KR20120108139A - Compositions for antioxidation comprising extract of agarum clathratum - Google Patents

Abstract

PURPOSE: A composition for antioxidation containing Agarum clathratum extract is provided to remove free radical or reactive oxygen and to treat diseases caused by oxidative stress. CONSTITUTION: A composition for antioxidation contains Agarum clathratum extract as an active ingredient. The extract is prepared by polar organic solvent or non-polar organic solvent. The polar solvent is water, alcohol, acetic acid, DMFO(dimethyl-formamide), DMSO(dimethyl sulfoxide), or a mixture thereof. The non-polar solvent is actone, acetonitrile, ethyl acetate, methyl acetate, fluoroalkane, pentane, hexane, 2,2,4-trimethylpentane, decane, cyclohexane, cyclopentane, diisobutylene, 1-pentene, 1-chlorobutane, 1-chloropentane, -xylene, diisopropyl ether, 2-chloropropane, toluene, 1-chloropropane, chlorobenzene, benzene, diethyl ether, diethyl sulfide, chloroform, dichloromethane, 1,2-dichloroethane, aniline, diethyl amine, ether, carbon tetrachloride, THF, or a mixture thereof. A pharmaceutical composition for antioxidation contains the extract as an active ingredient.

Description

Composition for Antioxidation, Comprising Antipersimmon Seaweed Extract as Active Ingredient {Compositions for Antioxidation Comprising Extract of Agarum Clathratum}

The present invention relates to a composition for antioxidant comprising a hole seaweed extract as an active ingredient.

Seaweed ( Undaria pinnatifida ) is a specialty of Japan and Korea, and it is distributed in Korea from Baeknyeong Island on the west coast to the end of the east coast. Photosynthetic seaweeds are exposed to light and high concentrations of oxygen that can generate free radicals and other peroxides, making them more prone to photodynamic damage in the case of unsaturated fatty acids, which are chloroplast membrane components. Despite the presence of oxygen produced by photosynthesis and unsaturated fatty acids in the chloroplast membranes, the lack of such oxidative damage in algae suggests the presence of antioxidant mechanisms and antioxidant components in algae (Matsukawa, R. et al., J. Appl). . Phycol., 9:29, 1997) .

In normal metabolic processes in the human body, additional chemical particles called free radicals are produced. These free radicals constantly attack and damage the body's constituent cells, called oxidation. Damage caused by free radicals is repaired by the body's natural healing power, but if damages beyond the restorative ability are continuously applied, chronic harmful effects occur throughout life. These adverse effects reduce the function of cells and tissues and cause disease and aging.

Free radicals are molecules that have one or more unpaired electrons and have the property of stabilizing electrons from other molecules. In addition, free radicals have a lifespan as short as a few thousandths of a second, which makes them very unstable, aggressive and strong in oxidizing power. Most of the free radicals are reactive oxygen species, which include hydroxyl radicals, superoxide radicals, singlet oxygen and hydrogen peroxide (H ₂ O ₂ ). And alkoxyl radicals, which are produced by lipids and oxidation, have two-sided properties that cause fatal oxygen toxicity in living organisms. That is, these free radicals have non-selective and irreversible destruction effects on cell components such as lipids, proteins, sugars, and DNA. It is known to cause various diseases such as skin diseases, digestive diseases, inflammation, rheumatism and autoimmune diseases.

To protect itself from damage caused by these free radicals, the body is equipped with a series of oxidative protection systems. These protective systems include enzymes such as superoxide dismutase (SOD) and catalase, alpha-tocopherol (vitamin E), beta-carotene and ascorbic acid. fat-soluble and water-soluble antioxidants or radical scavengers such as (ascorbic acid, vitamie C) and glutathione.

Recently, research on antioxidants can be used in various fields such as food industry, fermentation and pharmaceutical industry, and agricultural field, and thus can have a very large ripple effect in terms of national economy and industry.

In particular, since the known antioxidants have many problems in their use as active substances due to their toxicity and limitations in their use, the development of new natural antioxidants with safer and stronger activity from natural products is required.

Numerous papers and patent documents are referenced and cited throughout this specification. The disclosures of the cited papers and patent documents are incorporated herein by reference in their entirety to better understand the state of the art to which the present invention pertains and the content of the present invention.

