KR20210069810A - Pharmacology or food composition of antioxidant and anti-inflammatory activity on the aomponents of eggplants rind containing chlorogenic acid - Google Patents

Abstract

The present invention relates to a pharmaceutical or food composition having antioxidant and anti-inflammatory activity on eggplant peel components, and more particularly, by including an eggplant peel extract through an organic solvent as an active component, there is an effect of removing active oxygen and has an effect of protecting DNA damage, and thus it is possible to suppress the generation of NO that induces inflammation, and it can have the antioxidant enhancing effect the eggplant peel extract.

Description

A pharmaceutical or food composition having antioxidant and anti-inflammatory activity on eggplant peel components containing chlorogenic acid {Pharmacology or food composition of antioxidant and anti-inflammatory activity on the aomponents of eggplants rind containing chlorogenic acid}

The present invention relates to a pharmaceutical or food composition having antioxidant and anti-inflammatory activity against eggplant peel components, and more particularly, to a pharmaceutical or food composition having antioxidant or anti-inflammatory activity using an eggplant bark extract containing chlorogenic acid.

In general, oxygen is the most important molecule indispensable to sustain life. For example, brain cells begin to destroy if oxygen is not supplied for only 30 seconds. Mitochondria are the organs that play a key role in making energy in cells. Oxygen transported to the mitochondria produces energy by decomposing glucose made by humans ingesting nutrients (carbohydrates, proteins, and fats). Water, carbon dioxide and oxygen are generated in the energy production process, and this oxygen is called reactive oxygen species (ROS). Reactive oxygen species are generated during normal cell activity and are involved in a variety of biological processes, including cell differentiation, gene expression, and response to cytokines. Oxidative stress is a phenomenon in which ROS in cells increases due to the imbalance between the generation of ROS and the antioxidant reaction that removes it, which reacts with DNA, proteins, and lipids to damage it. This is known as a key cause of aging and heart-related diseases. .

Most ROS are produced as by-products between electron transport in mitochondria. In addition, ROS forms an intermediate necessary for metal-catalyzed oxidation reactions. The oxygen atom has two unpaired electrons in different orbitals of the outer electron shell. This electronic structure makes oxygen susceptible to radical formation. The sequential degradation of oxygen by the addition of electrons leads to the formation of ROS such as superoxide, hydrogen peroxide, hydroxyl radical, hydroxyl ion, and nitric oxide.

The detoxification of reactive oxygen species is the most important for the survival of all aerobic organisms. Many of the usual defense mechanisms have evolved to meet this need and provide a balance between the elimination and creation of ROS. The imbalance of ROS production and elimination is referred to as oxidative stress. Cells have a variety of defense mechanisms to ameliorate the deleterious effects of ROS. Superoxide dismutase (SOD) catalyzes the conversion of two superoxide anions inserted into _{hydrogen peroxide (H 2} O ₂ ) and oxygen (O _{2 ) molecules.}

2O ₂ + 2H → H ₂ O ₂ + O ₂

In eukaryotic peroxisomes, catalase enzymes _{convert H 2} O ₂ into water and oxygen, and complete the detoxification of free radicals initiated from SOD.

2H ₂ O ₂ → 2H ₂ O + O ₂

There are many non-enzymatic small molecule antioxidants that play an important role in this detoxification. Glutathione plays the most important intracellular defense against the harmful effects of reactive oxygen species. Reaction with ROS molecules oxidizes glutathione, but the reduced form is regenerated in redox by NADPH-dependent reductase. The reduced form of GSH as a percentage of the oxidized form of glutathione (GSSG) is a dynamic indicator of oxidative stress in organisms.

