KR101590546B1 - Antioxidant Composition Comprising 1-Methyl-1H-Benzimidazole Derivatives As An Effective Ingredient - Google Patents

Abstract

The present invention relates to a composition for antioxidant containing 1-methyl- 1H -benzimidazole derivative or a pharmaceutically acceptable salt thereof as an active ingredient. The 1-methyl- 1H -benzimidazole derivative of the present invention A pharmaceutical composition and a health food for prevention and treatment of related diseases due to oxidative stress, having excellent DPPH radical scavenging ability, reducing ability, OH radical scavenging ability and polyphenol oxidase inhibiting activity and excellent antioxidative effect by inhibiting xanthine oxidase Can be usefully used.

Description

An antioxidant composition comprising 1-methyl-1H-benzimidazole derivative as an active ingredient (Antioxidant Composition Comprising 1-Methyl-1H-Benzimidazole Derivatives As An Effective Ingredient)

The present invention relates to a composition for antioxidant containing 1-methyl- 1H -benzimidazole derivative as an active ingredient.

Free radicals, which are active oxygen species, include hydroxy, peroxy, superoxide radicals, hydrogen peroxide, singlet oxygen and various lipid peroxides. The lipid peroxides may react with various metabolic enzymes and small molecules of membrane lipids, proteins, nucleic acids and biological systems. Free radicals play an important role in the initiation and progression of various diseases such as atherosclerosis, cardiovascular diseases, aging, respiratory diseases, cancer and gout (see Non-Patent Document 1 and Non-Patent Document 2).

An antioxidant is a substance capable of preventing or delaying the oxidation of a molecule. Oxidation reaction refers to damage to a cell by transferring electrons from a substance to an oxidant to generate free radicals. However, most cells contain a complex network of antioxidants and enzymes to prevent oxidative damage to intracellular components such as DNA, proteins and lipids. Antioxidants remove or inhibit active species before such active species damage essential cellular components of cells or tissues.

Gout, one of the diseases caused by oxidative stress, is caused by immersion of uric acid in the formation of uric acid nodules in the lubricated joint during purine catabolism by xanthine oxidase (see Non-Patent Document 3). The xanthine oxidase enzyme acts to convert hypoxanthine to xanthine and xanthine to uric acid. At this time, it is accompanied by the generation of hydrogen peroxide and superoxide anion as by-products, which causes oxidative stress to the cells or generation of gout in the human body (see Non-Patent Document 4).

Recently, it has been reported that secondary metabolites have important biological characteristics such as electron transfer function, free radical scavenging, blocking ability, oxidase inhibition and anti-inflammatory activity, and clinically, trans- retinoic acid, ATRA) and retinol (vitamin A) have been used in the treatment of proliferative skin diseases and some tumors, but the application of retinoid therapy has been limited by the toxicity of natural retinoids. Therefore, there is a need to develop a novel substance having an activity capable of removing active oxygen species with low toxicity and side effects for the treatment of the above-mentioned diseases.

On the other hand, benzimidazole and derivatives thereof are considered to be very important compounds due to their wide use in the biological field. Several benzimidazole derivatives have been reported to exhibit anticancer, antihypertensive, antiviral, vasodilatation and antimicrobial activity, and these compounds are also useful as antitumor agents, antihistamines, anti-HIV and selective neuropeptide YY1 receptor antagonists, Has been found to act as a 5-lipoxygenase inhibitor, a factor Xa (FXa) inhibitor, for use as an agent. Thus, a new method for the synthesis of this essential part is being developed by this property of the benzimidazole centers. In addition, the benzimidazole is an important intermediate in many organic reactions and acts as a ligand to the transition metal for modeling the biological system. Several methods for the synthesis of these benzimidazole derivatives have been developed, of which two protocols follow. The first is to couple the O -phenylenediamine with the carboxylic acid or its derivative, and the second involves condensation of the O -phenylenediamine and the aldehyde to carry out the oxidative dehydrogenation (cyclo-dehydrogenation) . Among them, the second method is mainly used because of easy accessibility of various substituted aldehydes.