The present inventors have made diligent research efforts to develop substances which have antioxidant activity and are safe for humans, especially seaweed-derived substances. As a result, the present invention was completed by elucidating that Agarum Clathratum extract has the efficacy of antioxidant activity.

Therefore, it is an object of the present invention to provide an antioxidant composition comprising a hole seaweed seaweed extract as an active ingredient.

It is another object of the present invention to provide an antioxidant pharmaceutical composition comprising an extract of pomeson seaweed as an active ingredient.

In addition, another object of the present invention to provide an antioxidant improvement food composition comprising an extract of pomegranate seaweed as an active ingredient.

Other objects and advantages of the present invention will become more apparent from the following detailed description of the invention, claims and drawings.

According to an aspect of the present invention, the present invention is Agarum Clathratum ) provides an antioxidant composition comprising the extract as an active ingredient.

The present inventors made an intensive study to develop a substance which has antioxidant activity and is safe for the human body, in particular, algae-derived substance, and as a result, it was found that the perilla wakame extract has the efficacy of the antioxidant activity.

Agarum Clathratum ( Agarum Clathratum ), which is used as an active ingredient in the composition of the present invention, is distributed in Fukkaido , Kuril Islands, the Americas and the Bering Sea in Japan, and is distributed mainly in the north-eastern region of the east coast in Korea. Roots are thin fibrous, stems are columnar to oblong. The length is about 2-3 cm, the diameter is 3-4 mm, the lobes are elliptical, the length is 30-60 mm, the width is 20-30 cm, the base is heart-shaped, and the waves are wrinkled. There are large and small holes on the leaf surface, and the one near the middle is large and the one near the edge is small. The midrib is rather thick and raised on both sides of the leaf.

As used herein, the term “perforated seaweed extract” refers to an extract obtained from various organs or parts (eg, foliar, midrib, stem, and root) of perforated seaweed.

The composition of the present invention includes a hole seaweed extract as an active ingredient. As used herein, the term 'extract', which is used while referring to poached seaweed, is formulated (eg, powdered) so that the poultry seaweed itself can be administered to the animal as well as the extraction result obtained by treating the extraction solvent in the poultry seaweed. It also has the meaning to include a hole seaweed seaweed processed product.

When obtained by treating the extraction solvent in the poultry seaweed extract used in the composition of the present invention, a variety of extraction solvents may be used. Preferably, a polar solvent or a nonpolar solvent can be used. Suitable polar solvents include (i) water, (ii) alcohols (preferably methanol, ethanol, propanol, butanol, normal-propanol, iso-propanol, normal-butanol, 1-pentanol, 2-butoxyethanol Or ethylene glycol), (iii) acetic acid, (iv) dimethyl-formamide (DMFO) and (v) dimethyl sulfoxide (DMSO). Suitable as nonpolar solvents are acetone, acetonitrile, ethyl acetate, methyl acetate, fluoroalkane, pentane, hexane, 2,2,4-trimethylpentane, decane, cyclohexane, cyclopentane, diisobutylene, 1- Pentene, 1-chlorobutane, 1-chloropentane, o -xylene, diisopropyl ether, 2-chloropropane, toluene, 1-chloropropane, chlorobenzene, benzene, diethyl ether, diethyl sulfide, chloroform, dichloro Methane, 1,2-dichloroethane, anneal, diethylamine, ether, carbon tetrachloride and THF.

More preferably, the extraction solvent used in the present invention is (a) water, (b) anhydrous or hydrous lower alcohol having 1 to 4 carbon atoms (methanol, ethanol, propanol, butanol, etc.), (c) the lower alcohol and water Mixed solvent with (d) acetone, (e) ethyl acetate, (f) chloroform, (g) butyl acetate, (h) 1,3-butylene glycol, (i) hexane and (j) diethyl ether Include. Most preferably, the extract of the present invention is obtained by treating methanol with poached seaweed.

As used herein, the term "extract" has the meaning commonly used as a crude extract in the art as described above, but also broadly includes a fraction additionally fractionating the extract.

