JP5089025B2 - Antioxidant composition - Google Patents

Description

The present invention relates to an anti-oxidative pairs Narubutsu having antioxidant activity derived from animal natural extracts and vegetable natural extracts, and more particularly, for a variety of reactive oxygen species produced in vivo those related to effective antioxidative sets forming compound capable of exhibiting antioxidant capacity in vivo. In particular, the present invention eliminates oxidative stress caused by active oxygen generated in the human body to prevent lifestyle-related diseases in a broad sense, specifically, for prevention of diabetes-related diseases in diabetic patients whose body active oxygen is significantly high. it is intended to provide a novel anti-oxidative pairs Narubutsu that enables creation of Kyoseru antioxidant in foods and food materials.

  In recent years, in Japan, an increase in various adult diseases accompanying an increase in the elderly population and an increase in human and economic burdens related to medical care have been taken up as extremely serious problems. Since the aging of living organisms and the occurrence of diseases are inseparable, it is inevitable that the number of sick people will increase as the number of elderly people increases. Therefore, it is important to slow down the aging rate of humans as much as possible and suppress the occurrence of diseases.

  The first cause of human aging is that the number of divisional proliferation of the cells that make up the body is limited because of tissue regeneration. The ability to regenerate the tissue is reduced. And aging is accelerated | stimulated by the effect | action by the factor which injures a cell, for example, irradiation of an ultraviolet-ray or a radiation, a medicine etc., or the active oxygen which generate | occur | produces in the living body. In particular, active oxygen is inevitably generated in organisms that perform energy metabolism using oxygen, and in this sense, it can be said that our aging inevitably occurs. Therefore, it is important to suppress the harmful action of active oxygen generated in the body as a main measure for delaying aging and suppressing the occurrence of diseases associated with aging.

Until now, various antioxidants have been supplied to the market, but most of them have been prepared from a single component or a single raw material. However, there are various types of active oxygen generated in the living body. Specifically, for example, superoxide (O ₂ ⁻ ) generated from oxygen gas taken into the body, O ₂ ⁻ , superoxide. Hydrogen peroxide (H ₂ O ₂ ) is produced by an enzyme called dismutase (SOD), which further generates hydroxyl radicals (OH.) By the catalytic action of transition metals. In addition, leukocytes produce active oxygen in order to sterilize bacteria that have entered the living body. Among these, in addition to H ₂ O _2, there are hypochlorous acid radicals, nitrogen oxide radicals, and the like.

  On the other hand, various antioxidants are used to eliminate these active oxygens. Are these antioxidants effective for all these active oxygens, or are the active oxygen species that act? Limited? There hasn't been enough consideration. The present inventors have examined the action of antioxidants on the protein oxidative degradation action of various active oxygens. Antioxidants include types of active oxygens that they act on and are effective against all active oxygens in vivo. It has been shown that there are very few antioxidants.

  That is, peptides such as anserine, carnosine, reduced glutathione (GSH), sulfur-containing amino acids or amino acid analogs present in various animal living bodies are vitamin C (L-ascorbine) against hypochlorite radicals (ClO.). Acid, VC) has a strong antioxidant action against peroxynitrite radicals (ONOO.), And vitamin E (α-tocopherol, VE) has a strong antioxidant action on hydroxyl radicals (OH). In order to completely remove the active oxygen injury that occurs in the living body, it is confirmed that the combination and composition of antioxidants using these in combination is effective rather than using them alone. (See Patent Document 1).

  Until now, various antioxidants have been tested to prevent various diseases, particularly lifestyle-related diseases. For example, V.I. C, V.D. Polyphenols such as E, β-carotene and catechin have been tested. However, when we look at disease prevention with these antioxidants, for example, prevention of diseases such as cancer, there are not many findings that it was always effective. Conversely, there are many findings that there is no clinical effect (see Non-Patent Document 1), and antioxidants have not been effective in preventing diseases and aging as expected.

  These causes can be considered as follows. First, there are many types of active oxygen generated in the living body, and it is difficult to eliminate their harmful effects with a single antioxidant. Except for C, many of the antioxidants derived from other plants have phenolic groups in their skeletons and exhibit fat-soluble properties, and polyphenols are poorly absorbed from the human intestinal tract. In some cases, the amount absorbed can be 1/1000 or less of the amount taken orally. Therefore, even if extremely strong antioxidant activity is shown in the test tube, it is considered that many of them did not sufficiently exhibit the effect in vivo.

  In order to eliminate these drawbacks, in particular, select an antioxidant composition that can handle a wide range of reactive oxygen species, and a material that can sufficiently handle OH · generated in the process of energy metabolism. This is very important. As described above, the antioxidant peptides anserine, carnosine and sulfur-containing amino acids obtained from animal raw materials have a strong antioxidant action against ClO. C is a fat-soluble vitamin for ONO. E has a strong antioxidant action against OH.

