KR20110102246A - Composition for treating aged-related macular degeneration comprising extract from vaccinium uliginosum as an active ingredient - Google Patents

Abstract

The present invention relates to a composition for the treatment, prevention or improvement of macular degeneration containing a blueberry (Vaccinium uliginosum) extract or a blueberry fraction as an active ingredient.
The blueberry extract and the blueberry fraction, which are the active ingredients of the present invention, have the effect of fundamentally preventing the pathogenesis of macular degeneration by inhibiting A2E accumulation in retinal pigment epithelial cells and inhibiting oxidation of A2E to effectively protect retinal pigment epithelial cells. Therefore, the composition of the present invention can be developed into a variety of products, such as drugs for the treatment or improvement of excellent macular degeneration and health functional food.

Description

Composition for Treating Aged-related Macular Degeneration Comprising Extract from Vaccinium Uliginosum as an Active Ingredient}

The present invention relates to a composition for the treatment, prevention or improvement of macular degeneration containing a blueberry (Vaccinium uliginosum) extract or a blueberry fraction as an active ingredient.

In general, the nerve tissue located in the central part of the inner retina of the eye is called the macular. Most of the cells are gathered here, and the place where the image of the object is also the center of the macular and plays a very important role in vision. Eye disease that causes degeneration of the macula due to various causes and causes visual impairment is called macular degeneration (macular degeneration), and macular degeneration is known as one of the three major blindness diseases along with glaucoma and diabetic retinal disease.

This macular degeneration affects central vision, causing blurred vision in the center of vision, making it difficult to perform sophisticated activities such as reading and driving due to central dark spots, schizophrenia (a symptom of being distorted) or loss of vision in the local area. It is a very serious disease that, in severe cases, can lead to blindness.

The most common cause of macular degeneration is the increase in age, and is known to be related to family history, race, and smoking to some extent. In particular, Age-related Macular Degeneration (AMD) has long been the most common disease causing blindness in older people over 55 in Western society, and in the United States alone, more than 8 million people are currently a form of senile macular degeneration. It has dry AMD and more than 3 million people are reported to lose sight each year. Recently, macular degeneration patients are increasing exponentially in Korea, which is expected to be caused by increased exposure to ultraviolet light due to ozone depletion in addition to westernized eating habits. In particular, macular degeneration was originally a disease mainly occurring in the elderly, but in recent years, the age of the invention has been lowered from the elderly in their 60s to the 4 and 50s in the elderly, and the incidence of middle-aged people has also increased rapidly in recent years.

Macular degeneration can be classified into two types, dry and wet, and about 90% of people with macular degeneration are patients with dry macular degeneration. Dry macular degeneration causes macular tissue thinning or atrophy, resulting in the accumulation of waste products to kill retinal cells, and wet macular degeneration results in damage to the macular area due to rupture of new neoplasms due to the accumulation of waste products in or below the retina. do. In the dry form, early diagnosis is difficult because there is no sign, and it progresses slowly, and the risk of blindness is relatively small.However, it is possible to convert into wet macular degeneration with high risk of blindness. It is very important to prevent this disease.

In particular, dry macular degeneration has not yet been established. In the case of wet macular degeneration, laser photocoagulation is performed when the boundaries of the degeneration site can be clearly identified. In addition, photodynamic therapy (Vijudine), There are treatments such as the removal of neovascularization by injecting VEGF inhibitors into the eye, but the complete treatment has not been developed so much that the need for prevention is emphasized more.

Meanwhile, the macular degeneration market is largely divided into medicines and health functional foods. Currently, the only drug that has been approved for the treatment of senile macular degeneration in Korea is Ranibizumab (Lucentis), a treatment for wet macular degeneration, and there is no cure for dry macular degeneration. Ranibizumab is a humanized monoclonal antibody that blocks and recognizes vascular endothelial growth factor (VEGF A), and is known to be effective in restoring or maintaining vision in wet macular degeneration.

However, Genentech, Inc. has warned that clinical trials may increase Ranibizumab's risk of stroke, and Ranibizumab is expensive and inconvenient to inject into the eye. have.

