JP2002226368A - Inhibitor against oxidative damage to erythrocyte - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain an inhibitor against oxidative damage to erythrocytes, an inhibitor against hardening of erythrocytes, and a stabilizer of erythrocytes, a blood-preservative, and a food having activities for inhibition of the oxidative damage to, prevention of the hardening of, and stabilization of the erythrocyte. SOLUTION: The inhibitor against the oxidative damage to, the inhibitor against the hardening of, and the stabilizer of the erythrocyte, and the blood- preservative contain astaxanthin or an ester thereof as an active ingredient. The food having activities for the inhibition of the oxidative damage to, the food for the prevention of the hardening of, and the food for the stabilization of the erythrocyte contain the astaxanthin as an active ingredient.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to an erythrocyte oxidative damage inhibitor, an erythrocyte sclerosis inhibitor, an erythrocyte stabilizing agent and a blood preservative containing astaxanthin or an ester thereof as an active ingredient, and astaxanthin or an ester thereof. The present invention relates to a food having an effect of suppressing oxidative damage of red blood cells, a food having an effect of preventing sclerosis of red blood cells, and a food having an effect of stabilizing red blood cells.

[0002]

BACKGROUND OF THE INVENTION Blood is constantly exposed to oxidative stress, whether or not its presence is in vivo. That is, it is known that, in the stored state, lipid peroxide in blood increases due to oxidative stress, the antioxidant ability decreases, and hemolysis eventually occurs. Is constantly subjected to oxidative stress, and its membranes (red blood cells) are constantly at risk of hardening or breaking down and hemolysis. It is believed that the hardening and destruction of these films reduces oxygen supply to brain tissue and peripheral sites and contributes to reduced function of these tissues. Preventing the sclerosis and destruction of red blood cells can reduce arteriosclerosis, hypertension, diabetes, cancer, hyperlipidemia, rheumatism, gout, stroke, ischemic heart disease, emphysema, gastric ulcer, pancreatitis, nephritis, cataract, Alzheimer's disease,
It is considered to be useful for preventing or treating allergic diseases and diseases related to aging. Heretofore, it has been often reported that vitamin E protects erythrocytes from these oxidative stresses, and is already widely used in fields such as health foods. In recent years, astaxanthin having an antioxidant power even stronger than this vitamin E has been found, but there is no report on its protective effect on erythrocytes. Astaxanthin is a long-chain fat-soluble compound, and is expected to not only increase the fluidity of the membrane by being incorporated into the membrane structure of erythrocytes, but also protect and stabilize erythrocytes from oxidative stress. Astaxanthin or its ester, which is a compound derived from a natural product and has high safety, can be obtained from low-density lipoprotein (LD) in serum.
It has been reported that L) has the effect of preventing oxidation, preventing or suppressing arteriosclerosis, ischemic heart disease or ischemic encephalopathy (Japanese Patent Application Laid-Open No. H10-155559). However, there is no report that astaxanthin or its ester protects and stabilizes erythrocytes.

[0003]

DISCLOSURE OF THE INVENTION The present invention relates to an erythrocyte oxidative damage inhibitor, an erythrocyte sclerosis inhibitor, an erythrocyte stabilizing agent and a blood preservative comprising astaxanthin or an ester thereof as an active ingredient. It is an object of the present invention to provide a food having the action of suppressing oxidative damage to red blood cells, a food having an effect of preventing sclerosis of red blood cells, and a food having an action of stabilizing red blood cells, which contains the ester as an active ingredient.

[0004]

Means for Solving the Problems In order to solve the above-mentioned problems, the present inventors have searched for various compounds having an action of preventing oxidative damage to erythrocytes. It has been found that it has the effect of preventing mechanical damage. Further, they have found that astaxanthin or its ester is contained in foods and drinks and has an action of suppressing oxidative damage of red blood cells. The present invention has been made based on such findings.

