CN112402496A

CN112402496A - Antioxidant composition

Info

Publication number: CN112402496A
Application number: CN202010776171.9A
Authority: CN
Inventors: 今井理恵; 高桥香织
Original assignee: Fancl Corp
Current assignee: Fancl Corp
Priority date: 2019-08-23
Filing date: 2020-08-05
Publication date: 2021-02-26
Also published as: JP7334379B2; TW202114724A; JP2021031433A

Abstract

The present invention addresses the problem of providing an antioxidant composition containing an apple extract and a natural extract. To solve the problem, an antioxidant composition comprising the following components A and B as active ingredients is provided: component A: an apple extract; and B component: is selected from more than 1 of flos Granati extract, oriental cherry flower extract, fructus crataegi extract, flos Matricariae Chamomillae extract, and flos Pruni Pseudocerasi extract.

Description

Antioxidant composition

Technical Field

The present invention relates to a composition exhibiting an antioxidant effect.

Background

An apple extract (apple extract) formulated in food additives or health foods is rich in polyphenols, and has various physiological effects such as promotion of collagen production (patent document 1), promotion of blood circulation (patent document 2), and promotion of neutral fat metabolism (patent document 3).

In addition, it is known that it exerts a strong antioxidant action, reduces superoxide such as radicals accumulated in the living body, and inhibits aging (patent document 4). In recent years, the antioxidant effect and physiological action exhibited by apple extract have been attracting attention.

In addition, the antioxidant effect exhibited by natural extracts such as compositions derived from natural sources and apple extracts, rather than chemically synthesized compounds, is attracting attention. And various naturally derived compounds, extracts or compositions are known to exhibit antioxidant effects.

Patent document 5 exemplifies various substances as such natural antioxidants or extracts derived therefrom. For example, vitamin A, vitamin C, vitamin E, ubiquinone, inorganic selenium, manganese, melatonin, alpha-carotene, beta-carotene, lycopene, lutein, zeaxanthin (Zeanthin), Cryptoxanthin (Cryptoxanthin), resveratrol, eugenol, quercetin, catechin, gossypol, hesperetin, curcumin, ferulic acid, thymol, hydroxytyrosol, turmeric, thyme, olive oil, lipoic acid, glutathione, glutamine, oxalic acid, tocopherol, pectin, tocotrienol, coenzyme Q10, zeaxanthin, astaxanthin, canthaxanthin, saponins, limonoids, kaempferol, myricetin, isorhamnetin, procyanidins, rutin, lignans, apigenin, hesperetin, naringenin, eriodictyol, flavan-3-ols (e.g., anthocyanidins), catechins, gallo, galloquinones, quercetin, xanthophyllin, luteolin, and the like are described, Epicatechin and its gallate forms, epigallocatechin and its gallate forms (ECGC), theaflavin and its gallate forms, thearubigins, isoflavone phytoestrogens, genistein, daidzein, anthocyanins, cyanidin, delphinidin, malvidin, pelargonidin, peoniflorin, petunidin, ellagic acid, gallic acid, salicylic acid, rosmarinic acid, cinnamic acid and its derivatives (e.g., ferulic acid), chlorogenic acid, chicoric acid, gallotannins, ellagitannins, anthocyanins, beta-cyanines and other phytochromes, silymarin, citric acid, lignin, antinutrients, bilirubin, uric acid, R-alpha-lipoic acid, N-acetyl cysteine, emblicin (embrocinin), apple extract, apple peel extract (apple polyphenol), Red extract from lewis (Rooibos), green extract from lewis (Rooibos), hawthorn berry extract, red raspberry extract, green coffee bean antioxidant (GCA), 20% cranberry extract, grape seed extract (VinOseed), cocoa extract, hops extract, mangosteen fruit extract, mangosteen nutshell extract, cranberry extract, pomegranate rind extract, pomegranate seed extract, pomlla (Pomella) pomegranate extract, cinnamon bark extract, grape rind extract, bilberry extract, pine bark extract, pycnogenol, elderberry extract, mulberry root extract, wolfberry (Gogi) extract, blackberry extract, blueberry leaf extract, raspberry extract, turmeric extract, citrus bioflavonoids, blackcurrant, ginger, acai berry powder, green coffee bean extract, green tea extract, rose extract, and phytic acid.

