.

The following reference example is provided to provide a reference example commonly applied to each embodiment according to the present invention.

<Referential Example>

1. Animals

The mice used in the present invention were purchased from Orient Bio (Seoul, Korea), and the management and experimental protocols of the purchased animals were performed according to the guidelines of the Institutional Animal Care and Use Committee (IACUC) of the Catholic University of Korea (permission) # 2012-5-001).

The mice were adapted to sterile conditions for at least a week before the experiment and maintained a temperature of 23 ± 3 ° C and 40-60% relative humidity.

2. Reagents

LPS purified from E. coli was obtained from List Biological Laboratory Inc (Hornby, Canada) and dissolved in the toxin solution. MSU was purchased from Invivogen (San Diego, USA), and interleukin-1β (IL-1β) ELISA kit was purchased from R & D Systems (Minneapolis, MN, USA).

< Example  1> Biotin Combined  Caffeine phosphate compound synthesis

Biotin-tagged dimethoxycinnamic acid (BTDMC), biotin-tagged caffeic acid (BTCA), and biotin-conjugated dihydroxyhydrocinnamic acid (Biotin-tagged dihydroxyhydrocinnamic acid ; BTDHHC) was synthesized by the following procedure.

1. N- Biotinyl - Hexane diamine  (N- Biotinoyl - hexanediamine ) synthesis

1.5 mmol of N, N'-carbonyldiimidazole (CDI) was added to 5 ml of dimethylformamide in which 1 mmol of biotin had been dissolved. After 30 minutes, 1.5 mmol of N-Boc-hexanediamine was added thereto at room temperature for 30 minutes Stirred and evaporated.

The residue after evaporation was washed with 0.1 M HCl, 10% -NaHCO ₃ solution and water, and reacted with ₃ ml of 3M HCl-containing acetic acid for 1 hour to obtain N-biotinyl-hexanediamine.

2. Biotin Combined Dimethoxycinnamic acid  (Biotin-tagged dimethoxycinnamic  acid; BTDMC ) synthesis

2.25 mmol of CDI was added to dimethylformamide in which 1.5 mmol of dimethoxycinnamic acid had been dissolved. After 30 minutes, a mixture of 1 mmol of N-biotinyl-hexanediamine and 2 mmol of triethylamine was added thereto at room temperature After stirring for 1 day, it was evaporated.

The remaining residue after evaporation was washed with 0.1 M HCl, 10% -NaHCO ₃ solution and water and dried in a vacuum oven to obtain biotin-bound dimethoxycinnamic acid as a white powder.

Total yield: 56%; mp .: 222-224 [deg.] C; IR (nujol mull):? _Max (cm ^-1 ) = 1703 (C = O, H ₂ NCONH ₂ ), 1670 (C = O, amide), 1637 (C = O, amide); ^{1 H-NMR (DMSO-d6} ): δ = 1.25-1.59 (m, 14H), 2.22 (t, 2H, J = 7.3 Hz) 2.55 (d, 1H, J = 12.5 Hz) 2.80 (dd, 1H, J 2H), 3.76 (s, 3H), 3.77 (s, 3H), 4.10 (m, 6.8 (d, 1H, J = 13.9 Hz), 6.96 (d, 1H, J = 8.3 Hz), 7.08 (dd, 1H , J = 8.3 Hz, 1.6 Hz ), 7.12 (d, 1H, J = 1.7 Hz), 7.32 (d, 1H, J = 15.6 Hz)

3. Biotin Combined  Caffeic acid ( BTCA ) And Biotin Combined Dihydroxyhydrocinnamic acid (BTDHHC) synthesis

α-Dichlorodiphenylmethane (1.5 mmol) was added to 20 ml of diphenyl ether in which 1 mmol of caffeophosphoric acid or 1 mmol of dihydroxyhydrocinnamic acid had been dissolved and dissolved. The reaction mixture was stirred at 175 ° C. for 30 minutes After cooling to room temperature, 40 ml of 0.1 M NaOH was added and washed three times with diethyl ether.

The aqueous layer was acidified with 1 M HCl to obtain catechol-protected caffeic acid and dihydroxyhydrocinnamic acid white precipitate.