Accordingly, there is a need to develop novel substances that can effectively treat diseases caused by oxidative stress by synthesizing benzimidazole derivatives having various substituents having antioxidative action.

 Ames B. N. et al., Ann New York Acad Sci, 663: 85-89, 1992.  Esterbauer H. et al., Free Rad Res. Comm., 6: 67-75, 1989.  Owen P. L. et al., J. Ethnopharmacol., 64: 149-160, 1999.  Park D. A. et al., Acta Phy Scand, 548: 87-99, 1986.  Sharma S.D. et al., Synthetic Communications 1, 39: 980-991, 2009.  Ji-Tai Li. et al., Ultrasonics Sonochem, 17: 356-358, 2010.  Langley-Evans S. C., Int J Food Sci Nutr, 51: 181-188, 2000.  Elizabeth K. et al., Int J Pharmaceut., 58: 237-240, 1990.  Gacche R. N. et al., J Enzyme Inhib Med Chem, 23: 28-31, 2008.  Khobragade C. N. et al., J Enz Inhib Med Chem, 23: 341-346, 2008.  Takno Y. et al., Life Sci, 76: 1835-1847, 2005.

An object of the present invention is to provide an antioxidative composition containing 1-methyl- 1H -benzimidazole derivative as an active ingredient.

In order to achieve the above object, the present invention provides a method for preventing and treating diseases caused by oxidative stress, comprising 1-methyl-1 H -benzimidazole derivative represented by Chemical Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient A pharmaceutical composition is provided.

The present invention also provides a health food for preventing and ameliorating diseases caused by oxidative stress, comprising 1-methyl-1 H -benzimidazole derivative represented by the formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient do.

Hereinafter, the present invention will be described in detail.

1. 1- methyl -One H - Benzimidazole (One- methyl -One H - 안이 메디azole ) Derivative

The present invention provides 1-methyl- 1H -benzimidazole derivatives.

The 1-methyl- 1H -benzimidazole derivative is a compound having a chemical structure represented by the following formula (1).

Wherein R ₁ is C ₅ -C ₂₄ aryl or C ₅ -C ₂₄ heteroaryl containing at least one nitrogen or oxygen atom, preferably C ₆ -C ₁₄ aryl or nitrogen Or a C ₅ -C ₁₄ heteroaryl containing at least one oxygen atom, more preferably a C ₆ -C ₁₀ aryl or a C ₅ -C ₁₀ heteroaryl containing at least one nitrogen or oxygen atom.

일 수 있다. 이때 상기 R₂는 수소, C₁-C₆의 알킬, 할로겐, 히드록시 및 C₁-C₆의 알콕시로 이루어진 군으로부터 선택되는 어느 하나의 작용기를 가질 수 있다. Specifically, in one aspect of the present invention, R ₁ in the formula (1)

Lt; / RTI > Wherein R ₂ may have any one functional group selected from the group consisting of hydrogen, C ₁ -C ₆ alkyl, halogen, hydroxy and C ₁ -C ₆ alkoxy.

More specifically, the 1-methyl- 1H -benzimidazole derivative represented by the formula (1) having R ₁ is represented by the following formulas (2) to (13)

일 수 있고, 상기 R₂는 수소, C₁-C₆의 알킬, 할로겐, 히드록시 및 C₁-C₆의 알콕시로 이루어진 군으로부터 선택되는 어느 하나의 작용기일 수 있으며, 이를 갖는 1-메틸-1H-벤즈이미다졸 유도체를 구체적으로 예시하면 하기 화학식 14와 같다:In another aspect of the present invention, R < ₁ >

And R ₂ may be any functional group selected from the group consisting of hydrogen, C ₁ -C ₆ alkyl, halogen, hydroxy and C ₁ -C ₆ alkoxy, and 1-methyl- The 1 H -benzimidazole derivative is specifically exemplified by the following chemical formula 14:

일 수 있고, 상기 A는 산소 또는 질소일 수 있으며, 상기 R₂는 수소, C₁-C₆의 알킬, 할로겐, 히드록시 및 C₁-C₆의 알콕시로 구성된 군에서 선택된 어느 하나의 작용기일 수 있고, 상기 A가 질소이고 R₂는 수소, C₁-C₃의 알킬, 할로겐, 히드록시 또는 C₁-C₃의 알콕시인 것이 바람직하다. 이러한 R₁을 갖는 1-메틸-1H-벤즈이미다졸 유도체는 구체적으로 하기 화학식 15로 예시할 수 있다:In still another aspect of the present invention, R < ₁ >

, Wherein A may be oxygen or nitrogen and R ₂ is any one selected from the group consisting of hydrogen, C ₁ -C ₆ alkyl, halogen, hydroxy and C ₁ -C ₆ alkoxy Preferably, A is nitrogen and R ₂ is hydrogen, C ₁ -C ₃ alkyl, halogen, hydroxy or C ₁ -C ₃ alkoxy. The 1-methyl- 1H -benzimidazole derivative having R ₁ can be specifically exemplified by the following formula (15)

The 1-methyl-1 H -benzimidazole derivatives of the present invention which can have various side chains as described above can be prepared by reacting O -phenylenediamine with various aldehyde groups using silica trichloroacetic acid as a catalyst in an ethanol / water mixture (See Fig. 1 and non-patent document 5). In the present invention, each of 1-methyl-1 H -benzimidazole derivatives can be synthesized by using 14 types of aromatic aldehydes in the synthesis (see Table 1).

2. Oxidative  Pharmaceutical composition for the prevention and treatment of diseases caused by stress

The present invention provides a pharmaceutical composition for preventing and treating diseases caused by oxidative stress, which comprises the compound of formula (I) or a pharmaceutically acceptable salt thereof as an active ingredient.

The compound represented by the above formula (1) is the same as described in the above item " 1- methyl- 1H- benzimidazole derivative & quot ;. Therefore, a detailed description thereof will be omitted for the sake of explanation of the above items, and only the constitution specific to the "pharmaceutical composition" will be described below.

The term "pharmaceutically acceptable salt" of the present invention means salts prepared by conventional methods, which are well known to those skilled in the art. Such "pharmaceutically acceptable salts" include, but are not limited to, salts derived from pharmacologically or physiologically acceptable inorganic and organic acids and bases.

The disease caused by oxidative stress is caused by an imbalance in the production of reactive oxygen species (ROS) such as free radicals such as superoxide, peroxide, and hydroxyl radical, the removal of reactive oxygen species, and the regeneration of damaged complex materials And can induce various cytotoxic reactions such as lipid peroxidation and cell death. The disease caused by such oxidative stress may be aging, gout, kidney disease, hypertension, diabetes, arteriosclerosis, obesity, liver disease, lung disease, heart disease, hyperlipidemia or stroke, aging or gout, but is not limited thereto .