That is, the invention according to claim 1 of the present invention is an oxidative damage inhibitor for erythrocytes containing astaxanthin or an ester thereof as an active ingredient, and the invention according to claim 2 comprises astaxanthin or an ester thereof as an active ingredient. The present invention according to claim 3 is an erythrocyte hardening inhibitor, the invention according to claim 3 is a red blood cell stabilizing agent containing astaxanthin or an ester thereof as an active ingredient, and the invention according to claim 4 is a blood preservative containing astaxanthin or an ester thereof as an active ingredient. The invention according to claim 5 is a food product having astaxanthin or an ester thereof as an active ingredient and having an oxidative damage inhibitory effect on erythrocytes, and the invention according to claim 6 is an erythrocyte containing astaxanthin or an ester thereof as an active ingredient The invention according to claim 7, which is a food having an anti-curing effect. Or is a food having a stabilizing effect of the red blood cells of the ester as an active ingredient.

[0006] In the present invention, "astaxanthin or an ester thereof" means a product derived from a natural product or obtained by synthesis. As those derived from natural products, for example, shrimp, krill, shells and eggs of crustaceans such as crabs, organs, various fish and shells, eggs, algae such as hematococcus, yeasts such as red yeast Phaffia, Examples include marine bacteria (Agrobacterium aurantiacum), or those obtained from seed plants such as Fukusou grass and Kimbong flower. Extracts from nature and chemically synthesized products are commercially available and easily available.

[0007] Astaxanthin or its ester is
For example, it can be obtained by culturing red yeast Phaffia, green alga Haematococcus, marine bacteria and the like in an appropriate medium according to a conventional method or a known method.

[0008] Various methods are known for extracting astaxanthin from the above culture or extracting and purifying from the above crustaceans. For example, since diester-type astaxanthin is an oil-soluble substance, an astaxanthin-containing component can be extracted from a natural product containing astaxanthin with an oil-soluble organic solvent such as acetone, alcohol, ethyl acetate, benzene, and chloroform. After the extraction, the solvent is removed according to a conventional method to obtain a diester-type astaxanthin concentrate. The concentrate obtained may be further purified if desired.

Astaxanthin includes 3,3′-dihydroxy-β, β-carotene-4,4′-dione or a stereoisomer thereof. Specifically, three stereoisomers of (3R, 3'R) -astaxanthin, (3R, 3'S) -astaxanthin, and (3S, 3'S) -astaxanthin are known, but the present invention is not limited thereto. Any of them can be used.

[0010] Astaxanthin or its ester is known to be a highly safe compound with no mutagenicity observed.

The agent for suppressing oxidative damage to erythrocytes containing astaxanthin or its ester as an active ingredient of the present invention, or the food or drink having the action of suppressing oxidative damage to erythrocytes containing astaxanthin or its ester as an active ingredient can be produced by releasing astaxanthin. Any of the compound, mono- and diester can be used. The diester has two hydroxyl groups protected by an ester bond, and therefore has higher physical stability than a free form or a monoester, and is less likely to be oxidatively decomposed in a preparation. However, it is considered that when taken into a living body, it is rapidly hydrolyzed to astaxanthin by an in-vivo enzyme to exhibit an effect.

The monoesters of astaxanthin include lower or higher saturated fatty acids or esters with lower or higher unsaturated fatty acids. Specifically, acetic acid, lauric acid, myristic acid, pentadecanoic acid, palmitic acid, palmitooleic acid, heptadecanoic acid, elaidic acid, ricinoleic acid, petroselinic acid,
Vaccenic acid, eleostearic acid, punicic acid, ricanoic acid, parinaric acid, gadoric acid, 5-eicosenoic acid, 5
-Docosenoic acid, cetolic acid, erucic acid, 5,13-docosadienoic acid, ceracholic acid, decenoic acid, sterling acid, dodecenoic acid, oleic acid, stearic acid, eicosapentaenoic acid, docosahexaenoic acid, linoleic acid, linolenic acid, arachidonic acid And the like. Examples of the diester of astaxanthin include diesters composed of the same or different fatty acids selected from the group of the above fatty acids.