On the other hand, if you look at the health functional food field, there are very few lutein cases approved as individual recognized forms. Lutein protects the macula of the retina and maintains macular pigment density to prevent macular degeneration, but studies show that prolonged use of supplements containing carotenoids, such as lutein, increases the risk of developing lung cancer, especially for smokers Appeared. Therefore, it can be dangerous for smokers to take lutein-related products.

In addition, the natural products reported by the KFDA as functional ingredients, but reported to be effective, include zeaxanthin, which is present as macular pigment, such as lutein, and anthocyanin contained in berries, but studies related to macular degeneration In fact, the current social phenomenon that macular degeneration patients are increasing not only in the elderly but also in the elderly, and the treatment for it is in need of development of medicines and dietary supplements to help prevent macular degeneration. It is true.

Therefore, the present invention is to recognize and improve the problems with the existing treatment and prevention methods for macular degeneration as described above, more specifically, the present invention is a natural plant mainly used for food blueberry (Vaccinium uliginosum It is an object of the present invention to provide a composition for the treatment or prophylaxis for macular degeneration with excellent therapeutic and prophylactic effects without side effects even when used for a long time using the extract extracted from the main component. In addition, the present invention is to provide a cosmetic composition, food composition and pharmaceutical composition for improving macular degeneration including blueberry extract.

The objects and advantages of the present invention as described above will become more apparent from the following detailed description, claims and drawings.

In order to achieve the above technical problem, the present invention provides a composition for improving or treating macular degeneration containing an extract derived from blueberry (Vaccinium uliginosum) as an active ingredient.

The cause of macular degeneration and its progression are known to date, the greatest risk factor of macular degeneration is pyridnium bis-retinoid (A2E) that accumulates in the retinal pigment epithelium as age increases. A2E is one of the most common lipofuscins found in the eye, where A2E accumulates in retinal pigment epithelial cells and kills them, leading to the death of photoreceptor cells, leading to the accumulation of photoreceptor cells in the macula. Loss of function leads to macular degeneration.

In addition, ultraviolet radiation increases the risk of macular degeneration or accelerates the progression of the disease, since radical changes in retinal pigment epithelial cells are dramatically increased when A2E is converted to A2E in the photooxidized form by ultraviolet radiation.

Thus, the present inventors have been studying to find a substance having inhibitory ability to accumulate A2E and A2E oxidation in retinal pigment epithelial cells for the fundamental treatment and prevention of macular degeneration, while extracts from blueberries inhibit A2E accumulation in retinal pigment epithelial cells. And inhibiting A2E oxidation by ultraviolet rays and found to have an effect of protecting retinal pigment epithelial cells, and scientifically proved this by experimental results to confirm that the treatment and prevention of macular degeneration can be confirmed. To complete.

Firstly used as a component for the treatment or improvement of macular degeneration in the present invention is a birch and deciduous shrub plant that grows in Halla, Geumgang, and Baekdusan of the Korean Peninsula, which blooms in June to July and bears fruit in August. to be. The main components of blueberry include sugar (8-11.8%), fruit acid (2-2.25%), tannin (0.15-0.25%), fiber, and the like.

Originally, the pharmacological use of blueberries began with the recommendation of the fruit as a medicine related to female physiology by a woman named St. Hildgard (1098-1179), a medieval German abbey. Blueberries have been used for a long time as health liquor in China and North Korea, but the scientific pharmacological effect of the pharmacological action is inadequate and the component analysis is not properly performed.

As described above, the present invention adopts blueberry, which has been mainly used for food, as a raw material for improving, preventing or treating macular degeneration, and the composition for preventing or treating macular degeneration of the present invention has less side effects to the human body. Formulations such as eye drops, ophthalmic ointments, etc. to treat or ameliorate macular degeneration are very suitable for direct use in the eye.

In particular, according to the present invention, it was established by experiment that the components extracted from blueberries using ARPE-19 cells have an effect of inhibiting A2E accumulation and inhibiting A2E oxidation, as described below. And the blueberry fraction can fundamentally cure the pathogenesis of macular degeneration, such as A2E accumulation and A2E oxidation, and can be very useful for the treatment, prevention and improvement of macular degeneration through a pharmaceutical composition or functional food containing the same as an active ingredient. It is expected to be. In particular, it can be usefully used for the improvement, treatment and prevention of senile macular degeneration.