After 2.25 mmol of N, N'-carbonyldiimidazole (CDI) was added to dimethylformamide in which the protected caffeic phosphoric acid or dihydroxyhydrocinnamic acid was dissolved and after 30 minutes, N-biotinyl-hexanediamine 1 mmol and 2 mmol of triethylamine were added thereto. The mixture was stirred at room temperature for 1 day and then evaporated.

The residue after evaporation was washed with 0.1 M HCl, 10% NaHCO ₃ solution and water, and reacted with 3 ml of acetic acid containing 3 M HCl to remove the protecting group.

Then, the reaction mixture was evaporated, washed with 10% NaHCO ₃ solution and diethyl ether, and then dried in a vacuum oven to obtain dihydroxyhydrocinnamic acid conjugated with biotin-conjugated caffeic acid and ivory-colored powder of gray powder .

Biotin-bound compounds synthesized through the above process were identified through thin layer chromatography, IR, and ¹ H-NMR.

[BTCA] Total yield: 31%; mp .: 166-169 [deg.] C; IR (nujol mull):? _Max (cm ^-1 ) = 1702 (C = O, -HNCONH-), 1651 (C = O, amide), 1638 (C = O, amide); ^{1 H-NMR (DMSO-d6} ):): δ = 1.25-1.58 (m, 14H), 2.02 (t, 2H, J = 7.3 Hz) 2.56 (d, 1H, J = 12.4 Hz) 2.80 (dd, 1H 1H, J = 12.4 Hz, 5.2 Hz), 2.99 (m, 2H), 3.07 (m, , J = 15.6 Hz), 6.68 (d, 1H, J = 7.9 Hz), 6.77 (dd, 1H, J = 8.0 Hz, 1.4 Hz), 6.88 (d, 1H, J = 1.6 Hz), 7.18 (d, 1H, &lt; / RTI &gt; J = 15.6 Hz)

[BTDHHC] Total yield: 38%; mp .: 140-143 DEG C; IR (nujol mull):? _Max (cm ^-1 ) = 1703 (C = O, H ₂ NCONH ₂ ), 1663 (C = O, amide), 1639 (C = O, amide); ^{1 H-NMR (DMSO-d6} ):): δ = 1.19-1.59 (m, 14H), 2.03 (t, 2H, J = 7.4 Hz), 2.23 (t, 2H, J = 6.9 Hz) 2.55-2.60 ( 1H), 4.29 (m, 1H), 6.39 (m, 1H), 2.80 (dd, 1H, J = 12.4 Hz, dd, 1H, J = 8.1 Hz , 2.0 Hz), 6.54 (d, 1H, J = 2.0 Hz), 6.58 (d, 1H, J = 7.8 Hz)

< Example  2> urate (monosodium urate monohydrate ) And production of gout animal model and drug treatment

C57BL / 6 mice were used to derive a model of air-pouch, a local inflammatory response model. First, 5 ml of sterilized air was injected into the subcutaneous tissues of mice and then 5 ml of the second sterilized air was injected by the same method 3 days later.

Seven days after the first air injection, MSU crystals suspended in 1 ml of sterilized non-toxin PBS were injected into the air bag to prepare a model animal model. One hour before the injection of MSU, BTCA 30 mg / kg or PBS Were injected into the oral cavity.

After 6 hours of injection of MSU crystals, the mice were euthanized, and the liquid in the air-pouch was collected by washing with 2 ml of PBS containing 5 mM EDTA, centrifuged at 12,000 rpm for 2 minutes, The sample was cooled and stored before measurement.

< Example  3> By MSU  Confirming the inhibitory effect of CAPE on induced inflammation

Mononuclear cells exposed to MSU crystals are induced to induce IL-1β activation and IL-1β activation is induced by NLRP3 inflammatory mediators (Martinon F, Petrilli V, Mayor A, Tardivel A, Tschopp J. Gout-associated uric acid crystals activated in NALP3 inflammasome. Nature 2006, 440 (7081): 237-241. The change in IL-1β secretion by biotin-conjugated compounds was confirmed by ELISA (Koo JE et al, 2012) analysis (R & D Systems; Minneapolis,

As a result, as shown in FIG. 1, it was confirmed that IL-1β secretion by MSU crystals was remarkably decreased in animals modeled by oral administration of BTCA and BTDHC among the biotin-conjugated compounds.