It is very important to maintain a healthy state at all times to maintain normal immune function related to inflammation. Although various immune-related cells and immunomodulatory factors are involved in the immune response, sufficient nutrients are required to function normally, and some immunomodulatory factors require environmental factors to significantly affect the expression of immune function. it is known The purpose of such inflammation is to remove necrotic cells and wounded tissues in the wounded area together with suppression of initial cellular damage and to regenerate tissues at the same time. Macrophages are known as major cells involved in the inflammatory response and are activated by cytokines secreted by immune cells or exposed to stimuli, and play an important role in body defense by producing cytokines at the early stage of infection. Activation of mitogen-activated protein kinase (MAPK) and NF-kB signaling pathways by toll-like receptor (TLR)-4 was inhibited when lipopolysaccharide (LPS), a component of the cell wall of Gram-negative bacteria, was treated in macrophages. induced

Recently, as it is recognized that the cause of adult disease and aging is due to ROS, research on the development of antioxidants known as substances that can control or remove oxygen-derived ROS has been actively conducted, and research on the development of many antioxidants has been conducted. has been reported However, among these antioxidants, α-tocopherol has a relatively low antioxidant effect, and synthetic antioxidants such as beta hydroxy acid (BHA) and dibutyl hydroxy toluene (BHT) have excellent antioxidant properties, so they are the most widely used phenol in commercial foods and pharmaceuticals. Although they are antioxidants, their use is gradually decreasing because of their mutagenicity and toxicity.

Therefore, research on the development of new natural antioxidants with excellent antioxidant power that can replace synthetic antioxidants by separating antioxidants from natural substances or medicinal plants, which are food sources that humans have eaten safely for a long time, is being actively conducted. Plants also produce a lot of reactive oxygen species, especially peroxides, in the process of photosynthesis, so it is expected that many enzymes that can protect against such oxidative damage and defense systems such as selectively developed secondary metabolites are well developed. Therefore, the plant itself can be an important resource containing antioxidants. Therefore, it is thought that the search for antioxidants for aging inhibition from plants will be meaningful.

From this point of view, studies on antioxidants of many plant resources and medicinal plants have been conducted on sesame, avocado, onion, cornflower, gold, mulberry, peony, shiho, eggplant, goji, yanggang, hyssop, psyllium, pogongyeong, hyangbuja, safflower seed, etc. Antioxidant studies have been reported.

ROS are generated during normal cell activity and are involved in a variety of biological processes, including cell differentiation, gene expression, and response to cytokines. Oxidative stress is a phenomenon in which ROS in cells increases due to an imbalance between the generation of ROS and the antioxidant reaction that removes it, which reacts with DNA to damage it, which is known as a key cause of aging and diseases. Research on the development of new materials using plums, natural products with antioxidant effects that remove these free radicals, is being actively conducted.

Eggplant (Solanum melongena L.) to be used in the present invention is one of the most produced cultivated vegetable crops in the world. It contains various phenolic compounds including nasunin, vitamin C, and chlorogenic acid, which are anthocyanins. Polyphenolic compounds have free radical scavenging activity of lipid peroxidation, and polyphenolic compounds extracted from loquat leaves, flavonoids and chlorogenic acid, are known to be very effective in antioxidant activity. In particular, in plant tissues, anthocyanins among polyphenolic compounds are known to have a function of blocking ultraviolet and visible light as an antioxidant. So far, studies using eggplant as a material have reported that it has anticancer effects, suppression of cardiovascular disease, and temporary improvement of hypercholesterolemia. In addition, chlorogenic acid, a major polyphenol component of eggplant, inhibits oxidation of low-density lipoprotein. has been reported to help.

Accordingly, the present inventors intend to provide a novel natural material with antioxidant and anti-inflammatory effects using by-products of plants. Currently, as interest in natural materials increases, research on eggplant is being conducted. There are no studies on the antioxidant and anti-inflammatory activity of eggplant bark and cedar bark produced in Korea.

Republic of Korea Patent Registration No. 10-1881016

1. Blois MA. (1958) Antioxidant determination by the use of a stable free radical. Nature. 181: 1199-1200. 2. Jeong CH, Choi GN, Kim JH, Kwak JH, Heo HJ, Shim KH, Cho BR, Bae YII, and Choi JS. (2009) In vitro antioxidant activities and phenolic composition of hot water extract from different parts of Cudrania tricuspidata. Journal of food science nutrition. 14: 283-289. 3. Dutta AK, Gope PR, Banik S, Makhnoon S, Siddiquee MA, Kabir Y. (2009) Antioxidant properties of ten high yielding rice varieties of Bangladesh. Asian Pacific Journal of Tropical Biomedicine. 2: 99-103. 4. Jung Y, Surh Y (2001) Oxidative DNA damage and cytotoxicity unduced by copper-stimulated redox cycling of salsolinol. a neurotoxic tetrahydroisoquinoline alkalooid. Free Radic Biol Med 30: 1407-1417

Therefore, the purpose of the present invention, which was devised to solve the above problems, is to provide a new antioxidant use of eggplant bark and cedar bark, which are by-products, away from the research focused on eggplant, extracting eggplant bark and measuring the chlorogenic acid content, To provide an antioxidant and anti-inflammatory composition of eggplant bark extract containing chlorogenic acid as an active ingredient by measuring various antioxidant activity and anti-inflammatory effects in macrophages of the extract.