Accordingly, the present invention relates to the use of the blueberry extract or the blueberry fraction for the treatment, prevention or improvement of macular degeneration, the composition for the treatment, prevention or improvement of macular degeneration according to a suitable embodiment of the invention to the treatment, prevention of macular degeneration Or a method for improvement.

As one embodiment of the invention, the present invention provides a composition for the treatment, prevention or improvement of macular degeneration, containing blueberry extract or fractions fractionated therefrom as active ingredients.

The blueberry extract may be extracted from the flowers, fruits, leaves or bark of the blueberry, preferably from the fruit of the blueberry.

The extraction method may be separated according to a conventional method known in the art for extracting natural products, that is, using a solvent conventionally used in the art under conditions of ordinary temperature, pressure, and time. As an extraction solvent for separating the blueberry extract of the present invention, a solvent which can be generally used in the extraction process may be used, and it is preferable to extract using water, anhydrous or hydrated lower alcohol having 1 to 4 carbon atoms. It may also be extracted using a solvent selected from the group consisting of acetone, ethyl acetate, butyl acetate and 1,3-butylene glycol.

The blueberry fraction may be obtained from the flowers, fruits, leaves or bark of the blueberry, preferably from the fruit of the blueberry. It can also be obtained from blueberry extract. In addition, the blueberry fraction which shows especially significant effect in this invention is a polyphenol fraction of a blueberry, the pigment fraction of a blueberry, or a mixture thereof, Preferably it is a polyphenol fraction of a blueberry.

The blueberry fraction may be fractionated by a fractionation method well known in the art, for example, may be separated by a solid phase extraction (SPE) as used in this example. Blueberry fractions contain carbohydrates / acid fractions containing a large amount of carbohydrates / acid components in a proper order by selecting an appropriate elution solvent, polyphenol fractions containing a large amount of polyphenolic compounds (flavonol, etc.), pigment components (anthocyanins, etc.). It can be separated into a pigment fraction containing a large amount. The elution solvent may be appropriately selected by those skilled in the art. As used in the following examples, ethyl acetate may be used to obtain a polyphenol fraction, and an aqueous HCl / methanol solution may be used to obtain a pigment fraction. Among the blueberry fractions, it was confirmed that the polyphenol fraction, the pigment fraction of the blueberry or a mixture thereof showed a significant effect, in particular the polyphenol fraction is most preferred.

In one embodiment of the present invention, the composition of the present invention may be provided in the form of a pharmaceutical composition comprising a pharmaceutically effective amount of a blueberry extract. As used herein, the term "pharmaceutically effective amount" means an amount sufficient for the above-mentioned effective ingredient compound to achieve the effect of treating or preventing macular degeneration.

The pharmaceutical composition of the present invention may comprise a pharmaceutically acceptable carrier in addition to the blueberry extract or blueberry fraction. Such pharmaceutically acceptable carriers are those commonly used in the art, such as lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, Calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil, and the like. In addition, the pharmaceutical composition of the present invention may include diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, surfactants, and other pharmaceutically acceptable additives.

Suitable dosages of the pharmaceutical compositions of the present invention may be prescribed in various ways depending on such factors as the formulation method, mode of administration, age, weight, sex, morbidity, food, time of administration, route of administration, rate of excretion and response to response of the patient. Can be. For example, it can be administered in a dose of 0.001 to 1000 mg / kg per day, the administration can be administered once or several times a day.

The pharmaceutical composition of the present invention can be administered orally or parenterally. Examples of the dosage form include eye drops, eye ointments, injections, tablets, capsules, granules, powders, and the like, and eye drops or eye ointments are particularly preferable. These can be formulated using techniques that are widely used.

For example, eye drops include isotonic agents such as sodium chloride and concentrated glycerin, buffering agents such as sodium phosphate and sodium acetate, surfactants such as polyoxyethylene sorbitan monooleate, polyoxyl stearic acid 40 and polyoxyethylene hydrogenated castor oil, and citric acid Stabilizers, such as sodium and sodium edetate, and preservatives, such as benzalkonium chloride and paraben, etc. can be used as needed. Although pH should just be in the range which is acceptable for ophthalmic preparations, the range of 4-8 is preferable.