The antioxidant effect of the combination of these antioxidant substances is considered to be merely a simple additive effect of the antioxidant effects of the respective substances.

Documents of the prior art

Patent document

Patent document 1: japanese patent laid-open publication No. 2012 and 62278

Patent document 2: japanese patent laid-open publication No. 2006 and 265220

Patent document 3: japanese laid-open patent publication No. 2006-151944

Patent document 4: japanese laid-open patent publication No. 10-75740

Patent document 5: japanese Kokai publication No. 2009-523407

Disclosure of Invention

As described above, although it is considered that only additive antioxidant effects can be exhibited, the present inventors have conducted preliminary experiments on the use of these natural antioxidant substances or plant extract, and found that only the combination of apple extract and specific natural extract can exhibit strong synergistic antioxidant effects, and further studied intensively, and as a result, completed the present invention.

That is, an object of the present invention is to provide an antioxidant composition containing an apple extract and a specific natural extract.

The main structure of the present invention is as follows.

(1) An antioxidant composition characterized by containing the following components A and B as active ingredients:

component A: an apple extract;

and B component: is selected from more than 1 of flos Granati extract, oriental cherry flower extract, fructus crataegi extract, flos Matricariae Chamomillae extract, and flos Pruni Pseudocerasi extract.

(2) The composition for antioxidation according to (1), characterized by containing 1 to 30 parts by mass of any one component selected as the component B in terms of dry mass relative to 1 part by mass of the dry mass of the component A.

(3) The composition for antioxidation according to (1) or (2), characterized in that the component A contains polyphenol.

(4) The composition for antioxidation according to (3), characterized in that the polyphenol content of the component A per dry mass is 50 mass% or more.

According to the present invention, a novel antioxidant composition is provided.

The composition of the present invention can inhibit the oxidation of body substances such as superoxide even in a small amount because the composition exhibits a synergistic effect of the antioxidant effect of an apple extract and the antioxidant effect of a specific flower extract and the like.

Thus preventing the occurrence of aging or diseases due to superoxide.

Drawings

FIG. 1 is a graph showing antioxidant effect of apple extract in combination with pomegranate flower extract at 1. mu.g/mL.

FIG. 2 is a graph showing antioxidant effect in apple extract 2. mu.g/mL in combination with pomegranate flower extract.

FIG. 3 is a graph showing antioxidant effect of apple extract 1. mu.g/mL in combination with cherry blossom extract.

FIG. 4 is a graph showing antioxidant effect of extract of fructus crataegi monogamae in combination with 1. mu.g/mL of extract of fructus Mali Pumilae.

FIG. 5 is a graph showing antioxidant effect of 1. mu.g/mL apple extract in combination with chamomile flower extract.

FIG. 6 is a graph showing antioxidant effect of apple extract in combination with white peach flower extract at 1. mu.g/mL.

Detailed Description

The present invention is an invention of an antioxidant composition, which is characterized by containing a component A: apple extract and component B: more than 1 selected from flos Granati extract, oriental cherry flower extract, fructus crataegi extract, flos Matricariae Chamomillae extract and flos Pruni Persicae extract as effective components.

The composition of the present invention can be used as a pharmaceutical product, health food, beverage, food, or food for oral administration or ingestion, and further can be used as a food additive.

The components contained in the composition of the present invention will be described below.

Component A:

examples of varieties of apple (Malus pumila) as a raw material of the apple extract include fuji, guguang, king forest, ruby, qiangan (Jonagold), marshal, san xia (Sansa), and qianqiu, but are not particularly limited thereto. The apple extract is not particularly limited, and examples thereof include fruits, leaves, trunks, flowers, and the like, and fruits are preferred. The fruit may be, for example, an unripe fruit (young fruit) or a well-ripe fruit, and is not particularly limited. The part of the fruit to be extracted is not particularly limited, and examples thereof include the whole fruit, pulp, pericarp, and seed. In the case of apple extract, these parts may be extracted alone or in combination of 2 or more.