In addition, the present invention provides an antioxidant and anti-inflammatory composition that measures the content of chlorogenic acid using an extract extracted from cedar bark, and improves antioxidant activity through the antioxidant effect of cedar bark extract and a mixture with eggplant bark.

Various problems to be solved in the present invention in the above are not limited thereto, but can be clearly understood by those of ordinary skill in the art to which the present invention pertains through the examples and claims to be described later. will be.

In order to achieve the above object, the present invention provides an antioxidant or anti-inflammatory composition comprising an eggplant bark extract.

Here, the composition is characterized in that it is a pharmaceutical or food composition.

In addition, the extract is characterized by reflux extraction using an organic solvent.

In addition, the organic solvent is characterized in that it is selected from the group consisting of water, an alcohol having 1 to 4 carbon atoms, or a mixture thereof.

In addition, the extract is characterized in that the concentration is 0.01 to 0.5 mg / ml.

In addition, the composition is characterized in that it has an active oxygen scavenging ability.

In addition, the composition is characterized in that it inhibits oxidative damage to DNA.

In addition, the composition is characterized in that it inhibits the generation of nitric oxide (NO) that induces inflammation in macrophages in mice.

In one aspect, the composition provides an antioxidant or anti-inflammatory composition further comprising a cedar bark extract.

Here, the cedar bark extract is characterized in that it is extracted with one organic solvent selected from water, an alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof.

In another aspect, (1) reflux extraction of the eggplant peel with an organic solvent to obtain an extract; (2) concentrating the extract under reduced pressure; and, (3) freeze-drying the extract concentrated under reduced pressure in step (2); It provides a method for preparing an antioxidant and anti-inflammatory composition comprising a.

Here, the preparation method, reflux extraction of the cedar bark with an organic solvent to obtain an extract, and preparing a mixture by mixing the cedar bark extract and eggplant bark extract; may further include.

According to the present invention as described above, eggplant bark extract containing chlorogenic acid as an active ingredient directly removes active oxygen species (DPPH, ABTS radical) and has a reducing power effect, and can remove active oxygen, oxidation Not only can it suppress DNA damage caused by stress, but it can also suppress inflammation by inhibiting NO, which causes inflammation in mouse macrophages.

In addition, by using the cedar bark extract containing chlorogenic acid mixed with the eggplant bark extract, it is possible to enhance the antioxidant activity effect such as the removal of active oxygen.

The effects according to the present invention in the above are not limited to the above, and other effects of the present invention can be clearly identified by those of ordinary skill in the art through the examples and claims to be described later. can be understood

1 shows the chlorogenic acid peak of the eggplant bark extract of the present invention.
Figure 2 shows the measurement result of the DPPH radical scavenging ability of the eggplant bark extract of the present invention.
3 shows the measurement results of the ABTS radical scavenging ability of the eggplant peel extract of the present invention.
Figure 4 shows the measurement result of reducing power (reducing power) of the eggplant peel extract of the present invention.
5 shows the experimental results of the protective effect of the eggplant bark extract of the present invention against oxidative DNA damage.
6 shows the experimental results for the inhibition of NO generation of eggplant peel extract of the present invention.
7 shows the measurement results of the DPPH radical scavenging ability of the mixture mixed with the bark extract of the present invention.
8 is a measurement result of the ABTS radical scavenging ability of the mixture mixed with the bark extract of the present invention;
9 shows the mass spectrometry peak result of the mixture mixed with the bark extract of the present invention.

Hereinafter, the present invention will be described in more detail.

In one aspect of the present invention, an antioxidant and anti-inflammatory composition of eggplant bark extract containing chlorogenic acid is disclosed.