In addition, ophthalmic ointment can be prepared using a common base such as white petrolatum, liquid paraffin. On the other hand, oral preparations such as tablets, capsules, granules, and powders include bulking agents such as lactose, crystalline cellulose, starch, vegetable oil, glidants such as magnesium stearate and talc, and binders such as hydroxypropyl cellulose and polyvinylpyrrolidone. Disintegrating agents such as calcium carboxymethyl cellulose, low-substituted hydroxypropyl methyl cellulose, coating agents such as hydroxypropyl methyl cellulose, macrogol, silicone resins, and coating agents such as gelatin coatings, etc. Can be.

Furthermore, as another embodiment of the present invention, the composition of the present invention may be provided in the form of a food composition, in particular a functional food composition. Functional food compositions of the present invention include ingredients that are commonly added in the manufacture of food, and include, for example, proteins, carbohydrates, fats, nutrients and seasonings. For example, natural carbohydrates include monosaccharides (eg, glucose, fructose, etc.); Disaccharides (eg maltose, sucrose, etc.); oligosaccharide; Polysaccharides (eg, dextrins, cyclodextrins, etc.); And sugar alcohols (eg, xylitol, sorbitol, erythritol, and the like). As the flavoring agent, natural flavoring agents (eg, taumartin, stevia extract, etc.) and synthetic flavoring agents (eg, saccharin, aspartame, etc.) can be used.

In one embodiment of the invention, the present invention provides a method for treating macular degeneration comprising administering the above-mentioned pharmaceutical composition or food composition to a patient with macular degeneration or a subject at risk for macular degeneration, or depending on the formulation of the composition, Provide a method of prevention or improvement. The subject is a mammal, including a human.

The blueberry extract and the blueberry fraction, which are the active ingredients of the present invention, have the effect of fundamentally preventing the etiology by inhibiting A2E accumulation in retinal pigment epithelial cells and inhibiting the oxidation of A2E. Therefore, the composition of the present invention can be developed into a variety of products, such as drugs for the treatment or improvement of excellent macular degeneration and health functional food.

Figure 1 is a graph showing the test results for the antioxidant activity of the blueberry extract and blueberry fraction for A2E oxidized by ultraviolet A.
Figure 2 is a graph showing the test results of the protective effect of the blueberry and its fractions against apoptosis by UVA in ARPE-19 cells accumulated A2E.
Figure 3 is a graph showing the test results for the inhibition of the accumulation of A2E and iso A2E by treatment with blueberry extract and blueberry fraction in ARPE-19 cells.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention in more detail, it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples in accordance with the gist of the present invention. .

[ Example  One] Blueberry  Preparation of Extract

Among the fruits of the blueberry, the good condition was selected and washed thoroughly. Then, 2000 ml of water was added to 100 g of blueberries, followed by extraction of hot water twice at 16C for 16 hours to obtain a 10% blueberry extract. The blueberry extract obtained as described above was freeze-dried, processed into a powder form, and then stored at -40 ° C. until used to prepare the blueberry fraction and used in the experiments described below.

[ Example  2] Blueberry Fraction  Produce

After the blueberry extract powder prepared in Example 1 was dissolved in distilled water, two C18 Sep-Pak cartridges were connected, and preconditioned with 10 ml ethyl acetate and absolute methanol, 0.01 N aqueous HCl was flowed into the cartridge.

After the whole extract was loaded into the cartridge, 6 mL of 0.01 N aqueous HCl was flowed to remove carbohydrates, acids, and other water-soluble compounds. The layer thus removed was used as a carbohydrate / acid fraction in the following experiment as a comparative example.

Next, nitrogen gas was passed through the connected C18 Sep-Pak cartridges for 10 minutes to dry the cartridge. 20 ml ethyl acetate was poured into the dried cartridge as described above, and phenolic compounds were fractionated in a test tube to collect polyphenol fractions.

