JP2002029975A - Free radical scavenger - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a free radical scavenger with the combined use of vitamin C, synergistically enhanced in free radical scavenging effect compared to the case with the single use of eriocitrin (eriodictyol 7-rutinoside), thereby widely usable in foods/drinks, pharmaceutical preparations, cosmetics, etc. SOLUTION: This free radical scavenger is obtained by including, as active ingredients, eriocitrin (eriodictyol 7-rutinoside) and/or eriocitrin-containing substance and vitamin C.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a free radical scavenger and a composition containing the same, such as food and drink, cosmetics and pharmaceuticals.

[0002]

2. Description of the Related Art Active oxygen radicals, which are free radicals generated in vivo, are superoxide anions (.
O ₂ ^-), hydroxy radicals (· OH) and the like, are very rich material reactive, they can cause tissue damage such as damage cells and DNA, cancer, arteriosclerosis, various diseases such as aging It is said that he is greatly involved.
Those that have the effect of scavenging free radicals are thought to lead to the prevention of these diseases.

In such a situation, Japanese Patent Laid-Open No. 6-14 / 1994
No. 5062, an active oxygen free radical scavenger comprising a green tea leaf extract and a carotenoid pigment, and JP-A-9-208461, an active oxygen radical scavenger containing a lignan compound or an analog thereof as an active ingredient Has been proposed. Certainly, while these proposals are excellent, they are not entirely satisfactory and leave room for improvement.

[0004]

The present invention has been made in view of such circumstances, and has as its object to provide a new type of radical scavenger which is more effective.

[0005]

Means for Solving the Problems The present inventors have conducted intensive studies for a substance capable of scavenging radicals. As a result, eriocitrin, a flavonoid derived from citrus fruits, and its contents have a free radical scavenging effect. As a result of further research, the present inventors newly found that the combination of eriocitrin and / or its content with vitamin C synergistically enhanced radical scavenging activity, and completed the invention. Hereinafter, the present invention will be described in more detail.

[0006] Eriocitrin is a compound whose structure name is Eriodictyol 7-rutinos.
ide), which is a flavonoid glycoside in which rutinose (a disaccharide: glucose and rhamnose) is bound to eriodictyol belonging to flavanones among flavonoid compounds. Eriocitrin is found in citrus fruits, especially in lemons and limes, and also in oranges. It has never been known that eriocitrin has excellent antioxidant properties, and this is a novel finding discovered by the present applicant for the first time, and a novel invention relating to an antioxidant substance derived from citrus fruit based on this new finding. Japanese Patent Application Laid-Open No. 9-48969
No. has already been filed. However, eriocitrin and / or eriocitrin-containing substances and vitamin C
It has not been known at all that the radical scavenging effect is synergistically enhanced by combining.

In the present invention, as eriocitrin, not only purified eriocitrin itself but also eriocitrin-containing substances can be used. Examples of the former include citrus peels proposed in Japanese Patent Application Laid-Open No. 9-48969 filed by the present applicant,
A method for obtaining eriocitrin by performing a purification treatment such as resin treatment and preparative chromatography after extracting with water and / or an organic solvent, for example, a polar solvent, from the pericarp-containing material, fruit juice, and squeezed cake, After extracting with a polar solvent from citrus peel, juice and squeezed lees proposed in Japanese Patent Application No. 10-352197, the mixture is applied to a synthetic adsorption resin, heated with flowing hot water, eluted with 30% ethanol, and concentrated. The resulting eriocitrin-containing material can be used. If necessary, a paste or a dried product can be used. This eriocitrin-containing material contains flavonoids in addition to eriocitrin, and specific examples thereof include 6,8-di-C-glucosyldiosmin,
hesperidin, naringin and the like. In the former, the case where the purification treatment is not performed at all or only partially performed after the extraction is also included in the eriocitrin-containing extract.

[0008] Glucosyl diosmin includes 6,8-
Di-C-glucosyldiosmin (Diosmin-6,8-
C-di-β-D-glucopyranoside), 6-C-glucosyldiosmin (diosmin-6-C-β-D-glucopyranoside) and the like. The fact that glucosyl diosmin has an excellent antioxidant action was previously discovered by the present inventors and is not known before.