  However, V. When comparing the antioxidant activity of E against OH · with other plant-derived antioxidants in terms of protein degradation inhibitory activity, 1/5000 of the carotenoids astaxanthin, 1/2000 to 1/1000 of the polyphenols catechin and quercetin, It was about 1/1000 to 1/500 of the caffeic acid analog (Table 1). From these facts, V.V. Even if E is excellent in absorbability in the intestinal tract and distributed throughout the body in the living body, in order to eliminate OH. However, adult V.D. E1 maximum daily intake is 300 mg; E. Overdose can be harmful, and E was oily, and had a problem in reducing antioxidant activity during storage and food processing.

Japanese Patent Application No. 2003-025210 J. et al. Kaikkonen et al., Free Radicals Research, 33, 329-340, 2000

Under such circumstances, the present inventors have conducted intensive studies in view of the above prior art, to develop a new anti-oxidative pairs Narubutsu capable of solving the problems of the prior art as the target In proceeding with V. As a result of searching for antioxidant materials with strong OH / eliminating ability to replace or enhance E, as described above, polyphenols such as catechin and quercetin, caffeic acid analogues such as curcumin and ferulic acid, etc. It was found that phenol compounds and carotenoids astaxanthin and β-carotene have a strong antioxidant action against OH · (FIG. 1).

These antioxidants having a strong OH-erasing action are V.V. Similar to E, it has the property of hardly exhibiting an antioxidant effect at a low physiologically regulatable concentration with respect to ClO · and ONOO ·, and ClO · eliminant anserine-carnosine and ONOO · V. It has been found that the combined use with C is also essential, and has led to the creation of the present invention. The present invention aims to provide a novel anti-oxidative pairs formed product can effectively exhibit antioxidant capacity in vivo to a variety of reactive oxygen species produced in vivo.

The present invention for solving the above-described problems comprises the following technical means.
(1) An antioxidant composition that exhibits antioxidant capacity in vivo against reactive oxygen species produced in vivo,
1) per 100 mg of a histidine-containing dipeptide or sulfur-containing amino acid or its analog as a hypochlorous acid-based active oxygen scavenger 2) Vitamin C (L-ascorbic acid or L-ascorbine as a peroxynitrite-based active oxygen scavenger) Acid salt) at least 20 mg, and 3) at least 2 mg for caffeic acid analogues as hydroxylated radical active oxygen scavengers, at least 10 mg for polyphenol compounds, and at least 1.5 mg for carotenoid compounds. The histidine-containing dipeptide as a hypochlorous acid-based active oxygen scavenger is a mixture of anserine and carnosine, and the sulfur-containing amino acid or its analog is reduced glutathione, cysteine, acetylcysteine. , Methionine, and alliin 1 Or more, and wherein the hypochlorite radical is a specific active oxygen species produced in vivo, that all the three hydroxyl radicals and peroxynitrite radical having an action to suppress in vivo Antioxidant composition.
(2) A mixture of anserine and carnosine contained in animal extracts obtained by hot water extraction of poultry, cattle, pigs, and migratory fish muscles as a histidine-containing dipeptide as a hypochlorous acid-based active oxygen scavenger. And the sulfur-containing amino acid or its analog is at least one of reduced glutathione, cysteine, acetylcysteine, methionine, and alliin obtained from an extract of the leeks or cruciferous plants. Oxidizing composition.
(3) The antioxidant composition according to the above (1), wherein the caffeic acid analog is extracted from a plant of the grass family or the grass family.
(4) The antioxidant composition according to (1), wherein the polyphenol compound is a catechin extracted from a plant of mulberry leaves or tea leaves and / or a quercetin extracted from a plant of the family Leek.
(5) The anti-carotenoid compound according to (1), wherein the carotenoid compound is astaxanthin extracted from fish meat or algae and / or β-carotene extracted from a plant of tomorrow, pumpkin, carrot, mugwort, perilla or kale. Oxidizing composition.

Next, the present invention will be described in more detail.
Using anti-oxidizing substance derived from the anti-oxidizing substances and plant extracts derived from animal extract, mutually reinforce their mutual disadvantage, completely specific three active oxygen generated in vivo the antioxidative set Narubutsu can be erased in order to configure in natural materials, were selected natural materials containing component as ClO · scavenger, OH · quencher and ONOO · scavengers described above. Anserine and carnosine, which have strong ClO-eliminating action, are abundant in fish extract of large migratory fish such as various meat extracts, chicken extracts, tuna, bonito and salmon. Abundant in cruciferous plant extracts such as. ONOO / V. C is abundant in cruciferous plant extracts such as citrus fruits and cabbage.