Next, anthocyanins adsorbed using 10 ml of anhydrous methanol containing 0.1% HCl were eluted to obtain a pigment fraction.

The solvent of the fraction was concentrated at 40 ° C. with a vacuum concentrator to remove all residual solvents, thereby preparing each blueberry fraction (carbohydrate / acid fraction; pigment fraction; polyphenol fraction).

The carbohydrate / acid fraction and pigment fraction obtained through the above process are dissolved in distilled water, respectively. The blueberry polyphenol fraction is dissolved in 50% aqueous ethanol, and then stored at -70 ° C. for the experiment and analysis described later. Used.

[ Example  3] Cell culture method

Human adult ARPE cells (ARPE-19; catalog no. CRL-2302) used in the experiment and analysis of the present invention were used from the Catholic Medical University Institute of Ophthalmology. The ARPE cells were used in the experiment by culturing in DMEM (Dulbecco's modified Eagle's medium) medium containing 10% fetal bovine serum (FBS), 100 U / ml penicillin, and 100 mg / ml streptomycin. , Incubator conditions were 5% CO ₂ , 37 ° C. When the cells were used for the experiment, seeded in 6-well plates with 5 × 10 ⁴ cells was used for the experiment.

[ Example  4] A2E synthesis method

Synthesis of A2E used in the experiments and analysis of the invention, all- trans- retinal and ethanolamine dissolved in ethanol to which a mixture of the (2 1 molar ratio) of acetic acid was added in a dark room conditions, the reaction for 2 days, and then, The mixture was concentrated in vacuo at 40 ° C. and purified by silica gel column chromatography for synthesis. The synthesized A2E was dissolved in DMSO (Dimethyl Sulfoxide) at a stock concentration of 20 mM and stored at -20 ° C until the experiment.

[ Experimental Example  One] For A2E oxidized by UV A Blueberry  Extracts Blueberry Fraction  Oxidation Inhibitory ability  Experiment

In order to measure the antioxidant activity of the blueberry extract and its fractions against A2E oxidation by ultraviolet A irradiation, 20 μl of 1 mM A2E (obtained by diluting the 20 mM A2E stock solution obtained in Example 4 with PBS) was 0.01% DMSO (Dimethyl). Dissolve in 160 μl of PBS (phosphate buffered saline) added with sulfoxide, add 180 μl to 96-well plate, and then control (control) or suggested concentration (blueberry extract: 100, 250, 500 mg / 20 μl of samples diluted with blueberry polyphenol / pigment fractions: 12.5, 25, 50, 100 mg / ml) were added.

Thereafter, the absorbance was measured at 430 nm using an ELISA microplate reader, and then irradiated with ultraviolet A (3 J / cm ² ), and the absorbance was measured in the same manner as before. The measured absorbance values were calculated using the A2E standard curve, and then the concentration of oxidized A2E was determined by the concentration difference before and after UV A irradiation.

On the other hand, the equipment used in the experiment used an ultraviolet A lamp (Sankyo Denki, Tokyo, Japan) having a wavelength between 355 nm ~ 375 nm. The irradiation method of ultraviolet A is to wash the well plate with PBS before irradiation, open the lid of the well plate with only PBS that the cells are not dried, and under the UV A lamp, 3 J / cm ² (Irradiation density: 2.35 ± 0.3 mW / cm ² ; irradiation time: 22 to 24 min). The UV A dose was measured using a UV A radiometer.

[ Experimental Example  2] A2E accumulated ARPE For cell death by UVA in -19 cells Blueberry  And Blueberry Fraction  cell Protection  Experiment

After dispensing 5 × 10 ⁴ ARPE-19 cells per well into 6-well plates, 20 μl A2E was accumulated for 5 days. Blueberry extract and blueberry fraction were then treated with each concentration (blueberry extract: 100, 250, 500 mg / ml; blueberry fraction: 12.5, 25, 50, 100 mg / ml) and UV A (3 J / cm ² ) After incubation for 24 hours.

Thereafter, the survival rate of the cells was measured using the MTT assay. This MTT assay is based on the principle that yellow yellow tetrazolium salt (MTT) reacts with mitochondrial reductase in living cells to form purple formazan crystals. The production of is increased and the absorbance is measured to be high.