In other words, not only obtained by using the above-mentioned extraction solvent, but also obtained by further applying the purification process. For example, fractions obtained by passing the extract through an ultrafiltration membrane having a constant molecular weight cut-off value, separation by various chromatography (manufactured for separation according to size, charge, hydrophobicity or affinity), etc. The fraction obtained through the purification method is also included in the extract of poultice seaweed of the present invention.

The poultice seaweed extract used in the present invention may be prepared in a powder state by an additional process such as vacuum distillation and freeze drying or spray drying.

The present invention is a persimmon seaweed extract that exhibits antioxidant activity through scavenging ability against free redicals contained in various substances.

According to a preferred embodiment of the present invention, the composition of the present invention has the activity of removing the cationic radical (radical) of ABTS (2,2'-azino-bis (3-ethylbenzthiazoline-6-sulphonic acid).

According to a preferred embodiment of the present invention, the composition of the present invention has the activity of removing radicals of DPPH (2,2-diphenyl-1- picrylhydrazyl).

According to a preferred embodiment of the present invention, the composition of the present invention has the activity of reducing Fe (III) to Fe (II).

According to a preferred embodiment of the present invention, the composition of the present invention has the activity of inhibiting the breakage of single strand or double strand nucleotides by oxidation.

The composition of the present invention may be prepared as a pharmaceutical composition.

According to another aspect of the present invention, the composition of the present invention provides a pharmaceutical composition comprising (a) a pharmaceutically effective amount of the above-mentioned seaweed wakame extract of the present invention.

More preferably, the pharmaceutical composition of the present invention comprises (a) a pharmaceutically effective amount of pomegranate extract; And (b) a pharmaceutically acceptable carrier. As used herein, the term “pharmaceutically effective amount” means an amount sufficient to achieve the antioxidant efficacy or activity of the above-mentioned seaweed wakame extract.

When the composition of the present invention is made into a pharmaceutical composition, the pharmaceutical composition of the present invention includes a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers included in the pharmaceutical compositions of the present invention are those commonly used in the preparation, such as lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia rubber, calcium phosphate, alginate, gelatin, Calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil, and the like It doesn't happen. In addition to the above components, the pharmaceutical composition of the present invention may further include a lubricant, a humectant, a sweetener, a flavoring agent, an emulsifier, a suspending agent, a preservative, and the like. 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 may be administered orally or parenterally, and preferably applied by oral administration.

The appropriate dosage of the pharmaceutical composition of the present invention may vary depending on factors such as the formulation method, administration method, age, body weight, sex, pathological condition, food, administration time, administration route, excretion rate, . Typical dosages of the pharmaceutical compositions of the invention are in the range of 0.001-100 mg / kg on an adult basis.

The pharmaceutical compositions of the present invention may be prepared in unit dose form by formulating with a pharmaceutically acceptable carrier and / or excipient according to methods which can be easily carried out by those skilled in the art. Or may be prepared by incorporation into a multi-dose container. The formulation may be in the form of solutions, suspensions, syrups or emulsions in oils or aqueous media, or in the form of extracts, powders, powders, granules, tablets or capsules, and may further comprise dispersants or stabilizers.

The composition of the present invention may be provided as a food composition.

According to another aspect of the present invention, the composition of the present invention (a) provides a food composition comprising a food-effective amount of the above-mentioned seaweed seaweed extract of the present invention.

More preferably, when the composition of the present invention is made of a food composition, as an active ingredient, as well as a hole seaweed extract, it contains components that are commonly added during food production, for example, proteins, carbohydrates, fats, Nutrients, seasonings and flavoring agents. Examples of the above carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose, oligosaccharides and the like; And sugars such as conventional sugars such as polysaccharides such as dextrin, cyclodextrin and the like and xylitol, sorbitol, erythritol. As flavoring agents, natural flavoring agents (tauumatin, stevia extract (for example rebaudioside A, glycyrrhizin, etc.)) and synthetic flavoring agents (saccharin, aspartame, etc.) can be used. For example, when the food composition of the present invention is prepared with a drink, the citric acid, liquid fructose, sugar, glucose, acetic acid, malic acid, fruit juice, tofu extract, jujube extract, licorice extract, etc. You can.