  On the other hand, carotenoids such as astaxanthin and β-carotene, which have a strong OH-erasing action, are abundant in carp extract and algae extract that are animal extracts in the former case, and in green and yellow vegetable extracts such as carrots in the latter case. Polyphenols such as catechin, epigallocatechin gallate, and quercetin are contained in plant extracts such as mulberry leaves, tea, and onions, respectively. Furthermore, in caffeic acid analogues with strong OH-erasing activity, curcumin is abundantly contained in turmeric extract, ferulic acid and the like are extracted from wheat and rice plants, for example, wheat young leaves and rice bran extract. . Actually, when these OHs were tested for OH · erasing ability, they showed strong antioxidant action against OH · (FIG. 2).

  There are no specific doses for administration or intake of these antioxidants to humans. In C, 60 mg or more and 2000 mg or less per day; In E, intakes of 3 mg or more and 300 mg or less per day are recommended. When these are used as a reference and from the test results of the present invention, the dosage or intake of various components is set as follows: It was.

  Anserine and carnosine, an anserine-carnosine mixture, or sulfur-containing amino acids, which are ClO · eliminating agents, have an antioxidant action that completely suppresses the proteolytic action of 10 mM ClO · at 5 mM concentration. On the other hand, 1 mM V.V. C or V. C sodium completely suppresses the oxidation action of ONOO. For 10 mM OH ·, 0.5 μM carotenoids, 2.5 μM caffeic acid analogs, or 2.5 μM polyphenols have the effect of completely suppressing the oxidation of OH ·. Indicated. Polyphenols have a problem of intestinal absorption, and OH · is easily produced quantitatively among the active oxygen species generated in the living body.

The concentration of the active oxygen generated in the body may vary with the physiological conditions, since it is estimated to reach the level of several 10μM~ number 100μM at most locally, antioxidative sets of the present invention Narubutsu When the minimum blood concentration of each antioxidant at the time of ingestion of 1 was standardized per 500 μM of ClO · eliminant anserine-carnosine mixture (approximately 600 mg intake calculated as blood volume of 5000 mL), ONOO · elimination In the agent, V.V. C is 200 μM (also about 90 mg as ingestion), and OH / eliminator is the amount of carotenoids at 0.5 μM (also as ingestion as the minimum blood concentration in consideration of the OH / generation ratio in vivo. 1.5 mg), caffeic acid analogs of 2.5 μM (also 5-10 mg), and polyphenols of 2.5 μM (also 4 mg) are considered to be able to sufficiently prevent the damaging action of each active oxygen. It is done.

  If the amount of each component is expressed per 100 mg of sulfur-containing amino acids such as an anserine-carnosine mixture of ClO / eliminator or reduced glutathione, the V.O. C is 20 mg or more, and in the case of OH / eliminator, carotenoids are 1.5 mg or more with the lowest blood concentration in consideration of intestinal absorption (M. Osterile et al., Journal of Nutritional Biochemistry, Vol. 11, 482-490). Page, 2000), caffeic acid analogs that have no particular problem with intestinal absorption are 2 mg or more, and polyphenols such as catechin are 1 mg or more, but considering intestinal absorption (F. Catterall et al., Xenobiotica, Vol. 33, 743-753, 2003), 10 mg of 10 times the amount is calculated as a desirable blending amount.

Antioxidative sets composition as the present invention constructed to improve these OH · erasing agent, anserine - carnosine mixture per 100 mg, V. C 20 mg, V.I. E20 mg has the same antioxidant activity as the composition of E20 mg, can be combined with a wide range of food material combinations from animal extracts to various plant extracts, preventing the harmful effects of active oxygen generated in vivo preparation of antioxidant foods and antioxidant of food material may found to be possible. In the present invention, the antioxidant of the food is intended to mean any type and form of the food, beverage, and food materials containing the anti-oxidation set Narubutsu, type of food, the whether the form like matter It is intended.

The antioxidative sets formed of the invention, the histidine-containing dipeptides or a sulfur-containing amino acids or their analogues as hypochlorite-based active oxygen scavenger, preferably, anserine and / or carnosine is incorporated, its Vitamin C as a peroxynitrite-based active oxygen scavenger is blended in an amount of 20 mg or more per 100 mg, and preferably as a hydroxyl radical active oxygen scavenger, ferulic acid, chlorogenic acid, cinnamic acid, caffeic acid, Caffeic acid analogs such as curcumin, polyphenol compounds such as catechins and quercetins, and carotenoid compounds such as astaxanthin and β-carotene are blended at a ratio of 1.5 mg or more.