After adding DMEM medium containing 0.5 mg / ml MTT according to the MTT assay, the light was blocked and reacted at 37 ° C. for 2 hours in an incubator. After completion of the reaction, cells were sufficiently dissolved in 2 ml of DMSO, and then the absorbance at 550 nm was measured using an ELISA microplate reader, and the percentage (%) of the group that did not accumulate both A2E and the sample (normal control). The viability of the cells is shown.

[ Experimental Example  3] ARPE On -19 cells Blueberry  And that Fraction  A2E / by treatment isoA2E Accumulation of Inhibition  Experiment

After dispensing 5 × 10 ⁴ ARPE-19 cells per well into a 6-well plate, the blueberry extract and the blueberry fraction were treated for each concentration, and then cultured for 2 days to accumulate A2E as in the previous experiment. Thereafter, the cells were lysed using 25 mM HEPES buffer including protease inhibitors (Complete ^™ , Roche, Mannheim, Germany) and quantitatively analyzed for accumulation of A2E using high performance liquid chromatography (HPLC). A2E uses a known concentration of A2E solution for quantification and A2E levels were corrected by measuring the protein by Bradford method (Bio-Rad Protein Assay, Bio-Rad, Hercules, CA). Samples were used after extracting A2E with CHCl ₃ from ARPE-19 cells before HPLC quantitation, and the conditions and specific experimental methods of HPLC quantitation were performed as shown below.

[HPLC Quantitative Assay]

A reverse phase C18 column (250 4.6 mm, Cosmosil 5C18, Nacalai Tesque, Osaka, Japan) was used as the Waters 600E HPLC equipped with a Waters 996 Photodiode Array Detector A instrument. A2E is dissolved by dissolving with acetonitrile in water containing 0.1% trifluoroacetic acid (TFA). When separated as described above, the absorbance at 430 nm was measured and expressed as a percentage (%) for the group without A2E accumulation.

[Experimental Result 1] For A2E oxidized by UV A Blueberry  Extracts and their Fraction  Oxidation Inhibitory ability

In order to confirm the antioxidant activity of the blueberry extract and blueberry fraction of the present invention against A2E, the concentration of A2E oxide after UV A irradiation was measured according to the same method as in [Experimental Example 1] (FIG. 1).

As shown in (a) of FIG. 1, it can be seen that A2E oxidized at concentrations of 125, 250, and 500 μg / ml of blueberry extracts showed 20, 32, and 41% lower than untreated cells, respectively. It was confirmed that the blueberry extract showed an antioxidant activity against A2E in a concentration-dependent manner.

In addition, as shown in (b) of Figure 1, blueberry fraction is 33, 71, 85% lower than the untreated A2E treated cells at 25, 50, 100 ㎍ / ㎖ concentration of the polyphenol fraction, respectively, It was found that the highest inhibitory ability of A2E oxidation. In addition, it can be seen that the pigment fraction is 32, 53, 66% lower for the untreated A2E at the same concentration as the polyphenol fraction, respectively. Carbohydrate / acid fractions, on the other hand, were found to have no antioxidant activity. Therefore, the blueberry fraction showed the ability to inhibit the A2E oxidation in the order of polyphenol fraction> pigment fraction >> carbohydrate / acid fraction.

[Experiment 2] A2E accumulated ARPE For cell death by UVA in -19 cells Blueberry  Extract and Blueberry Fraction  cell Protection

In order to confirm the cell protective ability of the blueberry extract and blueberry fraction of the present invention, after irradiation with ultraviolet A according to the same method as in the above [Experimental Example 2] was shown by measuring the degree of self-killing of cells by MTT assay (Fig. 2).

Referring to FIG. 2A, cells irradiated with ultraviolet A without accumulating A2E (Non A2E-UV) and cells not irradiated with ultraviolet A after A2E accumulation (A2E-non UV) do not accumulate A2E. There was no significant difference between the cells that did not irradiate with UV A (normal control), and after A2E accumulation, there was a significant difference between the cells that were irradiated with UV A (negative control) and normal control.