As the glucosyl diosmin, a pure product can be used, and the above-mentioned contents can be used. Further, for example, citrus juice, pericarp, and the like proposed in Japanese Patent Application Laid-Open No. Hei 10-245552 of the present applicant can be used. Extracting the pericarp-containing material and / or pomace with water, an organic solvent or a mixture thereof, and performing the obtained extract in combination with a purification treatment such as reverse-phase resin treatment and liquid chromatography. You may use what was manufactured by the manufacturing method of the antioxidant substance 6,8-di-C-glucosyl diosmin or 6-C-glucosyl diosmin.

As eriocitrin (similarly for glucosyl diosmin and other flavonoids), a chemically synthesized product or an extract can be used as a purified product by further purifying it. Refers to all substances containing eriocitrin,
The present invention also encompasses eriocitrin containing water and other liquids, solid carriers and the like. Therefore,
Eriocitrin-containing substances also include eriocitrin-containing materials such as lemon juice, orange juice, lime juice, and other citrus juices or concentrates thereof.

In carrying out the present invention, for example, in the case of flavonoid-containing substances, contained materials, extracts, etc., flavonoids are inevitably isolated and purified into respective components in view of economy and the like. In some cases, it is not necessary to use the flavonoids in a mixed state.

As vitamin C, any of those obtained by extracting from various raw materials such as citrus fruits and acerola, those obtained by fermentation sugar biosynthesis, and those obtained by chemical synthesis can be used, and commercially available products can be used as appropriate. It is. As the vitamin C, not only purified products but also crude products or vitamin C-containing substances can be used depending on uses such as food and drink.

In the free radical scavenger of the present invention, the content of eriocitrin and / or eriocitrin-containing material or vitamin C, and the combination thereof, can be appropriately used as appropriate. The content is preferably 0.00001 to 50% by weight, more preferably 0.0001 to 40% by weight, and particularly preferably 0.005 to 30% by weight. When the content of the active ingredient is lower than the above range, the effect is not sufficiently exhibited, and the content may be more than the above range.

The ratio of eriocitrin and the combination of eriocitrin-containing substance and vitamin C is 1: 2.
000 to 2000: 1 is preferable, and more preferably,
The ratio is 1: 100 to 100: 1.

[0015] In the present invention, the free radical, superoxide anion as the active oxygen free radicals generated in the body (· O ₂ ^-), hydroxy radicals (· OH), and a stable free radical 1,1
-Diphenyl-2-picrylhydrazyl (DPPH) and the like.

When used as a food or drink type composition, the present active ingredients, vitamin C and eriocitrin (or content), may be used as they are, or may be used in combination with other foods or food ingredients. It can be used according to the usual method. The composition according to the present invention using the present active ingredient may be in the form of a solid (powder, granule, or the like), a paste, a liquid or a suspension, and is used for producing sweeteners, sour agents, vitamins, and other drinks. Using various commonly used components,
It is suitable to be formulated into a health drink. At that time, it is particularly preferable that the active ingredient is blended at 0.1 to 0.0001%.

When used as a pharmaceutical type composition, the active ingredient is administered in various forms. Examples of the administration form include tablets, capsules, granules, powders,
Oral administration using a syrup or the like can be mentioned. These various preparations are commonly used in the pharmaceutical preparation technical field such as excipients, binders, disintegrants, lubricants, flavoring agents, solubilizing agents, suspending agents, coating agents, etc. in the main drug according to the usual method. It can be formulated using known adjuvants. The amount used depends on the symptoms, age, body weight, administration method, dosage form, etc., but usually about 0.1 mg to 1,000 mg can be administered to an adult once.

When used as a cosmetic-type composition, in addition to the active ingredient, various components generally used in cosmetics, quasi-drugs, and pharmaceuticals, as long as the effects of the present invention are not impaired, if necessary. , An aqueous component, a humectant, a thickener, a preservative, an antioxidant, a fragrance, a coloring agent, a drug, and the like. For example, solid or liquid paraffin, crystal oil, hydrocarbons such as ceresin, olive oil,
Vegetable or animal fats and waxes such as carnauba wax and lanolin, and further fatty acids such as stearic acid, palmitic acid and oleic acid or esters thereof, alcohols, polyhydric alcohols or esters thereof, nonionic surfactant Examples of the surfactant include surfactants such as anionic surfactants and cationic surfactants.