The features and advantages of the present invention are summarized as follows:

(Iii) The present invention provides an antioxidant composition, a pharmaceutical composition, and a food composition for improving antioxidant, comprising an extract of pomeson seaweed as an active ingredient.

(Ii) The puncture seaweed extract in the present invention is characterized by exhibiting antioxidant activity through the effect of scavenging free radicals or free radicals.

(Iii) In addition, the present invention is based in the health functional food and pharmaceutical fields using natural extracts that can improve and improve the diseases caused by oxidative stress of free radicals, especially active oxygen free radicals, because of their excellent antioxidant activity. Provide historical data.

Figure 1 shows the results in ABTS, DPPH radical scavenging activity and reducing power activity of the oleracea extract of the present invention.

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these embodiments are only for describing the present invention in more detail and that the scope of the present invention is not limited by these embodiments in accordance with the gist of the present invention .

Example

Throughout this specification, "%" used to denote the concentration of a particular substance is intended to include solids / solids (wt / wt), solid / liquid (wt / The liquid / liquid is (vol / vol)%.

Materials and Methods

Of Agarum clathratum  Extract manufacturer

After measuring the dry weight of the hole seaweed (National Institute of Biological Resources, Korea), it was immersed in methanol, ground in a blender and extracted at room temperature for 24-48 hours. After filtering the extracted seaweed seaweed using Whatman No. 4 filter paper, methanol was concentrated under reduced pressure to measure the weight of the seaweed seaweed extract (Table 1). Thereafter, the perilla seaweed extract was placed in a container and refrigerated.

division Dry weight (g) Extract amount (g) One  19.74 2.69

ABTS (2,2'- azino - bis (3- ethylbenzthiazoline -6- sulphonic acid ) Radical  Cation scavenging activity

ABTS radical scavenging activity was measured using the characteristic that the ABTS radical (Sigma-aldrich, A3219) is colorless by an antioxidant having radical scavenging activity. Positive controls include butylated hydroxyanisol (Sigma-Aldrich, B1253), a synthetic antioxidant, and trolox (Sigma-Aldrich, 238813), a derivative of the natural antioxidant caffeic acid (Sigma-Aldrich, 60018), and tocopherol. Was used.

ABTS solution was prepared and used for the experiment. Potassium persulfate was added to a solution of ABTS at a concentration of 7 mM in tertiary distilled water at a concentration of 2.45 mM to be dissolved well, and then stored in a dark room at room temperature for 12 hours. 10 μl sample or positive control was added to 190 μl ABTS solution and reacted for 7 minutes, and then the absorbance was measured at UV-734 nm.

DPPH (2,2- 피덴 -One- picrylhydrazyl ) Radical  Scavenging activity

Synthesized DPPH (Sigma-aldrich, 257621) with nitrogen in the center is a chemically stabilized free radical, which encounters an anti-oxidant material and gives off electrons, which causes radicals to disappear and change color. Positive controls were butylated hydroxyanisol (BHA), caffeic acid (natural antioxidant), and trolox (a derivative of tocopherol).

For DPPH radical scavenging activity analysis, the DPPH reagent was dissolved in ethanol at a concentration of 6.3 mg / 50 mL to prepare a DPPH solution. 10 μl sample or positive control was added to 90 μl DPPH solution, and then reacted for 10 minutes, and the absorbance was measured at UV-517 nm.

Reducing power ( reducing power activity ) Measure

Reducing power by proton donation was measured by the antioxidant activity evaluation method which measures the ability to reduce Fe (III) to Fe (II).

As the absorbance value measured at 700 nm, it was confirmed that the antioxidant compound has a high reducing power. Positive controls were butylated hydroxyanisol (BHA), caffeic acid (natural antioxidant), and trolox (a derivative of tocopherol).

20 μl of 50 μl phosphate buffer (0.2M, pH 6.6), 50 μl potassium ferricyanide (1% potassium ferricyanide; Sigma-Aldrich, 702587), 20 μl of sample or positive control for 20 minutes at 50 ° C. After the reaction, 50 μl TCA (trichloroacetic acid, 10%; Sigma-Aldrich, T9159) was added and centrifuged at 3,000 × g for 20 minutes. 100 μl centrifuge supernatant, 100 μl tertiary distilled water, 20 μl ferric chloride (ferric chloride 0.1%; Sigma-Aldrich, 702587) were added, and the absorbance was measured at UV-700 nm.