In order to examine the antioxidative activity of 1-methyl- 1H -benzimidazole derivatives of the present invention, the inventors of the present invention found that DPPH (2,2-diphenyl-1-picryl hydrazine) radical scavenging ability, reducing ability, OH radical scavenging ability, The inhibition of polyphenol oxidase (PPO) and the inhibition of xanthine oxidase (XO) were examined. As a result, the benzimidazole derivatives of the present invention showed excellent DPPH radical scavenging activity and remarkable reducing ability (see Table 2). Herein, the DPPH radical scavenging activity is mainly used for measuring the antioxidant activity of DPPH because of its unique electrons exhibiting a strong absorption peak at 517 nm, and DPPH radical scavenging ability and reducing ability can be measured by hydroxylation , Which may be involved in providing electrons and thus stabilizing the radical compound. In addition, the benzimidazole analogues of the present invention have OH radical scavenging activity (see Table 2). OH radicals are known to play a very important role in the physiological regulation of cells, and in the inflammatory diseases such as rheumatoid arthritis and gout, the reaction of nitric oxide with superoxide generates nitric oxide, which is commonly found in inflammatory and ischemic areas, This produces OH radicals. Thus, the OH radicals produced by this reaction cause membrane damage to the cells at the site of inflammation. In addition, the benzimidazole derivatives were found to strongly inhibit PPO activity (see Table 2). The PPO is a copper-containing enzyme that catalyzes oxidized phenols with reactive orthoquinone, and the free radicals generated by the oxygen-induced transition metal mediates the development of diseases such as iron overload, rheumatoid arthritis and cancer. In general, cellular systems using molecular oxygen and transition metal ions produce free radicals and exhibit altered physiological effects. In addition, the benzimidazole analogues of the present invention were found to have a very potent XO inhibitory activity (see Table 3). Therefore, the 1-methyl-1 H -benzimidazole derivative of the present invention can be effectively used as an antioxidant through the above-mentioned activity, and can be used as an effective substance for ventilation by acting as an XO inhibitor.

The compounds of Formula 1 may be included in the pharmaceutical composition as such, as well as in the form of pharmaceutically acceptable salts of the compounds of Formulas 1 to 14. As the salt, an acid addition salt formed by a pharmaceutically acceptable free acid is useful. The acid addition salts may be prepared by reacting an inorganic acid such as hydrochloric acid, nitric acid, phosphoric acid, sulfuric acid, hydrobromic acid, hydroiodic acid, nitrous acid or phosphorous acid with an aliphatic mono- and dicarboxylate, a phenyl-substituted alkanoate, a hydroxyalkanoate and Alkanedioates, aromatic acids, non-toxic organic acids such as aliphatic and aromatic sulfonic acids. Such pharmaceutically non-toxic salts include sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, nitrate, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate chloride, bromide, But are not limited to, iodide, fluoride, acetate, propionate, decanoate, caprylate, acrylate, formate, isobutyrate, caprate, heptanoate, propiolate, oxalate, malonate, succinate, But are not limited to, but are not limited to, butyric acid, succinic acid, succinate, sebacate, sebacate, fumarate, maleate, butyne-1,4-dioate, hexane-1,6-dioate, benzoate, , Methoxybenzoate, phthalate, terephthalate, benzenesulfonate, toluene sulfonate, chlorobenzene Propionate, naphthalene-1, < / RTI > maleate, tartrate, methanesulfonate, propanesulfonate, -Sulfonate, naphthalene-2-sulfonate or mandelate.

When the benzimidazole derivative compound of the present invention is prepared from a pharmaceutical composition, the pharmaceutical composition may further comprise a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier to be contained in the pharmaceutical composition of the present invention is one commonly used in the present invention and includes lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia rubber, calcium phosphate, alginate, gelatin, But are not limited to, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrups, methylcellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. It does not. The pharmaceutical composition may further contain a lubricant, a wetting agent, a sweetening agent, a flavoring agent, an emulsifying agent, a suspending agent, a preservative, etc. in addition to the above components. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington ' s Pharmaceutical Sciences (19th ed., 1995).

The pharmaceutical composition of the present invention can be administered in various formulations, oral and parenteral, at the time of actual clinical administration. Solid formulations for oral administration may include tablets, pills, powders, granules, capsules, and the like. Liquid form preparations for oral use may include suspensions, solutions, emulsions and syrups. Agents for parenteral administration may include sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, and freeze-drying agents. Parenteral administration, it is preferable to select an injection method such as extracorporeal or intraperitoneal injection, intramuscular injection, subcutaneous injection, intravenous injection, intramuscular injection or intrathoracic injection.