From the above results, it was found that bifunctional caffeic acid compound (BTCA) strongly inhibited the secretion of IL-1β induced by MUS, indicating that the BTCA compound inhibited the induction of IL- Lt; RTI ID = 0.0 &gt; 1beta &lt; / RTI &gt;

Therefore, the caffeic acid compound bound with biotin can inhibit the activity of IL-1? Induced by the inflammose to inhibit the inflammatory reaction induced by the urate crystals, thereby treating gout.

Hereinafter, formulation examples of the pharmaceutical composition of the present invention will be described, but the present invention is not intended to be limited thereto but is specifically explained.

< Formulation example  1> Preparation of injection

10 mg of BTCA, 3.0 mg of sodium metabisulfite, 0.8 mg of methylparaben, 0.1 mg of propylparaben and 0.1 ml of sterile distilled water for injection are mixed and mixed to a final volume of 2 ml by a conventional method. Filled and sterilized to prepare an injection.

&Lt; Formulation Example 2 > Preparation of tablet

10 mg of BTCA, 100 mg of lactose, 100 mg of starch and an appropriate amount of magnesium stearate were mixed and tableted according to a conventional tablet preparation method.

&Lt; Formulation Example 3 > Preparation of capsules

10 mg of BTCA, 50 mg of lactose, 50 mg of starch, 2 mg of talc, and an appropriate amount of magnesium stearate were mixed and filled in gelatin capsules according to a conventional capsule preparation method to prepare capsules.

Hereinafter, a production example of health food using CAPE according to the present invention will be described, but it is not intended to limit the present invention but to describe it specifically.

&Lt; Preparation Example 1 > Preparation of health food

BTCA 10 mg, Vitamin A acetate 70,, Vitamin E 1.0 mg, Vitamin B 1 0.13 mg, Vitamin B 2 0.15 mg, Vitamin B 6 0.5 mg, Vitamin B 12 0.2,, Vitamin C 10 mg, Biotin 10 (Ferrous sulfate 1.75 mg, zinc oxide 0.82 mg, magnesium carbonate 25.3 mg, potassium phosphate monobasic 15 mg, dibasic calcium phosphate 55 mg, nicotinate amide 1.7 mg, folic acid 50 mg, calcium pantothenate 0.5 mg) Mg of calcium citrate, 90 mg of potassium citrate, 100 mg of calcium carbonate, and 24.8 mg of magnesium chloride) were mixed to prepare a granule, and a health food was prepared according to a conventional method.

&Lt; Preparation Example 2 > Preparation of health drink

10 mg of BTCA, 1000 mg of citric acid, 100 g of oligosaccharide, 2 g of a plum concentrate, 1 g of taurine and purified water were added to make a total of 900 ml, and the above components were mixed according to a conventional health drink manufacturing method, The solution was filtered and sterilized in a sterilized 2 L container, and then refrigerated.

While the present invention has been particularly shown and described with reference to specific embodiments thereof, those skilled in the art will appreciate that such specific embodiments are merely preferred embodiments and that the scope of the present invention is not limited thereby. something to do. It is therefore intended that the scope of the invention be defined by the claims appended hereto and their equivalents.

Claims (5)

A pharmaceutical composition for prevention or treatment of gout comprising, as an active ingredient, a compound (BTCA) represented by the following formula (1), wherein caffeic acid and biotin are combined:
[Chemical Formula 1]

In Formula 1,
R ¹ or R ² may be the same or different and is any one of OH, NH _2, and SH,
Solid lines with dashed lines indicate single or double bonds. The pharmaceutical composition for preventing or treating gout according to claim 1, wherein said gout is caused by monosodium urate monohydrate (MSU) crystals. The pharmaceutical composition for prevention or treatment of gout according to claim 1, wherein the compound inhibits the production of IL-1? Activated by a urate (MUS) crystal. [Claim 2] The pharmaceutical composition for preventing or treating gout according to claim 1, wherein the pharmaceutical composition comprises 0.01 to 90 parts by weight of the compound per 100 parts by weight of the total amount of the pharmaceutical composition. A health food for prevention or improvement of gout comprising caffeine acid and biotin combined with a compound represented by the following formula (1) as an active ingredient:
[Chemical Formula 1]

In Formula 1,
R ¹ or R ² may be the same or different and is any one of OH, NH _2, and SH,
Solid lines with dashed lines indicate single or double bonds.

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