In this invention, the said component is mix | blended as a natural raw material containing those components other than being mix | blended as those compounds, and it is also possible to mix and mix them arbitrarily. In the present invention, a specific anti-oxidant having an action of simultaneously suppressing in vivo the three types of hypochlorite radical, hydroxyl radical and peroxynitrite radical, which are specific reactive oxygen species produced in the living body. are those where there is significant technological significance in building the oxidation sets Narubutsu, these are simply not been gathered known antioxidants, and usually knowledge conventional antioxidants Nor is it easily predictable from the prior art. Antioxidative sets composition as the present invention has the essential element to be added to the specific ingredients in specific proportions, as long as it meets these requirements, be formulated with other optional ingredients, it can be arbitrarily designed depending on the intended use of the anti-oxidative pairs Narubutsu.

Next, test examples of the present invention will be described.
(1) Ovalbumin (Ovalbumin, hereinafter sometimes abbreviated as Ova) by three kinds of active oxygens Inhibiting action of various antioxidants against decomposition action As typical active oxygen produced in vivo, ClO. , OH · and NO ₃ · were prepared. Among these, ClO. Is a method of diluting sodium dichlorite in distilled water of pH 6.5, and OH. Is a Fenton method (B. Halliwell et al., Analytical Biochemistry, 165, 215-219). 1987) partially modified, and NO ₃ · was prepared by modifying the quenching flow reactor method (R. Radi et al., Journal of Biological Chemistry, 266, 4244-4250, 1991), respectively. did.

Next, Ova was dissolved in buffered saline, pH 7.2 to 2.5 mg / mL, and each concentration of antioxidant was added at a volume ratio of 10: 1 to the protein solution, Allow to stand at room temperature for 30 minutes, and then add active oxygen at each concentration in the same volume ratio of 10: 1 to the protein solution. ClO / oxidation is 37 ° C., 30 minutes, OH / oxidation is 37 ° C. The NO ₃ · oxidation was allowed to react at 37 ° C for 120 minutes for 60 minutes. After the reaction, each sample solution was electrophoresed with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and the gel stained with Coomassie Brilliant Blue R250 staining solution was subjected to densitometer to calculate the Ova decomposition inhibition rate. . Moreover, the exact decomposition | disassembly inhibitory rate applied the reaction sample liquid directly to GPC-HPLC of a TSKG-3,000SW column, and computed the proteolysis inhibition activity with the following formula | equation.

  The inhibitory action of various antioxidants against Ova oxidative degradation by the above three active oxygens is as shown in FIG. For ClO., Purified anserine-carnosine mixture derived from chicken extract completely prevents oxidative degradation by ClO. Slightly inhibitory activity was observed with C and β-carotene and epigallocatechin gallate (EGCG). However, the concentrations of β-carotene and EGCG are 20 μM and 500 μM, which are equivalent to more than 100 times the concentration that can be actually physiologically adjusted, and were found to be impractical.

  For OH ·, 5 mM V.V. To the same extent as E, 10 μM astaxanthin and β-carotene, 5 μM catechin, quercetin, EGCG, caffeic acid analogs curcumin, chlorogenic acid, and ferulic acid also completely blocked Ova oxidative degradation. For ONOO., 1 mM V.V. Except for C completely preventing its oxidative degradation, β-carotene, catechin and EGCG showed the activity of suppressing their oxidative degradation, and these concentrations were 10 μM, 500 μM and 500 μM, respectively. It exceeded the physiologically adjustable concentration. These degradation inhibitory activities are summarized in Table 1.

(2) Comparison of action of antioxidant component-containing natural extract against three types of active oxygen In the same manner as in the test method of (1) above, the inhibitory activity of various natural extracts against Ova oxidative degradation action was compared. The concentration of the natural extract tested was 5 mg / mL as a solid. The results of SDS-PAGE are as shown in FIG.

  There were three types of natural extracts that showed an inhibitory effect on the oxidative degradation of ClO. The chicken extract contains anserine and carnosine, the koji extract contains anserine, the cabbage extract contains sulfur-containing amino acids and V. C is included. For oxidative decomposition of OH, potato extract, carrot extract, tea extract, turmeric extract, cabbage extract show strong inhibitory action, followed by wheat young leaf extract, rice bran extract, onion extract, mulberry leaf extract The effect was shown. The inhibitory action of chicken extract and lemon juice was extremely weak. In addition to anserine, the koji extract contains astaxanthin, the carrot extract contains β-carotene, the tea extract contains EGCG, the turmeric extract contains curcumin, the cabbage extract contains sulfur-containing amino acids, In addition to C, catechins and caffeic acid analogs are contained.