The amount of the active ingredient in the total effective skin external preparation (cosmetic) of the present invention is 0.005 as a dry product.
~ 30% by weight is preferred. If the compounding amount is less than 0.005% by weight, the radical scavenging effect is not sufficiently exhibited, and if the compounding amount is 30% by weight or more, the cost becomes high and it is not practical.

Eriocitrin and an eriocitrin-containing substance are components derived from a natural product, and eriocitrin is added to a citrus juice such as lemon or lime which has already been eaten in an amount of 10 mg / ml.
It is contained in a large amount of 100 ml, and there is no problem in safety to the living body. Mutagenicity tests by the Rec-assay method and the Ames method were negative, confirming safety.

Hereinafter, embodiments of the present invention will be described.

[0022]

Example 1 3 kg of lemon peel was pulverized with a homogenizer, 12 L of water was added, and the mixture was boiled for 2 hours. Thereafter, the extract was filtered to obtain a filtrate. This was concentrated under reduced pressure by an evaporator to obtain an extract. Next, it was dissolved in 200 ml of water, and the reverse phase resin column (4 cm × 50 cm, cosmos)
il 175C18-OPN; Nacalai Tesque, Inc.)
Shed. After dissolving in 1.5 L of water, the fraction eluted with 40% methanol was concentrated under reduced pressure, and the concentrate was dissolved in 5.1 of water. Eriocitrin was collected by preparative high performance liquid chromatography, and concentrated to dryness to obtain powder 5.0.
g (Eriocitrin; contained 90% or more). Using this, a sample aqueous solution used for the test was prepared.

Regarding the radical scavenging activity, DPPH (1,1-diphenyl-2-picrylhydrazyl) which is a stable radical
, The reduction rate of DPPH in each sample was quantified using HPLC to determine the radical scavenging activity. The measurement method was as follows.

100 mM Tris-HCl buffer (pH =
7.4) 1 ml of a 500 μM ethanol solution of DPPH and 200 μl of a sample solution were added to 800 μl, and the mixture was stirred well and allowed to stand at room temperature in a dark place for 20 minutes. The amount of DPPH in this solution was quantified using HPLC, and the DP
The radical scavenging activity was calculated by determining the DPPH scavenging rate when the PH amount was 100. HPLC analysis conditions were: detection wavelength: 517 nm, mobile phase: methanol 70%; water 30
%, Flow rate: 1 ml / min, column: Octyl-80
The test was performed under the conditions of Ts (4.6 mm x 150 mm) and a column temperature of 30 degrees. Based on the above method, the obtained eriocitrin was used to confirm the radical scavenging activity. The obtained results are shown in Table 1 below and FIG.

(Table 1) ──────────────────────────────── Each sample and final concentration Radical scavenging activity (%) ──────────────────────────────── Eriocitrin 20 ppm 31.0 Vitamin C 5 ppm 10.4 Eriocitrin 20 ppm 71.0 + Vitamin C 5ppm ────────────────────────────────

Thus, the radical scavenging activity of eriocitrin alone is 31.0%, and that of vitamin C alone is 11.0%.
It is 0.4%, which is 41.4% in total. By combining eriocitrin and vitamin C, the radical scavenging activity was 71.0%. That is, a very strong synergistic effect was observed by combining eriocitrin and vitamin C.

[0027]

Example 2 2 kg of lemon peel was crushed, 10 L of water was added, and the mixture was immersed at room temperature for 30 minutes, and then the extract was filtered to obtain a filtrate. This is centrifuged (9000 rpm, 2
0 min), and the supernatant is mixed with a synthetic adsorption resin (200 m
l; Amberlite XAD-16; manufactured by Rohm and Haas Company). Then, after flowing 1 L of water, 6
After flowing into 2 L of hot water at 0 ° C., 30% ethanol 1
L eluted. The eluate was concentrated to dryness to obtain 10.5 g of a lemon flavonoid-containing extract (Eriocitrin:
30%). Using this, a sample aqueous solution used for the test was prepared. The radical scavenging activity was confirmed in the same manner as in Example 1 using the obtained lemon flavonoid-containing extract. The results obtained are shown in Table 2 below and FIG.