In one embodiment of the present invention, an antioxidant and anti-inflammatory composition further comprising a cedar bark extract containing chlorogenic acid is disclosed. According to the present invention, an antioxidant enhancing effect of a mixture of eggplant bark extract and cedar bark extract is disclosed.

In a specific embodiment of the composition according to the present invention, the composition may be used as a pharmaceutical composition. It can be used as a pharmaceutical composition for the prevention or treatment of diseases caused by oxides produced by active oxygen, specifically cancer, arteriosclerosis, diabetes, stroke, myocardial infarction, hepatitis, nephritis, atopic dermatitis, Parkinson's disease, It is effective in the prevention or treatment of diseases caused by ultraviolet rays and radiation. The pharmaceutical composition may be administered through various routes including oral, transdermal, subcutaneous, intravenous or intramuscular during clinical administration, and may be applied as an external preparation for the skin. In addition, it can be formulated in various formulations, and in the case of formulation, it can be formulated using diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrants and surfactants. Solid preparations for oral administration include tablets, pills, powders, granules, and capsules, and such solid preparations include at least one excipient such as starch, calcium carbonate, sucrose, lactose, and gelatin in soybean extract and the like. prepared by mixing Liquid formulations for oral administration include suspensions, solutions, emulsions, and syrups. In addition to water and liquid paraffin, which are commonly used simple diluents, various excipients such as wetting agents, sweeteners, fragrances and preservatives are used. may be included. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations and suppositories. Non-aqueous solvents and suspensions may include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate. As the base of the suppository, Witepsol, macrogol, tween 61, cacao butter, laurin, glycerol and gelatin may be used.

When the composition of the present invention is used as a food additive, the eggplant peel extract may be added as it is or used together with other foods or food ingredients, and may be appropriately used according to a conventional method. The mixing amount of the active ingredient may be appropriately determined depending on the purpose of use (prophylactic, health, or therapeutic treatment), and may further include a food pharmaceutically acceptable food supplement additive.

The food composition of the present invention may include all foods in a conventional sense, and may be used interchangeably with terms known in the art, such as functional food and health functional food.

The term "functional food" as used in the present invention means a food manufactured and processed using raw materials or ingredients useful for the human body according to Health Functional Food Act No. 6727, and "functionality" refers to the structure of the human body. And it refers to ingestion for the purpose of obtaining useful effects for health purposes such as regulating nutrients for function or physiological action.

In addition, the term "health functional food" as used herein refers to a food manufactured and processed using a specific ingredient as a raw material for the purpose of health supplementation or by extracting, concentrating, refining, or mixing a specific ingredient contained in the food raw material, It refers to food designed and processed to sufficiently exert biological control functions such as biological defense, regulation of biological rhythm, prevention and recovery of disease, etc., by the above ingredients, and the composition for health food is used for prevention of diseases and prevention of diseases. It can perform functions related to recovery, etc.

There is no limitation on the kind of food in which the composition of the present invention can be used. In addition, the composition comprising the eggplant bark extract of the present invention as an active ingredient can be prepared by mixing other suitable auxiliary ingredients that may be contained in food and known additives according to the selection of those skilled in the art. Examples of foods that can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gums, dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages and There are vitamin complexes and the like, and it can be prepared by adding the extract according to the present invention as a main component to juice, tea, jelly, juice, and the like.

In addition, foods that can be applied to the present invention include, for example, special nutritional foods (eg, formula milk, infant food, etc.), processed meat products, fish meat products, tofu, jelly, noodles (eg, ramen, noodles, etc.), health supplements , seasoned foods (eg soy sauce, soybean paste, red pepper paste, mixed soy sauce, etc.), sauces, sweets (eg snacks), dairy products (eg fermented milk, cheese, etc.), other processed foods, kimchi, pickled foods (various kimchi, pickles, etc.) ), beverages (eg, fruit, vegetable beverages, soy milk, fermented beverages, etc.), and natural seasonings (eg, ramen soup, etc.).

When the health functional food composition of the present invention is used in the form of a beverage, it may contain various sweetening agents, flavoring agents, or natural carbohydrates as additional ingredients, as in a conventional beverage. In addition to the above, the health functional food composition of the present invention includes various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin , alcohol, a carbonation agent used in carbonated beverages, and the like. In addition, it may contain the pulp for the production of natural fruit juice, fruit juice beverage and vegetable beverage.