Wheat leaf extract and rice bran extract contain caffeic acid analogues such as ferulic acid, onion extract contains quercetin, and mulberry leaf extract contains catechins and caffeic acid analogues.
Lemon extract showed the strongest inhibitory action against the oxidative degradation of ONOO. Next, the oxidative decomposition inhibitory action was also observed in the koji extract, onion extract, cabbage extract and chicken extract. The main components of these extracts are V.V. for lemon juice and cabbage. C and the like, and onions and mulberry leaves include V.V. In addition to C, reduced glutathione and catechins are included, and the chicken extract and koji extract contain various amino acids in addition to anserine and carnosine. These antioxidant activities are summarized in Table 2.

(3) Comparison of reactive oxygen scavenging action of each antioxidant Comparison of the strength of antioxidant activity of peptides and sulfur-containing amino acids with antioxidant activity against ClO. The strength of antioxidant activity of various antioxidants with activity was compared. When compared with the concentration of each antioxidant having a ClO · scavenging action that completely inhibits the protein oxidative degradation exhibited by 10 mM ClO ·, it is as shown in Table 3, and reduced glutathione, L-cysteine, N-acetyl- L-cysteine was strongest, followed by anserine and carnosine dipeptides, followed by alliin and methionine.

  Similarly, for antioxidants having antioxidant activity against OH ·, the concentrations of each antioxidant that completely inhibits the proteolytic action exhibited by 10 mM OH · are as shown in Table 4. It was. The carotenoids astaxanthin and β-carotene, the polyphenols quercetin, the catechin, the caffeic acid analogs curcumin, ferulic acid, and chlorogenic acid showed the strongest antioxidant effect against the oxidative degradation of proteins by OH. , Caffeic acid order, V. E had the weakest antioxidant activity among the tested.

(4) Impaired ability of human erythrocytes to pass through microchannels due to active oxygen and prevention effect by various antioxidants Microchannel (abbreviated as MC), a microvascular model, measures blood rheology, that is, microvascular circulation ability of blood. It is a device to do. Factors affecting blood rheology include diet and oxidative stress. In the present invention, the effect of three types of active oxygen produced in vivo on the ability of human erythrocytes to pass through MC was tested, and the effects of various antioxidants on the prevention of active oxygen damage were compared.

In the test method, healthy volunteer blood is separated into plasma, leukocyte and red blood cell fractions by Ficoll-Paque centrifugation (Pharmacia), and each blood fraction is analyzed by MCFAN KH-3A (channel width 7 μm, MC Laboratory). MC transit time was measured. As a result, leukocytes (the number of nucleated cells 2 × 10 ^{5) are obtained} by three kinds of active oxygens of ClO., OH. And ONOO.
Cells / mL), red blood cells (hematocrit 15%) were impaired in passage, but plasma (diluted 1: 1) was not affected at all by the passage ability (FIG. 3).

  Next, using red blood cells, the effect of preventing various antioxidants against MC passage disorder caused by various active oxygens was measured. One example is shown in FIG. 4, but in this test, purified extract-derived anserine-carnosine mixture (2.5 mM) as ClO · eliminator, EGCG (50 μM) as OH · eliminator, and ONOO · eliminator V. C (2.5 mM) was used. The antioxidant composition is a mixture of these three types. As shown in FIG. 4, erythrocyte passage obstruction due to various active oxygens was completely prevented by the respective antioxidants, and the combination of antioxidants resulted in complete inhibition of any active oxygen obstruction.

(5) Effect of reducing blood glycated hemoglobin concentration in diabetic model rats GK rats (Y. Ihara et al., FEBS Letters, 473, 24-26, 2000), which are diabetic model animals, were administered with an antioxidant alone. And the effect of combined administration of antioxidants were compared by reducing blood glycated hemoglobin (HbA1c) concentration. It is known that the mortality rate of cardiovascular disease associated with diabetes is highly correlated with the blood HbA1c concentration. When the HbA1c concentration is decreased by 0.1%, the mortality rate is decreased by 5%, 0.2 % Reduction in mortality is expected to be 10% (KT. Khaw et al., BMJ, 322, 1-6, 2001).

  As a ClO · eliminator, chicken extract-derived anserine-carnosine mixture 200 mg / kg, as an OH · eliminator, young wheat leaf extract (total amount of caffeic acid analog 10 mg / kg), as an ONOO · eliminator, V. C100 mg / kg was used, and an antioxidant composition containing all of them was orally administered to GK rats for 28 days, and the blood HbA1c concentration at the end of the administration was measured. As shown in FIG. 5, the results are as follows: a decrease of 0.3% with ClO · erasing agent alone administration, a reduction of 0.2% with OH · alone administration, a reduction of 0.3% with ONOO · erasing agent alone administration, In the case of a formulation containing all these antioxidants, the reduction is 0.4%, and the blood HbA1c concentration formed by the action of hyperglycemia and active oxygen is determined by the combination of three antioxidants. The best result.