Examples of astaxanthin esters include, for example, amino acid esters such as glycine and alanine, mono- or poly-carboxylic esters such as acetates and citrates and salts thereof, and phosphates and sulfates. Monoesters such as inorganic acid esters and salts thereof, sugar esters such as glucoside, sugar fatty acid esters, glycerosaccharide fatty acid esters, sphingose sugar fatty acid esters, glycerofatty acid esters, and glycerophosphate esters. it can. Or the above amino acids, carboxylic acids, phosphoric acids, sulfuric acids, sugars, unsaturated fatty acids, saturated fatty acids, polyunsaturated fatty acids, fatty acid esters, sugar fatty acid esters, glycerosaccharide fatty acid esters, sphingosaccharide fatty acid esters, glycerofatty acid esters Or the same or different diesters selected from glycerophosphate esters and the like.

Examples of the glycerophosphate diester include glycerophosphate saturated fatty acid esters and glycerophosphate esters containing fatty acids selected from highly unsaturated fatty acids, unsaturated fatty acids and saturated fatty acids.

Specific examples of astaxanthin diglycerophosphates include astaxanthin diglycerophosphate, astaxanthin glycerophosphate palmitic acid, astaxanthin glycerophosphatidylcholine palmitate, astaxanthin glycerophosphatidylcholine DHA, and astaxanthin glycerophosphatidyl inositol. Compounds such as palmitic acid, astaxanthin glycerophosphatidylinositol DHA, astaxanthin glycerophosphatidylinositol linoleic acid, and astaxanthin glycerophosphatidylcholine linoleic acid can be exemplified.

[0016]

The present invention will be described in detail with reference to the following reference examples and examples, but the present invention is not limited to these examples.

Example 1 Inhibitory effect of astaxanthin on rat erythrocyte membrane oxidative damage (i) Test method Astaxanthin or its ester is added to a rat erythrocyte suspension and incubated. Furthermore, when 2,2′-azobis (2-amidinopropane) dihydrochloride (hereinafter, AAPH), which is a free radical generator, is added, hemolysis occurs over time. It was investigated whether astaxanthin or its ester suppresses hemolysis. (Ii) Preparation of astaxanthin solution and control solution 6 mg of astaxanthin (manufactured by Sigma) was placed in a mortar, finely ground and mixed with 2 g of HCO-60 and a physiological saline solution.
00 ml. HCO-60 as control solution
The volume was adjusted to 10 ml using 0.2 g and a physiological saline solution. (Iii) Preparation of red blood cell suspension (RBC) Wistar male rat [Sankyo Lab Service Co., Ltd.]
Was bred for about one month after transport, and used after confirming that there was no abnormality in the general condition. Five animals are housed in wire mesh cages, temperature 22 ± 2 ° C, humidity 55 ± 10%, lighting 1
They were bred in a breeding room set for 2 hours (7 am to 7 pm). Chow and water were available ad libitum. The weight at the time of the experiment was 314 to 385 g. Blood was collected from the abdominal aorta under ether anesthesia and 1/10 volume of 2% EDTA-2K
To mix. After centrifugation at 1000 g for 10 minutes, the plasma and leukocyte layers were removed. Phosphate buffered saline (PBS: 125 mM sodium chloride, 10 mM sodium phosphate, pH 7.4) of 5 times the volume of the remaining red blood cells
Was added and mixed, and the supernatant was removed by centrifugation at 1000 g for 10 minutes. This operation was repeated three times to wash the red blood cells. Add 4 times the volume of red blood cells to PBS and add 20%
An erythrocyte suspension (RBC) was prepared.

(Iv) Measurement of the degree of hemolysis The experiment was performed in four groups. That is, astaxanthin was 0.1 μM, 0.5 μM, 1.0 μM, and 9 cases each in the control group (0 μM). Each liquid (ml) was placed in a test tube for each group as shown in Table 1 below.