The method for preparing a composition for antioxidant and anti-inflammatory of eggplant peel extract containing chlorogenic acid according to one embodiment of the present invention is (1) the eggplant peel of 'black rice', an improved eggplant variety in Japan, grown on a farm in Gunwi-gun, Gyeongbuk separating and preparing for extraction; (2) reflux extraction of eggplant skins with an organic solvent to obtain an extract, followed by filtering; (3) concentrating the extract under reduced pressure; (4) freeze-drying the extract concentrated under reduced pressure in step (3); In step (2), reflux extraction can be performed with 70% ethanol of a volume corresponding to 30 times (wt/v%) of eggplant skin, and concentration under reduced pressure at 40° C. in step (3) can be performed, The freeze-drying in step (4) may be performed at -80°C to -110°C.

In addition, the eggplant bark extract may be mixed with the cedar bark extract, preferably by reflux-extracting the cedar bark with an organic solvent to obtain an extract; and mixing the cedar bark extract and the eggplant bark extract in a volume ratio of 2:1, 1:1 or 1:2 to prepare a mixture.

Hereinafter, the present invention will be described in more detail through examples. These examples are only for illustrating the present invention, and it will be apparent to those of ordinary skill in the art that the scope of the present invention is not to be construed as being limited by these examples.

<Example 1> Preparation of eggplant peel ethanol extract

The peel was separated from the commercially available eggplant and used. Extracts were obtained by reflux extraction 3 times at 25°C for 2 hours each with 70% ethanol in a volume corresponding to 30 times (wt/v%) of the separated eggplant skins. The obtained extract was filtered twice under reduced pressure using filter paper (adventec Toyo 2, Japan). The ethanol extract was concentrated under reduced pressure at 40°C or lower using a rotary evaporator (RV10D, Ika), and the concentrate was freeze-dried at -80°C to prepare it. The final yield is 39.71%.

<Example 2> Analysis of components of eggplant peel extract chlorogenic acid (HPLC)

Measurements were made using High Performance Liquid Chromatography (HPLC). Syncronis C18 Colimns, 5 um, 4.6 mm, 250 mm [Thermo Fisher Scientific, Massachusetts, USA] column was used, A (10% MeOH: water (0.1% with formic acid = 1:1 (v/v)), B (water(ph3.5): MeOH: ACN = 20: 20: 60 (v/v)) ratio was maintained. Flow rate was maintained at 0.3 mL/min, Injection volume was 5 μl, Fluorescence Detector (Ex : 280 nm, Em: 325 nm).

As shown in Table 1, the chlorogenic acid content and yield of the eggplant bark ethanol extract extracted in Example 1 was 39.71%. The chlorogenic acid content was measured to be 24.6 mg/g (extract). Eggplant skins were found to contain chlorogenic acid. The peak as a result of HPLC measurement of eggplant peel is shown in FIG. 1 .

Extract chlorogenic acid
(mg/g of extract) Yield (%) eggplant peel 24.6 39.71

<Example 3> DPPH radical scavenging activity (scavenging activity)

DPPH radical scavenging activity was carried out by modifying the experimental method of Blois (Blois MA. 1958). After preparing the eggplant bark ethanol extract extracted in Example 1 to a concentration of 10 mg/ml, it was diluted to a concentration of 0.01, 0.05, 0.1, 0.25, and 0.5 mg/ml to prepare 1 ml each, and ascorbic acid used as a standard 10 mg/ml was diluted with 1 ml of DW to prepare 1 ml at a concentration of 2 mg/ml. After dispensing 80 μl each of eggplant bark extract and ascorbic acid by concentration into a 96 well plate, 80 μl each in the order of 70% EtOH, methanol, and DPPH, wrap it in foil to prevent it from reacting with light, and then at room temperature for 30 minutes. After standing for a minute, absorbance at 517 nm was measured using an ELISA reader. The formula for calculating the DPPH radical scavenging ability is as follows.

Scavenging activity (%) = (1- Absorbance of sample / Absorbance of control) ×100

The results are shown in FIG. 2 . Referring to FIG. 2 , it can be seen that the 70% ethanol extract of eggplant bark has a better DPPH radical scavenging activity compared to the standard ascorbic acid.