The method for extracting the apple extract is not particularly limited, and conventionally known methods can be used. Specific examples of the extraction method are shown below.

First, the whole apple fruit is washed with water and then pulverized by a grinder or the like. The crushed product can be subjected to pectinase treatment, centrifuged, and subjected to partition filtration with an extraction solvent to prepare an apple extract. The pectinase treatment is not particularly limited, and, for example, 10 to 50ppm of pectinase may be added at a temperature of 20 to 60 ℃. The extraction solvent is not particularly limited, and examples thereof include organic solvents such as water, alcohol, hexane, and chloroform. Removing the extraction solvent to obtain the apple extract.

The apple extract can be obtained by extracting commercially available apple extract or apple fruit, and is not particularly limited.

The content of the apple extract contained in the composition of the present invention is not particularly limited, and as the apple extract, an apple extract containing 50 mass% or more of apple-derived polyphenols is preferable.

Apple extract can also be a material containing high concentrations of polyphenols. The method of separating the polyphenol by components is not particularly limited, and conventionally known methods can be used. The polyphenol can be separated into components by, for example, feeding an apple extract to a column, eluting the adsorbate of the column, distilling the eluted component under reduced pressure to remove it, and concentrating it. Further, the concentrated solution may be added with a powder aid to prepare a powder containing polyphenol at a high concentration by freeze-drying or spray-drying.

When a substance containing a high concentration of polyphenols derived from an apple extract is used as the apple extract of the present invention, for example, a commercially available substance sold as polyphenols derived from apples can be used. Examples of such commercially available products include "APPLE polyphenol APPL 'IN AFPOMM 9051" and "APPL' IN-POLYPHENOLIC APPLE EXTRACT POWDER", which are sold by UNITEC FOODS corporation.

And B component:

the flos Granati extract is flower extract of Punica granatum (Punica granatum). The variety of pomegranate in the present invention is not particularly limited.

The method for extracting the pomegranate flower extract is not particularly limited, and conventionally known methods can be used. Specific examples of the extraction method include the following methods. First, the collected pomegranate flower is washed with water and then pulverized by a grinder or the like. The pulverized product was used as an extraction raw material. The extraction solvent is not particularly limited, and examples thereof include water, alcohols, hexane, chloroform and other organic solvents. Then removing the extraction solvent to obtain the pomegranate flower extract.

The pomegranate flower extract can also be used, for example, as commercially available pomegranate flower extract. Examples of such a flower extract include "pomegranate flower extract powder: manufactured by Xiangrongxing corporation.

In the composition of the present invention, 1 to 30 parts by mass of the pomegranate flower extract is blended, preferably 2 to 18 parts by mass, and particularly preferably 2 to 10 parts by mass, based on 1 part by mass of the dried apple extract.

The oriental cherry flower extract is an extract of flower of Prunus (Prunus) belonging to genus Prunus of family Prunoideae of family Rosaceae. The species of oriental cherry in the present invention is not particularly limited.

The method for extracting the oriental cherry extract is not particularly limited, and conventionally known methods can be used. Specific examples of the extraction method include the following methods. First, collected oriental cherry flowers are washed with water and then pulverized with a grinder or the like. The pulverized product was used as an extraction raw material. The extraction solvent is not particularly limited, and examples thereof include water, alcohols, hexane, chloroform and other organic solvents. Then removing the extraction solvent to obtain the oriental cherry flower extract.

The oriental cherry extract can be used, for example, as commercially available oriental cherry extract. Examples of such a flower extract include "flower extract of oriental cherry flower-P: manufactured by Oryza oiled Co., Ltd.

In the composition of the present invention, 1 to 30 parts by mass of the oriental cherry flower extract is blended, preferably 2 to 18 parts by mass, and particularly preferably 4 to 10 parts by mass, based on 1 part by mass of the dried apple extract.

The fructus crataegi extract is extract of Crataegus Laevigata (Crataegus laevigata) or Crataegus monogyna (C. monogyna) belonging to Crataegus, and is also called Hawthorn (English). In the present invention, an extract extracted from the whole plant is used.