(6) Human blood cholesterol lowering action It is known that an ingredient having an antioxidant action has an action of lowering blood cholesterol, but its action mechanism is various. For example, there are a mechanism that consumes cholesterol by promoting energy production from fat, a mechanism that disturbs intestinal absorption of cholesterol, a mechanism that inhibits cholesterol synthase, a mechanism that enhances the ability of macrophages to remove cholesterol. It is known that chlorogenic acid and ferulic acid, which are caffeic acid analogs of the OH • eliminator according to the present invention, also have an effect of lowering blood cholesterol (A. Herrera-Arellano et al., Phytmedicine, Vol. 11, pp. 561-566). 2004; HK Kim et al., Clinica Chimica Acta, 327, 129-137, 2003). However, since the cholesterol-lowering effect by oral administration of ferulic acid or chlorogenic acid is unknown, it was tested whether oral administration of the antioxidant beverage according to the present invention has a cholesterol-lowering effect.

  Antioxidant beverage prepared by the method of Example 5 above, that is, ClO · eliminator, chicken extract-derived purified anserine-carnosine 400 mg, OH · eliminator, rice bran-derived purified ferulic acid 20 mg, and ONOO · eliminator Mango juice (50 ml) containing 300 mg of vitamin C was allowed to be taken once a day for 4 weeks to 17 healthy male volunteers, and blood before and after drinking was collected to measure blood lipid content. The measured lipids were total cholesterol, HDL cholesterol, and neutral fat, and the amount of LDL cholesterol was calculated from the following formula. That is, LDL cholesterol = total cholesterol− (HDL + neutral fat × 0.2).

  The results are shown in FIGS. There were 12 healthy individuals whose blood total cholesterol concentration exceeded 200 mg / dL before the start of drinking, but the average blood total cholesterol ranged from 233.8 ± 19.9 mg / dL to 219.3 ± 24.2 mg. / DL (p <0.05). The number of bad cholesterol LDL before starting drinking exceeded 120 mg / dL was also 12, but the mean blood LDL value was 149.0 ± 16.4 mg / dL to 135.0 ± 16.4 mg / dL. (P <0.05). Neutral fat is 124.9 ± 57.1 mg / dL before drinking, and 91.6 ± after drinking when compared with 8 people with hypercholesterolemia whose blood cholesterol before starting drinking exceeds 220 mg / dL. The result decreased to 29.4 mg / dL (p <0.05).

  According to the present invention, (1) the active oxygen species generated in the living body can be roughly divided into three, and an antioxidant that effectively acts on these can be provided. (2) From the OH system generated from oxygen gas and leukocytes It is possible to identify and prepare antioxidants against the active oxygen of chlorinated and nitrogen oxides produced. (3) It is possible to provide antioxidant foods effective in suppressing the occurrence of aging and lifestyle-related diseases. The effect of.

  EXAMPLES Next, although this invention is demonstrated concretely based on an Example, this invention is not limited at all by the following Examples.

  Chicken extract 40L obtained by hot water extraction from 10 kg of whole chicken was concentrated under reduced pressure to adjust the solid content concentration to 20%. The content of the anserine and carnosine mixture in this concentrated chicken extract was 140 mg / g per solid. For 1200 g of concentrated chicken extract, 16.8 g of C (food additive), and concentrated wheat extract extracted from young wheat leaves by hot water extraction to adjust the solid content to 10% and caffeic acid analog concentration to 0.2% Add about 840 g of young leaf extract, add 60 g of indigestible dextrin, mix well, spray dry and add about 400 g of antioxidant-containing powder containing ClO · eraser, OH · eraser and ONOO · eraser. Was prepared. This powder had 84 mg of anserine-carnosine per gram, V.D. C. Containing 42 mg and caffeic acid analog 4 mg. This powder was confirmed to show a blood HbA1c concentration-reducing action in a test in the above-mentioned diabetes model rat.

  Add 15kg of water to 5kg of red crab from which internal organs and bones have been removed, boil at 100 ° C for 3 hours, homogenize with a mixer as it is, add protease at 0.2% concentration and hydrolyze at 45 ° C for 4 hours Then, a red yeast rice paste was prepared. 1 kg of concentrated whole chicken extract of Example 1, V. 60 g of C sodium, 100 g of tea leaf extract powder (catechin content 20%), 40 g of seasoning liquid, and 100 g of dextrin were added and freeze-dried to prepare about 2 kg of antioxidant-containing koji extract powder. To 1 g of the powder, 50 mg as anserine-carnosine, V.V. 25 mg as C, 1 mg astaxanthin as total carotenoid, and 10 mg catechin were contained. Further, the 1% powder solution completely suppressed the proteolytic action by the above three active oxygens.