<Example 4> ABTS radical scavenging activity (scavenging activity)

ABTS radical scavenging ability was measured by modifying the method of Jeong (Jeong CH. 2009) et al. After preparing the eggplant bark ethanol extract extracted in Example 1 to a concentration of 10 mg/ml, it was diluted to a concentration of 0.01, 0.05, 0.1, 0.25, and 0.5 mg/ml to prepare 1 ml each, and Ascorbic acid used as a standard 10 mg/ml is diluted with 1 ml of distilled water to prepare 1 ml at a concentration of 2 mg/ml. After mixing 7 mM ABTS and 2.45 mM Potassium persulphate dissolved in distilled water, it is left in the dark for 12 - 16 hours for stabilization. This solution can be used for 2 days at room temperature. The above mixture was diluted with PBS (0.01 M, pH 7.4) to 0.70 ± 0.02 at 734 nm. Then, 0.2 ml of eggplant bark extract and ascorbic acid standard were mixed with 0.8 ml of ABTS + solution, and reacted at room temperature for 5 minutes, and then the blank was measured at 734 nm. In the control group, a solvent was used instead of the extract of the sample, and the scavenging activity of ABTS radicals was calculated as follows.

Scavenging activity (%) = (1- Absorbance of sample / Absorbance of control) ×100

The results are shown in FIG. 3 . Referring to FIG. 3 , it can be seen that the 70% ethanol extract of eggplant peel has a superior ABTS radical scavenging activity compared to the standard Ascorbic acid.

<Example 5> Reducing power

The reducing power measurement was carried out with a slight modification of the method described by Dutta (Dutta AK. 2009). Eggplant peel extract of various concentrations at 0.01, 0.05, 0.1, 0.25, 0.5 mg/ml, 0.5 ml of standard Ascorbic acid (2 mg/ml), 0.5 ml of Sodium Phosphate buffer (PBS) and 0.5 ml of 1% Potassium Ferricyanide After mixing each, the mixture was left at 50°C for 20 minutes, and then 1 ml each of the reaction mixture was transferred to a new tube, 0.5 ml of Trichloroacetic acid (TCA) was added, and centrifuged at 3,000 rpm in a centrifuge at 4°C for 10 minutes. separated. After adding 0.5 ml of DW and 0.1 ml of ferric chloride to 0.5 ml of the centrifuged supernatant, aliquot 0.15 ml each in a 96 well plate, and then measure the absorbance at 700 nm using an ELISA reader. The measured values are shown in a graph.

The results are shown in FIG. 4 . Referring to FIG. 4 , it can be confirmed that the 70% ethanol extract of eggplant peel has an excellent reducing effect compared to the standard Ascorbic acid.

<Example 6> DNA protection activity (protection activity)

Prevention of oxidative damage to DNA caused by hydrogen peroxide (H ₂ O ₂ ) was carried out by slightly modifying the experimental method of Jung and Surh (Jung Y, Surh Y 2001). 1 μl of plasmid PBR 322 DNA (0.5 μg/μl) is a mixture of 2 μl (0.08 mm) _{of FeSO 4 , 3 μl of 30% H 2} O ₂ 3 μl (v/v) distilled water and 2 μl of extracts of different concentrations was reacted at 37 °C for 1 hour. After that, 2 μl of 6X DNA loading dye was added. It was reacted with 0.8% agarose gel electrophoresis at 70 V at room temperature. DNA bands (supercoiled, linear and open circular) were stained with ethidium bromide, and the effect of inhibiting DNA oxidation was evaluated by increasing and decreasing the supercoiled monomer based on the control value.

The results are shown in FIG. 5 . Referring to FIG. 5 , as a result of the protective effect of the eggplant bark extract against DNA damage, the super-helical DNA was completely transformed into a linear form due to DNA damage compared to the plasmid DNA control (Lane 2). When DNA was treated with hot water extract and 70% ethanol extract, DNA of Supercoiled was restored in a dose-dependent manner (Lanes 3 - 5). As a result, all of the 70% ethanol extracts of eggplant bark showed a protective effect against DNA damage.