The method for extracting the extract of hawthorn fruit is not particularly limited, and conventionally known methods can be used. Specific examples of the extraction method can be carried out, for example, as follows. First, the whole collected individual hawthorn is washed with water and then pulverized with a grinder or the like. The pulverized product was used as an extraction raw material. The extraction solvent is not particularly limited, and examples thereof include water, alcohols, hexane, chloroform and other organic solvents. Then removing the extraction solvent to obtain the extract of fructus crataegi.

The extract of Crataegus monogyna can also be obtained, for example, from commercially available Crataegus monogyna. Examples of such extracts include "dry extract of hawthorn fruit GMP 133: ASK drugs "products.

In the composition of the present invention, 1 to 30 parts by mass of the extract of hawthorn fruit, preferably 2 to 18 parts by mass, and particularly preferably 2 to 4 parts by mass, is blended in terms of dry mass relative to 1 part by mass of the dry mass of the apple extract.

Chamomile extract is known by the name chamomile: an extract of Matricaria recutita. The extract extracted from the flowers of chamomile is used in the present invention.

The method for extracting the chamomile flower extract is not particularly limited, and conventionally known methods can be used. Specific examples of the extraction method can be carried out as follows. First, collected chamomile flowers are washed with water and then pulverized with a grinder or the like. The pulverized product was used as an extraction raw material. The extraction solvent is not particularly limited, and examples thereof include water, alcohols, hexane, chloroform and other organic solvents. Then removing the extraction solvent to obtain the chamomile flower extract.

As the extract of Matricaria chamomilla, for example, a commercially available extract of Matricaria chamomilla can be used. Examples of such an extract include "chamomile extract powder H: manufactured by japan powdered drug co, or "dried chamomilla extract GMP742 obtained by extracting the flower parts of chamomilla with hydrous ethanol: ASK drugs "products.

In the composition of the present invention, 1 to 30 parts by mass of the chamomile flower extract, preferably 2 to 20 parts by mass, and particularly preferably 5 to 20 parts by mass is blended in terms of dry mass with respect to 1 part by mass of the dry mass of the apple extract.

The white peach flower extract is extract of flower of white peach (Prunus persica Batsch).

The method for extracting the extract of white peach blossom is not particularly limited, and conventionally known methods can be used. Specific examples of the extraction method may be as follows. First, the collected flowers of white peaches are washed with water and then pulverized by a grinder or the like. The pulverized product was used as an extraction raw material. The extraction solvent is not particularly limited, and examples thereof include water, alcohols, hexane, chloroform and other organic solvents. Then removing the extraction solvent to obtain the white peach blossom extract.

The extract of white peach blossom can be used, for example, as a commercially available extract of white peach blossom. Examples of such extracts include "white peach blossom extract powder N: manufactured by Nippon powdered drug Co., Ltd. ".

In the composition of the present invention, 1 to 30 parts by mass of the white peach blossom extract is blended, preferably 2 to 18 parts by mass, and particularly preferably 4 to 10 parts by mass based on 1 part by mass of the dried apple extract.

When the composition of the present invention is formulated into a pharmaceutical product, a supplement, a beverage, a jelly or other food or beverage for ingestion, it is expected that the composition exerts an antioxidant effect in vivo by orally ingesting 1mg to 10g, preferably 1mg to 5g, particularly preferably 1mg to 3g per day.

The form of the food or beverage is not particularly limited. In addition, other additive components to be blended in the composition of the present invention when prepared into foods and beverages are not particularly limited.

Examples

< determination of antioxidant Effect >

The antioxidant effect of the composition of the present invention in vivo was confirmed by the ORAC method. ORAC was an evaluation method developed by the National Institute of Aging (National Institute on Aging) in 1992.

ORAC is a method of measuring the fluorescence intensity of fluorescein decomposed by using fluorescein as a fluorescent substance as a fluorescent probe in the presence of 2, 2' -azobis (2-methylpropylammonium) dihydrochloride (AAPH) as a radical initiator over time, and measuring the change as an index of the oxidation resistance. Since the rate of decrease in fluorescence intensity of fluorescein is delayed in the presence of the antioxidant substance in the reaction system, the antioxidant activity of the sample converted into the standard substance can be calculated by comparing the degree of delay in the rate of decrease of fluorescein in the presence of the standard substance. Accordingly, when the ORAC test shows a high value, the stability against oxidation can be said to be high.