The dosage of the pharmaceutical composition of the present invention varies depending on the patient's body weight, age, sex, health condition, diet, administration time, administration method, excretion rate, and severity of disease. The dosage ranges vary depending on the patient's body weight, age, sex, health condition, diet, time of administration, method of administration, excretion rate and severity of disease, and the daily dosage is 0.00001 To 10 g / kg, preferably 0.0001 to 1 g / kg, and can be administered 1 to 6 times a day.

The pharmaceutical composition of the present invention can be used alone or in combination with methods for the treatment and prevention of oxidative stress related diseases or using surgery, hormone therapy, chemotherapy and biological response modifiers.

The pharmaceutical composition of the present invention may be formulated into a unit dosage form by using a pharmaceutically acceptable carrier and / or excipient according to a method which can be easily carried out by those having ordinary skill in the art to which the present invention belongs. Or by intrusion into a multi-dose container. The formulations may be in the form of solutions, suspensions, syrups or emulsions in oils or aqueous media, or in the form of excipients, powders, powders, granules, tablets or capsules, and may additionally contain dispersing or stabilizing agents.

3. Oxidative  Health food for preventing and improving diseases caused by stress

Another aspect of the present invention is a pharmaceutical composition for preventing or ameliorating diseases caused by oxidative stress, comprising 1-methyl-1 H -benzimidazole derivative represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient Provide food.

Also, the compound represented by the above formula (1) will be explained with reference to the item " 1. 1- methyl- 1H- benzimidazole derivative ", and a detailed description thereof will be omitted. Hereinafter, Only the configuration will be described.

The disease caused by the oxidative stress is preferably one selected from the group consisting of aging, gout, kidney disease, hypertension, diabetes, arteriosclerosis, obesity, liver disease, lung disease, heart disease, hyperlipidemia and stroke, It is more preferable that it is ventilation, but is not limited thereto.

When the benzimidazole derivative compound of the present invention is prepared as a health food, it may further include components that are ordinarily added in the manufacture of a food, for example, a protein, a carbohydrate, a fat, a nutrient, a seasoning agent, . Examples of the above-mentioned carbohydrates include conventional saccharides such as monosaccharides such as glucose and fructose, disaccharides such as maltose, sucrose and oligosaccharides, and polysaccharides such as dextrin and cyclodextrin, and xylitol such as sorbitol, erythritol And the like. Natural flavors such as tau martin, stevia extract and synthetic flavoring agents can be used as the flavoring agent. For example, when the health food of the present invention is prepared as a drink, citric acid, liquid fructose, sugar, glucose, acetic acid, malic acid, fruit juice, natural extract and the like may be further added in addition to the benzimidazole derivative of the present invention.

Since the 1-methyl- 1H -benzimidazole derivative of the present invention is excellent in antioxidative and XO inhibitory activity, the benzimidazole derivative or a pharmaceutically acceptable salt thereof is useful for prevention of diseases caused by oxidative stress such as gout Or as an active ingredient of a health food for improvement.

Since the 1-methyl-1 H -benzimidazole derivatives or pharmaceutically acceptable salts thereof according to the present invention have DPPH radical scavenging activity, reducing ability, OH radical scavenging activity and PPO inhibiting activity, they can be used not only as antioxidants , A powerful XO inhibitory action, and thus can be effectively used as a therapeutic agent for diseases caused by oxidative stress such as XO inhibitor or gout.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a diagrammatic view illustrating a method for synthesizing 2-aryl-1-arylmethyl-1 H -benzimidazole derivatives of 1-methyl- 1H -benzimidazole derivatives of the present invention.

Hereinafter, the present invention will be described in more detail by way of examples.

However, the following examples are for illustrative purposes only and are not intended to limit the scope of the present invention.

Example  1. 1- methyl -One H - Benzimidazole  Derivative synthesis

In order to synthesize 1-methyl-1 H -benzimidazole derivatives, 2-aryl-1-methyl- 1H -benzimidazole derivatives such as 1-methyl- 1H -benzimidazole derivatives were synthesized according to the synthesis and characterization methods shown in the prior art 1-arylmethyl-1 H -benzimidazole derivatives were synthesized (Fig. 1).