<Example 7> Evaluation of inhibition of NO generation in macrophages

In the NO production amount measurement experiment, the NO production amount of the cell culture supernatant was measured using the Griess reaction. The cultured RAW264.7 cells ^{were aliquoted at a concentration of 5 × 10 4} cells/well, incubated for 24 hours at 37°C, 5% CO₂, and then stabilized. Samples diluted by concentration and LPS (1 μg/ml) were added. and incubated for an additional 24 hours. After that, 80 μl of the supernatant from the plate is transferred to a new 96-well plate, and 80 μl of Griess reagent is dispensed and allowed to react at room temperature for 10 minutes. Then, the amount of NO generated was measured by measuring the absorbance at 540 nm using an ELISA reader (16039400, Tecan, Mannedorf, Switzerland).

The results are shown in FIG. 6 . As a result of measuring the cell viability of the eggplant bark extract, the cell viability was high at the concentration of 100 μg/ml, so the NO generation inhibition experiment was performed from the concentration of 100 μg/ml (see FIG. 6A ). As a result of the NO generation inhibition test of eggplant peel extract, it was confirmed that the eggplant peel extract could inhibit the induction of inflammation caused by NO generation because it showed a significant NO generation inhibitory effect at a concentration of 10 μg/ml. (See Figure 6B).

<Example 8> Preparation of cedar bark extract

Commercially available red cedar bark was used. The cedar bark was crushed to a size of 0.1 to 1 cm, and the extract was obtained by reflux extraction 3 times at 25° C. for 2 hours with 70% ethanol in a volume corresponding to 30 times (wt/v%). The extracted extract was filtered twice under reduced pressure using filter paper (adventec Toyo 2, Japan), and concentrated under reduced pressure at 40 ° C or lower using a rotary evaporator (RV10D, Ika), and then freeze-dried at -80 ° C. prepared. The final yield was 39.31%.

<Example 9> Analysis of chlorogenic acid component of cedar bark extract

The chlorogenic acid component of the cedar bark extract was measured using High Performance Liquid Chromatography (HPLC) in the same manner as in Example 2.

The chlorogenic acid content and yield of the cedar bark extract extracted in Example 8 are shown in Table 2 below. The chlorogenic acid content was measured to be 16.6 mg/g (extract).

Extract chlorogenic acid
(mg/g of extract) Yield (%) cedar bark 16.6 11.05

<Example 10> Preparation of a mixture of eggplant bark extract and cedar bark extract

The eggplant bark extract extracted in Example 1 and the cedar bark extract extracted in Example 8 were each prepared at a concentration of 10 mg/ml, and then mixed and mixed in a 1:1 (v/v) volume ratio. Thereafter, the prepared mixture was used for the experiment.

<Example 11> DPPH radical scavenging activity of the mixture

DPPH radical scavenging activity (scavenging activity) was carried out in the same manner as in Example 3. Specifically, the mixture of eggplant bark extract and cedar bark extract prepared in Example 10 was diluted to concentrations of 0.01, 0.05, 0.1, 0.25, and 0.5 mg/ml to prepare 1 ml each, and the DPPH radical scavenging ability was measured.

The results are shown in FIG. 7 . Referring to FIG. 7 , it was confirmed that the DPPH radical scavenging ability of the eggplant bark extract and cedar bark extract mixture was improved compared to ascorbic acid and eggplant bark extract alone.

<Example 12> ABTS radical scavenging activity of the mixture

ABTS radical scavenging ability was carried out in the same manner as in Example 4. A mixture of the eggplant bark extract and cedar bark extract prepared in Example 10 was diluted to concentrations of 0.01, 0.05, 0.1, 0.25, and 0.5 mg/ml to prepare 1 ml each, and the ABTS radical scavenging ability was measured.

The results are shown in FIG. 8 . Referring to FIG. 8 , it was confirmed that the ABTS radical scavenging ability was improved in the mixture of the eggplant bark extract and the cedar bark extract compared to the eggplant bark extract alone.

<Example 13> Analysis of antioxidant active ingredients of the mixture

In Examples 11 and 12, the mixture had the same concentration of chlorogenic acid as the eggplant peel extract alone, but it was confirmed that the mixture had improved DPPH radical and ABTS radical scavenging ability compared to the eggplant peel extract alone. Therefore, it was attempted to analyze the component that increased the antioxidant effect.