  30 kg of water is added to 10 kg of pork, and the extract obtained by heating at 100 ° C. for 2 hours is filtered to remove the precipitate, and then concentrated under reduced pressure to prepare about 4 kg of concentrated pork extract with a solid concentration of 10%. did. To this, 1 kg of onion extract (solid content 10%), V. 100 g of sodium C and 100 g of wheat leaf extract powder (caffeic acid analog content 1.5%) were added and spray-dried to prepare about 600 g of anti-oxidant-containing stew pork extract powder. 1 g of the powder contains 85 mg of an anserine-carnosine mixture, 140 mg of C, 1 mg of quercetin as a polyphenol, and 2 mg of caffeic acid analog ferulic acid were contained. The 1% aqueous solution of the powder completely suppressed the proteolytic action by the above three active oxygens.

  The chicken extract is filtered through an ultrafiltration membrane (SEP-3013 manufactured by Asahi Kasei) with a molecular weight cut off of 3,000, the filtrate permeate is concentrated, and the anserine-carnosine mixture is purified by a cation exchanger chromatograph of Dowex 50WX8. did. In addition, the rice bran extract obtained by hot water extraction from rice bran is added with activated carbon for decolorization and filtration, and further concentrated to a solids concentration of 20% to obtain a concentrated rice bran extract containing caffeic acid analogues. Prepared. Furthermore, the grapefruit fruit was applied to a mixer, and the obtained fruit juice was filtered to remove insoluble matters. Per 100 mL of grapefruit juice, 500 mg of the purified anserine-carnosine mixture, 10 mL of rice bran extract (containing 30 mg of ferulic acid), V. An appropriate amount of C200 mg, sweetener 1 g and natural grapefruit flavor was added to prepare an antioxidant-containing soft drink. The 10-fold dilution of the soft drink completely prevented the proteolytic action of the three types of active oxygen.

  400 mg of purified anserine-carnosine mixture prepared by the method of Example 4 per 50 mL of mango juice, 20 mg of purified ferulic acid extracted from rice bran (manufactured by Tsukino Rice Fine Chemicals), and 300 mg of vitamin C (food additive) are added, About 3000 beverages containing antioxidants were prepared.

  As described above in detail, the present invention relates to an antioxidant composition, and according to the present invention, an antioxidant composition that effectively acts on a wide range of active oxygen species can be provided. Until now, foods containing a single antioxidant have been widely used for the purpose of preventing lifestyle-related diseases and aging. For example, arteriosclerosis, hypertension, cancer, various diseases associated with diabetes and diabetes. However, in the clinical trials for preventing lifestyle-related diseases and aging prevention of antioxidants that have been conducted so far, there is a reality that has not necessarily been sufficiently effective. There are many possible causes for this, but first, if the clinical effects obtained by inhalation in vitro or in animals are actually taken orally by humans, problems with intestinal absorption and biodistribution may occur. There was a problem. In particular, plant-derived polyphenols are applicable.

  Second, reactive oxygen in the living body that causes aging and lifestyle-related diseases is not a single one, but various types of oxygen are generated depending on human lifestyle and physiological conditions. . Conventionally, the strength of the activity of an antioxidant has been evaluated for active oxygen that is poorly related to the active oxygen generated in the living body, and the order of strength has been defined by that standard. However, as confirmed in the test examples of the present invention, the action of various antioxidants at physiologically adjustable concentrations has specificity for reactive oxygen species, and conventionally, the antioxidant activity is weak. It has been clarified that even those that have been evaluated exert a strong antioxidant action against specific active oxygen, and conversely, even those that have been evaluated as strong antioxidants have active oxygen that has a weak action. From these facts, it is considered that foods mainly composed of a single antioxidant could not sufficiently prevent various diseases caused by various active oxygens generated in the living body.

  In the present invention, the active oxygen species generated in the living body are classified into three categories, and the antioxidants for OH · system generated from oxygen gas, chlorine-based and nitrogen oxide-based active oxygen generated by white blood cells were specified. . As a result, peptides and sulfur-containing amino acids constituting the peptides are chlorinated active oxygen, plant-derived carotenoids, polyphenols and phenols having a hydrophobic group are OH-based, and vitamin C It became clear that each showed strong antioxidant activity against nitrogen oxide-based active oxygen. From these findings, it is important to formulate antioxidants that act effectively on a wide range of reactive oxygen species, and by actually blending antioxidants, the final glycation products in vivo caused by oxidative damage As a result, it was possible to provide an antioxidant food that is more effective than the conventional one for suppressing the occurrence of aging and lifestyle-related diseases. In addition, a more effective antioxidant food material can be produced by blending a natural material containing these antioxidant components.