[Table 1]

The test tubes were incubated at 37 ° C. for 30 minutes with shaking in the dark. Next, 1000g
And centrifuged for 10 minutes at +10 to remove the supernatant.
BS was added and the mixture was inverted, and then centrifuged in the same manner, the supernatant was removed, and the erythrocytes were washed. This washing operation was repeated three times. After washing, PBS was added to make up to 5 ml, 5 ml of 150 mM AAPH PBS solution was added,
Incubation was performed at 37 ° C. for 30 minutes with shaking in the dark. After incubation for 90 minutes, 0.1 ml was collected into two test tubes, one of which was added with 4 ml of PBS (solution A) and the other with 4 ml of distilled water for complete hemolysis (solution B). ). Both were centrifuged at 2000 g for 5 minutes, and the absorbance at 540 nm of the supernatant was measured. Hemolysis was calculated by the following equation. Hemolysis (%) = (absorbance of solution A / absorbance of solution B) ×
100

The effect of astaxanthin on oxidative damage to rat erythrocyte membrane is shown in Table 2.

[0021]

[Table 2] Each value is the mean ± standard deviation **: p <0.01 administration group vs. control group (t-test)

From the results in Table 2, it can be seen that astaxanthin inhibits hemolysis induced by adding AAPH to rat erythrocytes in a dose-dependent and statistically significant manner.

According to the present invention, it has been found that astaxanthin has an inhibitory effect on oxidative damage of erythrocyte membrane and stabilizes erythrocytes. That is, it is found that the composition is useful as an oxidative damage inhibitor for erythrocytes containing astaxanthin or an ester thereof as an active ingredient.

The oxidative damage inhibitor for erythrocytes of the present invention can be used for stabilizing erythrocytes during blood storage and for diseases caused by oxidative damage to erythrocytes, such as arteriosclerosis, hypertension, diabetes, cancer, It is useful as a preventive or therapeutic agent for hyperlipidemia, rheumatism, gout, stroke, ischemic heart disease, emphysema, gastric ulcer, pancreatitis, nephritis, cataract, Alzheimer's disease, allergic disease, aging and the like. It is also useful as a blood preservative.

The drug of the present invention may be prepared according to a conventional method using appropriate sugars such as lactose and saccharose, amino acids such as glycine, excipients such as cellulose, starch, gelatin,
Mixing binders such as methylcellulose and polyvinylpyrrolidone, or disintegrants such as starch and agar, or lubricants such as silicon dioxide, talc, magnesium stearate, and polyethylene glycol, flavoring agents, and sweeteners to form various formulations. Can be. For example, they are administered in solid dosage forms such as powders such as tablets, granules and fine granules, and liquid dosage forms such as elixirs, syrups and suspensions. The astaxanthin or ester used as an active ingredient may be a chemically synthesized product, an extract derived from a natural product or a crude extract, and these may be used alone or in a suitable mixture.

Astaxanthin or its ester is
It is susceptible to oxidative degradation by oxygen in the air, has poor physical stability such as temperature and light, and is decomposed and deactivated over time during the storage period when it is formulated. In order to suppress the decomposition of this astaxanthin or its ester, if necessary, a substance having antioxidant ability can be added to the composition as a stabilizer. For example, existing antioxidants such as vitamin A, vitamin B, vitamin C, vitamin E or vitamin derivatives thereof, cysteine, glutathione, glutathione peroxidase, citric acids, phosphoric acids, polyphenols, nucleic acids, herbal medicines, seaweeds, inorganic substances, etc. One or a mixture of two or more selected from the agents can also be added. In order to improve the absorption of astaxanthin alone or monoester, it is preferable to administer it in a fine powder state.

The amount of astaxanthin or an ester thereof used as a drug is expressed in terms of astaxanthin.
In an adult, the dosage is 0.5 mg to 100 mg, preferably 1 mg to 20 mg per day, orally or parenterally. The dosage depends on the age, weight,
It depends on the severity of the symptoms and the mode of administration.

The present invention also includes foods and drinks having astaxanthin or an ester thereof as an active ingredient and having the effect of suppressing oxidative damage to erythrocytes.