Moreover, ORAC is considered to be useful as an index for evaluating the in vivo antioxidant ability of food.

1. Method for measuring antioxidation effect

The principle of measurement of ORAC is a method of measuring the change in the fluorescence intensity of fluorescein decomposed by the radical initiator AAPH over time using fluorescein as a fluorescent substance as described above as a fluorescent probe, and measuring the antioxidant capacity using the change as an index. Specifically, the fluorescence intensity (RFU) in the absence of Antioxidant substances was measured to prepare a change curve of fluorescence intensity (Blank), and then a change curve of fluorescence intensity (antitixidant) in the presence of Trolox standard solution or sample was prepared to calculate AUC of each curve. The AUC obtained by subtracting the AUC value in the absence of antioxidant substance from the AUC value in the presence of Trolox standard solution or sample is taken as the net AUC. Based on a calibration curve prepared by measurement of the Trolox standard solution, the Trolox Equivalent (TE) as a value converted into Trolox was calculated from the net AUC of the sample, and the antioxidant ability was evaluated based on the calculated value.

2. Test specimen

The composition to be measured was prepared from commercially available extract products described in table 1, in which the antioxidant effect was determined to have a synergistic effect in a screening conducted in advance.

[ Table 1]

Chamomile extract (W): camomile hot water extract camomile extract (E): 50% ethanol extract of chamomile

In addition, the apple extract shown in table 1 had a polyphenol content of 80 mass% per dry mass.

Among the reagent substances used in the test, Trolox ((±) 6-hydroxy-2, 5, 7, 8-tetramethylchroman-2-carboxylic acid) and Fluorescein (fluoroscein) were purchased from SIGMA-ALDRICH, AAPH was purchased from tokyo chemical industries co. Further, dipotassium hydrogen phosphate, potassium dihydrogen phosphate, dimethyl sulfoxide (DMSO), ethanol, and acetone were purchased from fuji film and mitsubishi corporation.

The experimental equipment is used: an microplate reader (INFINITE 200PRO, TECAN Japan K.K.), a 96-well plate (3881-096, IWAKI), and a multi-well plate sealing film (FG-BC100PCR, Japan Genetics).

3. Measurement method

(1) Preparation of test specimens

The samples of Table 1 above were dissolved in DMSO to 100mg/mL, respectively, and stored at-20 ℃. Then, 990. mu.L of phosphate buffer was added to 10. mu.L of the solution and dissolved to prepare a solution of 1mg/mL (1% DMSO). Further, the solution was diluted with a 1% DMSO phosphate buffer solution, and the following concentrations of the solution were prepared as test samples for measurement.

Component A: apple extract 1 μ g/mL and 2 μ g/mL

And B component:

pomegranate flower extract 8 μ g/mL

Oriental cherry flower extract 8 μ g/mL

Fructus crataegi extract 2 μ g/mL and 4 μ g/mL

The flos Matricariae Chamomillae extract is flos Matricariae Chamomillae extract (E)8 μ g/mL

Flos Matricariae Chamomillae extract (W)18 μ g/mL

8 mu g/mL of white peach blossom extract

In addition, the concentrations are based on the dry mass of the sample.

(2) Preparation of reagents

1) Trolox standard liquid

mu.L of 50mM Trolox, 45. mu.L of DMSO, 450. mu.L of phosphate buffer were added and a 500. mu.M solution of Trolox was prepared by dissolution. mu.L of 1% DMSO phosphate buffer solution was added to 160. mu.L of this 500. mu.M Trolox to dilute the solution, thereby preparing a 160. mu.M solution of Trolox. Hereinafter, a series (0, 6.25, 12.5, 25, 50 μ M, 1% DMSO phosphate buffer solution) diluted twice with 1% DMSO phosphate buffer solution was prepared as a standard solution for preparing a calibration curve.