As a result, as shown in Table 1, 14 types of 1-methyl- 1H -benzimidazole derivatives having different aldehyde groups were synthesized (Table 1).

Example  2. DPPH Radical  Confirmation of scavenging activity

To confirm the DPPH radical scavenging activity of the 1-methyl- 1H -benzimidazole derivative synthesized in Example 1, each 1-methyl- 1H -benzimidazole derivative and DPPH were dissolved in the same molar concentration 1 mM in ethanol). The mixture was stirred vigorously and the absorbance at 515 nm against the blank was measured 45 minutes after incubation when the reaction reached steady state. The% inhibition of the radical scavenging activity was calculated according to the following formula.

[Equation 1]

% Inhibition = (A _C (0) - A _A (t) / A _C (0)) x 100

Where A _c (0) is the absorbance of the control at t = 0 and A _A (t) is the absorbance of the antioxidant at t = 1 h. Measurement was repeated three times and quercetin (1 mM) was used as a standard compound.

As a result, all the benzimidazole derivatives were found to have DPPH radical scavenging activity, as shown in Table 2 below. Compared with the standard compound quercetin (45.8%), the overall range of DPPH scavenging activity of the benzimidazole derivatives of the present invention was found to be 45.6 to 16.5%. In particular, the compounds B3, B10, B12 and B4 showed high antioxidative activity (Table 2).

Example  3. Reducible  Confirm

The Fe ^{3 +} reducibility of benzimidazole derivatives was evaluated by the slightly modified Langley-Evans method (See Non-Patent Document 7). Specifically, a solution containing each 1-methyl- 1H -benzimidazole derivative 500 ml (1 mM in 0.5% v / v dimethyl sulfoxide) was mixed with 3 ml of 1 mM potassium ferricyanide solution and cultured in a water bath at 50 ° C for 20 minutes. After incubation, 0.5 ml of TCA (10%) was added to terminate the reaction. 1 ml of distilled water was mixed with 1 ml of the supernatant of this solution, and 0.1 ml of FeCl ₃ solution (0.01%) was added. The reaction mixture was left at room temperature for 10 minutes and absorbance was measured at 700 nm for the appropriate blank solution. All tests were repeated 6 times, and the higher the absorbance of the reaction mixture, the greater the reducing power, and glutathione (1 mM) was used as a standard compound.

As a result, the benzimidazole derivative showed a reducing power ranging from 31.2 to 12.8% as compared with glutathione (38.8%). In particular, Compounds B10, B3 and B13 were found to be highly effective reducing agents as compared to other benzimidazole derivatives (Table 2).

Example  4. OH Radical  Confirmation of scavenging activity

The OH radical scavenging activity was slightly modified by Elizabeth and Rao (See Non-Patent Document 8). This method was based on quantifying 2-deoxyribose degradation products as they were condensed with TBA. Hydroxyl radicals were produced by the Fe ^{3 +} - ascorbate - EDTA - H ₂ O ₂ system (pantone reaction). The reaction mixture was added to 1 ml of the final volume with 2-deoxy-2-ribose (2.8 mM), KH ₂ PO ₄ -KOH buffer (20 mM, pH 7.4), FeCl ₃ (100 μM) H ₂ O ₂ (1.0 mM), ascorbic acid (100 μM) and 0.1 ml of each 1-methyl- 1H -benzimidazole derivative (1 mM). The reaction mixture was incubated at 37 ° C for 1 hour, added with 1 ml of 2.8% TCA and then added with 1 ml of 1% aqueous TBA and incubated at 90 ° C for 15 minutes for color development. After cooling, the absorbance was measured at 532 nm against the appropriate blank solution. All tests were repeated 6 times and glutathione (1 mM) was used as a positive control. % Inhibition was determined by comparing the test and blank solutions.