FIG. 1 shows the inhibitory effect of various antioxidants on ovalbumin degradation by active oxygen. A is 10 mM ClO radical, B is 10 mM OH radical, and C is oxidative decomposition of ONOO radical. Symbols on each panel are as follows: R is untreated ovalbumin, C is oxidatively decomposed ovalbumin control, 1 is purified anserine-carnosine mixture, 2 is vitamin C, 3 is vitamin E, 4 is astaxanthin, 5 is β-carotene , 6 is catechin, 7 is quercetin, 8 is epigallocatechin gallate (EGCG), 9 is curcumin, 10 is chlorogenic acid, and 11 is a ferulic acid-added sample (each concentration is as shown in Table 1). FIG. 2 shows the inhibitory effect on the ovalbumin degradation action by active oxygen of various natural raw materials. A is 10 mM ClO radical, B is 10 mM OH radical, and C is oxidative decomposition of ONOO radical. Symbols on each panel are as follows: R is untreated ovalbumin, C is oxidatively decomposed ovalbumin control, 1 is chicken extract, 2 is lemon juice, 3 is rice bran extract, 4 is koji extract, 5 is carrot extract, and 6 is mulberry Leaf extract, 7 onion extract, 8 tea leaf extract, 9 turmeric extract, 10 cabbage extract, and 11 wheat leaf extract (each concentration is as shown in Table 2). FIG. 3 shows the ability of human blood components to pass through microchannels. A is ClO radical treatment, B is OH radical treatment, and C is ONOO radical treatment. The graphs in the figure are plasma, white blood cell, and red blood cell from the left. FIG. 4 shows the effect of various active oxygens and various antioxidants on human red blood cells on the ability to pass through microchannels. A is ClO radical treatment, B is OH radical treatment, and C is ONOO radical treatment. From the left of the graph in the figure, there are a control, ClO / erasing agent addition, ONOO / erasing agent addition, OH / erasing agent addition, and three kinds of antioxidants. FIG. 5 shows blood HbA1C concentration lowering action of various antioxidants in diabetes model GK rats. FIG. 6 shows the blood total cholesterol concentration of 12 healthy volunteers. FIG. 7 shows the blood LDL cholesterol levels of 12 healthy volunteers. FIG. 8 shows blood triglyceride concentrations of 8 high cholesterol volunteers.

Claims (5)

An antioxidant composition that exerts antioxidant capacity in vivo against reactive oxygen species produced in vivo,
1) per 100 mg of a histidine-containing dipeptide or sulfur-containing amino acid or its analog as a hypochlorous acid-based active oxygen scavenger 2) Vitamin C (L-ascorbic acid or L-ascorbine as a peroxynitrite-based active oxygen scavenger) Acid salt) at least 20 mg, and 3) at least 2 mg for caffeic acid analogues as hydroxylated radical active oxygen scavengers, at least 10 mg for polyphenol compounds, and at least 1.5 mg for carotenoid compounds. The histidine-containing dipeptide as a hypochlorous acid-based active oxygen scavenger is a mixture of anserine and carnosine, and the sulfur-containing amino acid or its analog is reduced glutathione, cysteine, acetylcysteine. , Methionine, and alliin 1 Or more, and wherein the hypochlorite radical is a specific active oxygen species produced in vivo, that all the three hydroxyl radicals and peroxynitrite radical having an action to suppress in vivo Antioxidant composition.   The histidine-containing dipeptide as a hypochlorous acid-based active oxygen scavenger is a mixture of anserine and carnosine contained in animal extracts obtained by hot-water extraction of poultry, cattle, pigs, and migratory fish muscles. The antioxidant composition according to claim 1, wherein the sulfur-containing amino acid or an analog thereof is at least one of reduced glutathione, cysteine, acetylcysteine, methionine, and alliin obtained from an extract of a leece family or a cruciferous plant.   The antioxidant composition according to claim 1, wherein the caffeic acid analog is extracted from a plant of the family of grasses or grasses.   The antioxidant composition according to claim 1, wherein the polyphenol compound is a catechin extracted from a plant of mulberry leaves or tea leaves and / or a quercetin extracted from a plant of the family Leek.   The antioxidant composition according to claim 1, wherein the carotenoid compound is astaxanthin extracted from fish meat or algae and / or β-carotene extracted from plants of tomorrow, pumpkin, carrot, mugwort, perilla or kale.