2) Fluorescein

A1.2 mM solution was prepared by adding 100mL of phosphate buffer to 39.8mg of fluorescein. Next, 2mL of phosphate buffer was added to 10. mu.L of the 1.2mM fluorescein to prepare a 6.0. mu.M stock solution.

3)110.7nM fluorescein solution

The stock solution was diluted and dissolved by adding 25mL of phosphate buffer to 470. mu.L of 6.0. mu.M fluorescein stock solution.

31.7mM AAPH solution (prepared at the time of use)

To 129mg of AAPH, 15mL of phosphate buffer was added and dissolved.

(3) Measurement procedure

1) A 96-well plate was prepared, and the following measurement solutions containing the test samples prepared above were injected, respectively.

Trolox standard solution: 35 μ L

Measurement solution of component a: a mixed solution of 17.5. mu.L of apple extract solution and 17.5. mu.L of 1% DMSO/phosphate buffer solution as a test sample of the component A

Measurement solution of component B: one of 17.5. mu.L of pomegranate flower extract, cherry flower extract, hawthorn fruit extract, chamomile extract and white peach flower extract solution as the test sample of the component B and 17.5. mu.L of 1% DMSO/phosphate buffer solution

Measurement solution of component a + component B: mixture solution of apple extract solution 17.5 μ L as the test sample of the component A and one of pomegranate flower extract, cherry flower extract, fructus crataegi extract, flos Matricariae Chamomillae extract, and flos Pruni Pseudocerasi extract solution 17.5 μ L as the test sample of the component B

2) mu.L of a 110.7nM fluorescein solution was added and a multiwell plate seal was applied.

3) The 96-well plate was placed in a microplate reader, and the fluorescence intensity was measured at Ex485nm and Em538nm, followed by incubation at 37 ℃ for 10 minutes.

4) The enzyme-linked immunosorbent assay was performed again for Ex485nm and Em538nm, and the 96-well plate was removed.

5) Add 50. mu.L of 31.7mM AAPH solution, attach the sealing film to the multi-well plate again and place in the microplate reader. The measurement was carried out at 2-minute intervals for 90 minutes. The measurement conditions were as follows.

Setting a porous plate: temperature: at the temperature of 37 ℃ and the like,

wavelength: ex485nm/Em538nm,

stirring: for 10sec of the time period of the operation,

and (3) circulation: the number of the 45 times is 45,

spacing: the time for the reaction is 2min,

gain: and (5) manually operating by 100%.

4. Measurement and evaluation results of antioxidant Effect

The measurement results of Trolox equivalent as an antioxidant effect and the synergistic effects are shown in fig. 1 to 6 and tables 2 to 3 below. The representations of the graphs shown in fig. 1 to 6 are also summarized in parentheses in tables 2 to 3.

In the determination of the synergistic effect, a case where the calculated value of (measured value/theoretical value) × 100% is 120% or more is regarded as "present", and it is determined that the synergistic effect is obtained by the combination of the a component and the B component. The theoretical value is a total value of a measurement result of the Trolox equivalent of the component A alone and a measurement result of the Trolox equivalent of the component B alone, and shows an additive effect.

That is, the measured value is the measured value of a solution in which component B is mixed with component A

The theoretical value is the measured value of the a component alone + the measured value of the B component alone.

[ Table 2]

[ Table 3]

As shown above, it is clear that when the ratio of component a: when any one of the pomegranate flower extract, the cherry flower extract, the fructus crataegi extract, the chamomile flower extract and the white peach flower extract of the component B is used in combination with the apple extract, the antioxidant effect is enhanced in a synergistic manner.

Claims

1. An antioxidant composition characterized by containing the following components A and B as active ingredients:

component A: an apple extract;

2. The antioxidant composition as set forth in claim 1, wherein 1 to 30 parts by mass of any one component selected as the component B is contained in a dry mass basis relative to 1 part by mass of the component A in dry mass.

3. The composition for antioxidation according to claim 1 or 2, characterized in that the component A contains polyphenol.

4. The composition for antioxidation according to claim 3, characterized in that the polyphenol content per dry mass of the component A is 50 mass% or more.