The results showed that compound B6 (15.2%) was the strongest OH radical scavenging agent followed by compound B12 (14.5%) and B3 (14.2%) with high OH radical scavenging activity (Table 2).

Example  5. Polyphenol oxidase ( polyphenol oxidase , PPO ) Inhibition activity confirmation

The PPO inhibition assay was performed according to the method described by Gacche et al. (See non-patent reference 9), and the PPO enzyme was extracted from the apple by ammonium sulfate precipitation. The reaction mixture contained L-DOPA (0.75 ml solution) and each 2-aryl-1-arylmethyl-1 H -benzimidazole derivative (1.5 ml, 2 mM), citrate buffer (0.75 ml, pH 4.8, ). After reaction for 10 minutes, a brown chromophore (dopachrome) was measured at 470 nm. L-cysteine (1 mM), a known PPO inhibitor was used as a standard inhibitor.

As a result, compounds B10 (46.2), B11 (43.5), B1 (41.4) and B6 (40.3) strongly inhibited the activity of PPO and other benzimidazole derivatives inhibited PPO in the range of 20.8-38.6% (Table 2).

The results are expressed as the mean value ± S of n = 3.

^* Standard compound

NR: no reaction under experimental conditions, ND: not measured.

Example  6. Xanthine  Oxidase ( xanthine oxidase , XO ) Confirmation of inhibitory activity

Bovine milk XO (Grade 1, ammonium sulfate suspension, Sigma-Aldrich) activity was analyzed spectrophotometrically by measuring uric acid formation at 293 nm using a UV-visible spectrophotometer (see non-patent documents 10 and 11). The reaction mixture contained 50 mM potassium phosphate buffer (pH 7.6), 75 [mu] M xanthan and 0.08 units of XO. The XO inhibitory activity of each of the 1-methyl-1 H -benzimidazole derivatives was measured according to the decrease of uric acid formation at 293 nm at 25 ° C. The XO enzyme was first incubated with each benzimidazole derivative dissolved in DMSO (1% v / v) for 5 minutes, and the reaction was started by adding xanthine. The final concentration of DMSO (1% v / v) did not affect enzyme activity. All experiments were repeated three times and the measured values were expressed as the mean value of three experiments. At this time, allopurinol was used as a positive control, and% inhibition was measured by comparing test solution and blank solution.

As a result, all of the benzimidazole derivatives are excellent XO inhibition appeared to exhibit activity, in particular the control of allopurinol (IC ₅₀ = 5.5 μM) and the compound B3 (4.3 μM), B10 ( 4.4 μM) compared, and B6 ( 4.5 μM) was found to be most effective. The benzimidazole derivatives of the present invention exhibit IC ₅₀ in the range of 5.1-8.7 μM, indicating that they are potent XO-inhibiting substances (Table 3).

^a Results are expressed as mean ± SD of n = 3.

^* Standard compound

The preparation examples for the composition of the present invention are illustrated below.

Manufacturing example  1. Preparation of pharmaceutical preparations

<1-1> Sanje  Produce

200 mg of 1-methyl- 1H -benzimidazole derivative

20 mg of lactose

After mixing the above components, the mixture was packed in an airtight container to prepare a powder.

<1-2> Preparation of tablets

500 mg of 1-methyl- 1H -benzimidazole derivative

Corn starch 100 mg

Lactose 100 mg

2 mg of magnesium stearate

After mixing the above components, tablets were prepared by tableting according to a conventional method for producing tablets.

&Lt; 1-3 > Preparation of capsules

500 mg of 1-methyl- 1H -benzimidazole derivative

3 mg of crystalline cellulose

Lactose 14.8 mg

Magnesium stearate 0.2 mg

After mixing the above components, the capsules were filled in gelatin capsules according to the conventional preparation method of capsules.