DNA Single chain  Destruction( breakage Inhibitory activity

In the reaction system included, iron catalyzes the Fenton reaction to generate a hydroxy radical, a powerful oxidant. Therefore, compounds that inhibit the Fenton reaction can eliminate hydroxy radicals, or chelate highly reactive iron, preventing the Fenton reaction from proceeding. Fenton reaction inhibitory activity was used in the single strand breakage (SSB) method in the presence of iron and hydrogen peroxide. The amount of sample was used at a concentration of 0.5 mg / ml.

Mix 6 μl Tris-HCl buffer (10 mM), 4 μl FeSO ₄ (0.25 mM), 4 μl H ₂ O ₂ (1 mM), 4 μl DNA (supercoilded DNA from pBR322; Roche) and 2 μl sample at 25 ° C. After 30 minutes of reaction, 10 μl stop solution (4 M urea, 50% sucrose, 50 mM EDTA and 0.1% bromophenol blue) was added to stop the reaction. After the reaction was completed, 10 μl sample and 2 μl loading star (5: 1) were mixed well, and the result was confirmed by electrophoresis.

Experiment result

Table 2 shows the results of the antioxidant activity evaluation of the extract of persimmon seaweed. Hole seaweed showed similar activities to BHA and trolox used as positive controls in ABTS, DPPH radical scavenging activity and reducing activity.

division ABTS (%)
(1 mg / ml) DPPH (%)
(1 mg / ml) Reducing power
(5 mg / ml) Boiled seaweed
( Agarum clathratum ) 6 14 0.58 BHA 70 68 1.74 Trolox 70 71 1.81

In addition, the DNA single chain breakage inhibitory activity of the oleracea extract was examined, and the overall DNA single chain breakage inhibitory activity was shown.

Having described the specific part of the present invention in detail, it is apparent to those skilled in the art that such a specific technology is only a preferred embodiment, and the scope of the present invention is not limited thereto. Therefore, the substantial scope of the present invention will be defined by the appended claims and equivalents thereof.

Claims (10)

Agarum seaweed clathratum ) Antioxidant composition comprising the extract as an active ingredient.
The composition of claim 1, wherein the pore seaweed extract is an extract extracted by treating a polar organic solvent or a non-polar organic solvent.
3. The polar solvent according to claim 2, wherein the polar solvent is (i) water, (ii) alcohol, (iii) acetic acid, (iii) dimethyl-formamide (DMFO), (iii) dimethyl sulfoxide (DMSO) or a combination thereof. Composition.
The method of claim 2, wherein the nonpolar solvent is acetone, acetonitrile, ethyl acetate, methyl acetate, fluoroalkane, pentane, hexane, 2,2,4-trimethylpentane, decane, cyclohexane, cyclopentane, diisobutyl Ethylene, 1-pentene, 1-chlorobutane, 1-chloropentane, o -xylene, diisopropyl ether, 2-chloropropane, toluene, 1-chloropropane, chlorobenzene, benzene, diethyl ether, diethyl sulfide , Chloroform, dichloromethane, 1,2-dichloroethane, anneal, diethylamine, ether, carbon tetrachloride, THF or a combination thereof.
The composition of claim 1, wherein the composition has an activity of removing cationic radicals of ABTS (2,2'-azino-bis (3-ethylbenzthiazoline-6-sulphonic acid).
The composition of claim 1, wherein the composition has an activity of removing radicals of DPPH (2,2-diphenyl-1-picrylhydrazyl).
The composition of claim 1, wherein the composition has an activity of reducing Fe (III) to Fe (II).
The composition of claim 1, wherein the composition has an activity of inhibiting breakage of single stranded or double stranded nucleotides by oxidation.
Agarum seaweed clathratum ) Antioxidant pharmaceutical composition comprising the extract as an active ingredient.
Agarum seaweed clathratum ) Antioxidant food composition comprising an extract as an active ingredient.

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