The mixture of eggplant bark extract and cedar bark extract prepared in Example 10 was subjected to GS-MS analysis after TMS derivatization (trimethylsilylation).

For GC-MS, TMS derivatization (Trimethylsilylation) reaction was performed as follows. TMS was performed by adding pyridine (JUNSEI, Japan) and Sylon BFT solution (BSTFA + TMCS (99:1), SUPELCO) to the sample at a ratio of 1:1 and reacting at 105°C for 2 hours. GC analysis was performed using Agilent's GC analyzer (model-Agilent 6890, USA) and DB-5 (25 m×0.32 mm×0.52 μm) column. Helium was used as a carrier gas, and an injector 260° C. and a detector 280° C. were used as temperature conditions. The oven temperature was analyzed by maintaining the initial temperature at 50° C. for 5 minutes, increasing the temperature at a rate of 3° C./min, and maintaining it at the final temperature of 320° C. for 5 minutes. Mass spectrometry (MS analysis) are Agilent's mass spectrometer (Agilent model 5973. USA) was used for mass data of the sample obtained by peak analysis by EI mode (mass data) with the standard data library ^{(library data, Willy 7 th ed} ) a By comparison, the compound structure of the peak was identified (FIG. 9).

The results are shown in FIG. 9 . Referring to FIG. 9 , the main components were detected as a sugar component and a phenolic acid component, and chlorogenic acid was identified as the highest peak in the eggplant peel extract. In addition, high peaks of shikimic acid, quinic acid, catechin, and chlorogenic acid were confirmed in the mixture of eggplant bark extract and cedar bark extract.

In general, chlorogenic acid is an ester compound of caffeic acid and quinic acid, and it was assumed that quinic acid contained in the mixture was derived from chlorogenic acid. As a result, it was confirmed that the mixture further contained quinic acid and catechin along with chlorogenic acid as components to improve the antioxidant effect.

<Example 14> Statistical processing

All data are presented as mean ± standard deviation (SDM). All experiments were repeated three times. For comparison of measured values between each group, a one-way analysis of variance (ANOVA) test was performed and the difference was verified at the significance level of p < 0.05. GraphPad Prism 5 and Microsoft Excel 2007 were used for graph evaluation and statistics.

As described above, although specific embodiments of the present invention have been described in detail, those skilled in the art who understand the spirit of the present invention may add, change, delete, etc. other components within the scope of the same spirit, and other degenerate inventions However, other embodiments included within the scope of the present invention may be easily proposed. Therefore, it should be understood that the embodiments described above are illustrative in all respects and not restrictive. The scope of the present invention is indicated by the following claims rather than the above detailed description, and all changes or modifications derived from the meaning and scope of the claims and their equivalents are included in the scope of the present invention. should be interpreted as

Claims (12)

An antioxidant or anti-inflammatory composition comprising eggplant bark extract.
The composition according to claim 1, wherein the composition is a pharmaceutical or food composition.
The composition of claim 1, wherein the extract is reflux-extracted using an organic solvent.
The composition according to claim 3, wherein the organic solvent is selected from the group consisting of water, an alcohol having 1 to 4 carbon atoms, or a mixture thereof.
The composition of claim 1, wherein the extract has a concentration of 0.01 to 0.5 mg/ml.
The composition according to claim 1, wherein the composition has an active oxygen scavenging ability.
The composition of claim 1, wherein the composition inhibits oxidative damage to DNA.
The composition according to claim 1, wherein the composition inhibits the generation of nitric oxide (NO) that induces inflammation in mouse macrophages.
The composition for antioxidant or anti-inflammatory according to claim 1, wherein the composition further comprises a cedar bark extract.
The composition according to claim 9, wherein the cedar bark extract is extracted with one organic solvent selected from water, an alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof.
(1) reflux extraction of eggplant skins with an organic solvent to obtain an extract;
(2) concentrating the extract under reduced pressure; and
(3) freeze-drying the extract concentrated under reduced pressure in step (2);
A method for producing a composition for antioxidant and anti-inflammatory comprising a.
12. The method of claim 11,
The method comprises the steps of obtaining an extract by reflux-extracting the bark of cedar with an organic solvent; and;
preparing a mixture by mixing the cedar bark extract and eggplant bark extract;
Method for producing a composition for antioxidant and anti-inflammatory further comprising.

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