<1-4> Liquid  Produce

1-Methyl-1 H -benzimidazole derivative 2 g

10 g per isomer

5 g mannitol

Purified water quantity

Each component was added to purified water in accordance with the usual liquid preparation method and dissolved, and the lemon flavor was added in an appropriate amount. Then, the above components were mixed, and purified water was added thereto. The whole was adjusted to 100 ml with purified water, And sterilized to prepare a liquid preparation.

<1-5> Preparation of Injection

1-methyl- 1H -benzimidazole derivative 10 mg / ml

Diluted hydrochloric acid BP until pH 7.6

Sodium chloride BP Up to 1 ml

The 1-methyl- 1H -benzimidazole derivative is dissolved in an appropriate volume of sodium chloride BP. The pH of the resulting solution is adjusted to pH 7.6 using dilute hydrochloric acid BP, the volume is adjusted using sodium chloride BP, Respectively. The solution is filled in a 5 ml Type I ampoule made of clear glass, sealed in the upper lattice of the air by dissolving the glass, and sterilized at 120 캜 for 15 minutes or longer to prepare an injection solution.

Manufacturing example  2. Manufacturing of food

Foods containing the 1-methyl- 1H -benzimidazole derivatives of the present invention were prepared as follows.

<2-1> Solar  Produce

Brown rice, barley, glutinous rice, and yulmu were dried by a known method and dried, and the mixture was granulated to a powder having a particle size of 60 mesh.

Black soybeans, black sesame seeds, and perilla seeds were steamed and dried by a known method, and then they were prepared into powders having a particle size of 60 mesh by a grinder.

The 1-methyl-1 H -benzimidazole derivative of the present invention was concentrated under reduced pressure in a vacuum concentrator, dried by spraying, and dried with a hot-air drier, and pulverized to a size of 60 mesh with a pulverizer to obtain a dry powder.

The grains, seeds and 1-methyl-1 H -benzimidazole derivatives prepared above were blended in the following proportions.

(30 parts by weight of brown rice, 15 parts by weight of yulmu, 20 parts by weight of barley)

Seeds (7 parts by weight of perilla, 8 parts by weight of black beans, 7 parts by weight of black sesame seeds)

1-methyl- 1H -benzimidazole derivative (12 parts by weight),

(0.5 part by weight),

(0.5 parts by weight)

<2-2> Manufacture of health drinks

1-methyl- 1H -benzimidazole derivative 10 g

Citric acid 100 mg

100 mg of oligosaccharide

2 mg of plum concentrate

100 mg taurine

Purified water was added to 500 ml

The above components were mixed according to a conventional health drink manufacturing method, and the mixture was heated at 85 DEG C for about 1 hour with stirring, and the resulting solution was filtered to obtain a sterilized 1 liter container, sealed sterilized, It is used in the production of the health beverage composition of the invention.

Although the composition ratio is a mixture of the components suitable for the preferred beverage as a preferred embodiment, the blending ratio may be arbitrarily varied according to the regional and national preferences such as the demand level, the demanding country, and the intended use.

Claims (9)

1-methyl-1H-benzimidazole derivative or a pharmaceutically acceptable salt thereof selected from the group consisting of compounds represented by formulas (4), (5), (7) and A pharmaceutical composition for preventing and treating diseases caused by oxidative stress selected from the group consisting of aging, gout, kidney disease, diabetes, arteriosclerosis, liver disease, lung disease, heart disease, hyperlipidemia and stroke,
[Chemical Formula 4]

,
[Chemical Formula 5]

,
(7)

,
[Chemical Formula 15]

. delete delete delete delete delete 1-methyl-1H-benzimidazole derivative selected from the group consisting of compounds represented by formulas (4), (5), (7) and (15) or a pharmaceutically acceptable salt thereof as an active ingredient, For the prevention and amelioration of diseases caused by oxidative stress selected from the group consisting of diabetes, atherosclerosis, liver disease, lung disease, heart disease, hyperlipidemia, and stroke.
[Chemical Formula 4]

,
[Chemical Formula 5]

,
(7)

,
[Chemical Formula 15]

. delete delete

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