(Table 2) ────────────────────────────────── Each sample and final concentration Radical scavenging activity ( %) レ モ ン Lemon flavonoid-containing extract 50 ppm 35.3 Vitamin C 5 ppm 10. 4 Lemon flavonoid-containing extract 50 ppm 69.7 + vitamin C 5 ppm ──────────────────────────────────

Thus, the radical scavenging activity of lemon flavonoid alone is 35.3%, and that of vitamin C alone is 10.4%, which is 45.7% in total.
By combining lemon flavonoid and vitamin C, the radical scavenging activity was 69.7%. In other words, by combining lemon flavonoids and vitamin C,
A very strong synergistic effect was observed.

[0030]

Example 3 1 g of the extract containing lemon flavonoid obtained in Example 2, 70 g of fructose, 2 g of citric acid, vitamin C
0.5 g and 1 g of lemon flavor were dissolved in 1 L of water to produce a beverage.

[0031]

Example 4 A free radical scavenger was produced by the following formulation. (1) A mixture of 4 g of the lyophilized eriocitrin obtained in Example 1 and 1 g of vitamin C, (2) 90 g of lactose, (3) 29 g of corn starch, and (4) 1 g of magnesium stearate. First, (1), (2),
(3) (but 17 g) was mixed and granulated together with the paste prepared from (3) (but 7 g). (3) (5 g) and (4) were added to the obtained granules and mixed well, and the mixture was compressed with a compression tablet machine to produce 100 tablets each containing 40 mg of the active ingredient.

[0032]

Example 5 A skin external preparation (cream) containing lemon flavonoid (Example 1) and vitamin C was prepared according to the following formulation.

(A. Oil phase) Stearic acid 10.0% Stearyl alcohol 4.0 Glycerin monostearate 8.0 Vitamin E acetate 0.5 Fragrance 0.4 Ethyl paraben 0.1 Butyl paraben 0.1 Propyl paraben 0.1 (b. Aqueous phase) lemon flavonoid 0.03 vitamin C 0.01 1,3-butylene glycol 10.0 propylene glycol 8.0 glycerin 2.0 potassium hydroxide 0.4 purified water residue

[0034]

According to the present invention, it has been found that eriocitrin and / or eriocitrin-containing substances (lemon flavonoids) have excellent radical scavenging ability, and as a result of further research, they were used in combination with vitamin C (ascorbic acid). By synergistically enhancing the radical scavenging ability for the first time, the present invention provides:
An effective radical scavenger can be obtained by combining vitamin C with eriocitrin or lemon flavonoid.

It is said that free radicals generated in the living body are greatly involved in diseases such as cancer, arteriosclerosis, and aging. For the present invention, preventive foods, drinks and health foods for these diseases are provided. Not only can it be used as medicines, but also can be widely used in cosmetics, toiletry products, industrial chemicals, etc.

[Brief description of the drawings]

FIG. 1 shows the radical scavenging activity in Example 1.

FIG. 2 shows the radical scavenging activity in Example 2.

Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat II (reference) A61K 7/00 A61K 7/00 H K31 / 375 31/375 35/78 K35 / 78 A61P 39/06 A61P 39 / 06 A23L 2/00 F (72) Inventor Yoshiaki Miyake 45-2, Kumanosho-ji, Shizukatsu-cho, Nishi-Kasugai-gun, Aichi Prefecture 45-2 Pokka Corporation R & D department F term (reference) 4B017 LC03 LG02 LK06 LK16 4B018 MD08 MD25 ME06 4C083 AA111 AA112 AB032 AC072 AC122 AC242 AC422 AC482 AC841 AC842 AD212 AD242 AD641 AD642 AD662 BB47 BB51 CC01 CC05 DD31 EE12 EE13 4C086 BA18 EA04 MA23 MA35 MA37 MA41 MA43 MA52 NA05 NA14 ZC28 ZC37 CA12 MA03 AB03CA03 MA41 MA43 MA52 NA05 NA14 ZC28 ZC37 ZC75

Claims (5)

[Claims] 1. A free radical scavenger comprising eriocitrin and / or an eriocitrin-containing substance and vitamin C as active ingredients. 2. A free radical scavenger composition comprising the free radical scavenger according to claim 1. 3. The composition according to claim 2, wherein the use of the composition is food or drink. 4. The composition according to claim 2, wherein the composition is used for cosmetics. 5. The composition according to claim 2, wherein the use of the composition is a pharmaceutical.