Examples of addition to foods and drinks include margarine,
Butter, butter sauce, cheese, fresh cream, shortening, lard, ice cream, yogurt, dairy products, sauce meat products, fish products, french fries, potato chips, popcorn, sprinkle, chewing gum, chocolate, pudding, jelly, gummy candy, Candy, drop, caramel, castella, cake,
Donuts, biscuits, cookies, crackers, etc.
Macaroni, pasta, salad oil, instant soup, dressing, eggs, mayonnaise, miso, etc., or non-alcoholic beverages such as fruit juice beverages, soft drinks, sports beverages, etc., non-alcoholic beverages such as tea, coffee, cocoa, etc. Examples of addition to general foods such as alcoholic beverages such as liqueurs and medicinal liquors can be given.

The food and drink of the present invention can be processed and manufactured by mixing astaxanthin or its ester together with the raw materials of general food and according to a conventional method. Although the amount of astaxanthin or its ester varies depending on the form of the food, it is generally 0.001% to 10%, preferably 0.0% to 10%.
It is adjusted to be 1 to 5% and contained in an amount necessary to exert the action of suppressing oxidative damage to the erythrocyte membrane. The amount used can be appropriately selected by those skilled in the art according to the type of food or drink.

When the food or drink of the present invention is used as a dietary supplement or a functional food, the form may be the same as the above-mentioned pharmaceutical preparation. It is also possible to use a mixture of milk protein, soy protein, egg albumin protein and the like, or their degradation products such as egg white oligopeptide, soy hydrolyzate and amino acid alone. In addition, processed foods such as natural liquid foods, semi-digestive nutritional foods and nutritional foods, drinks, capsules, and enteral nutritional supplements containing saccharides, fats, trace elements, vitamins, emulsifiers, flavors, etc. it can. When provided in a drink form, nutritional additives such as amino acids, vitamins, and minerals, sweeteners, spices,
Perfumes and pigments may be blended. The form of the food of the present invention is not limited to these.

[0032]

According to the present invention, an oxidative damage inhibitor for erythrocytes, an erythrocyte hardening inhibitor, a erythrocyte stabilizer and a blood preservative containing astaxanthin or an ester thereof as an active ingredient, and astaxanthin or an ester thereof as an active ingredient It is possible to provide a food having an action of suppressing oxidative damage to red blood cells, a food having an action of preventing hardening of red blood cells, and a food having an action of stabilizing red blood cells.
The erythrocyte membrane oxidative damage inhibitor of the present invention can prevent sclerosis and destruction of erythrocytes, enhance its stability in stored blood, and in vivo, atherosclerosis, hypertension, diabetes, cancer, and high fat. Pharmaceuticals for the prevention or treatment of diseases related to blood, rheumatism, gout, stroke, ischemic heart disease, emphysema, gastric ulcer, pancreatitis, nephritis, cataract, Alzheimer's disease, allergic disease, aging, or
It is useful as a material for functional foods.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) A61P 3/10 A61P 3/10 7/00 7/00 9/10 9/10 101 101 9/12 9 / 12 11/00 11/00 13/12 13/12 19/06 19/06 25/28 25/28 27/12 27/12 29/00 101 29/00 101 35/00 35/00 37/08 37 / 08 39/06 39/06 43/00 105 43/00 105 F term (reference) 4B018 MD08 ME06 4C206 AA01 AA02 CB25 MA01 MA04 NA14 ZA15 ZA16 ZA33 ZA36 ZA42 ZA45 ZA51 ZA59 ZA66 ZA68 ZA81 ZA96 ZB13 ZB15 ZB21 ZB26 ZC35

Claims (7)

[Claims] An erythrocyte oxidative damage inhibitor comprising astaxanthin or an ester thereof as an active ingredient. 2. An erythrocyte hardening inhibitor comprising astaxanthin or an ester thereof as an active ingredient. 3. An erythrocyte stabilizer comprising astaxanthin or an ester thereof as an active ingredient. 4. A blood preservative comprising astaxanthin or an ester thereof as an active ingredient. 5. A food containing astaxanthin or an ester thereof as an active ingredient, which has an effect of suppressing oxidative damage to red blood cells. 6. A food product having astaxanthin or an ester thereof as an active ingredient and having an effect of preventing erythrocyte hardening. 7. A foodstuff having astaxanthin or an ester thereof as an active ingredient and having an effect of stabilizing erythrocytes.