In addition, as shown in Figure 2 (a), the survival rate of the cells treated with the blueberry extract 125, 250, 500 ㎍ / ㎖ concentration was 63, 134, 167%, respectively, based on the cell survival rate of the negative control (negative control) By appearing high, blueberry extract demonstrated concentration dependent protection of ARPE-19 cells.

In addition, as shown in (b) of FIG. 2, the viability of the cells treated with the polyphenol fraction 12.5, 25, 50, and 100 ㎍ / ml concentration in the blueberry fraction was 63, 80, 161 based on the cell survival rate of the negative control, respectively. The highest cell protection was shown by 207%, and the cell viability of the pigment fraction treated was 14, 26, 44, 75% higher than the negative control at the same concentration as the polyphenol fraction. . Blueberry carbohydrate / acid fractions, on the other hand, were found to be incapable of protecting. Therefore, the blueberry fraction was found to protect the ARPE-19 cells from oxidation by UVA in the order of polyphenol fraction »pigment fraction> carbohydrate / acid fraction.

[Experimental Result 3] ARPE In -19 cells Blueberry  Extract and Blueberry Fraction  Accumulation of A2E and iso A2E by Treatment Inhibition

In order to confirm the accumulation inhibitory ability against A2E and iso A2E of the blueberry extract and the blueberry fraction of the present invention, the concentration of A2E and iso A2E was measured and shown in FIG. 3.

As shown in (a) of FIG. 3, the concentration of A2E / iso A2E accumulated in ARPE-19 cells treated with 50, 100, 250, and 500 μg / ml concentrations of blueberry extract was measured. 18, 26, 37, 34% / 8, 11, 18, 13% (A2E / iso A2E) lower than the cells, respectively, it was confirmed that the blueberry extract inhibited the accumulation of A2E / iso A2E in a concentration-dependent manner.

In addition, as shown in (b) of FIG. 2, the blueberry fraction is a cell in which A2E / iso A2E concentration accumulated in ARPE-19 cells treated with pigment fractions 25, 50, and 100 μg / ml was not treated with the blueberry fraction. It showed 36, 45, 51/13, 14, 40% lower than the highest A2E / iso A2E accumulation inhibitory ability, and the concentration of A2E / iso A2E accumulated in cells treated with polyphenol extract was 19, 20 at the same concentration. , 40 / 0.1, 3.3, 15% lower. Carbohydrate / acid fractions, on the other hand, were found to be incapable of inhibiting A2E / isoA2E accumulation. Therefore, the blueberry fraction demonstrated the effect of inhibiting A2E and isoA2E accumulation in ARPE-19 cells in the order of pigment fraction> polyphenol fraction »carbohydrate / acid fraction.

From the above experimental results, the blueberry extract of the present invention and fractions fractionated therefrom protects ARPE-19 cells by inhibiting A2E accumulation in ARPE-19 cells and inhibiting oxidation of A2E by UV light. It was found to be effective. Therefore, according to the composition of the present invention can fundamentally prevent the pathogenesis of macular degeneration, it is expected that the composition of the present invention can be very useful for improving or alleviating macular degeneration.

Having described the specific part of the present invention in detail, it is apparent to those skilled in the art that such a specific technology is only a preferred embodiment and the scope of the present invention is not limited thereto. Accordingly, the substantial scope of the invention will be defined by the claims appended hereto and their equivalents.

Claims (13)

Pharmaceutical composition for the treatment, prevention or improvement of macular degeneration containing blueberry extract or blueberry fraction as an active ingredient. The pharmaceutical composition of claim 1, wherein the blueberry extract is extracted with water, a lower alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof. The pharmaceutical composition for treating, preventing or ameliorating macular degeneration of claim 1, wherein the blueberry fraction is fractionated by solid phase extraction (SPE). The pharmaceutical composition for treating, preventing or improving macular degeneration according to claim 1, wherein the macular degeneration is senile macular degeneration. The pharmaceutical composition for treating, preventing or ameliorating macular degeneration according to claim 1, wherein the blueberry fraction is a polyphenol fraction, a pigment fraction, or a mixture thereof. According to claim 5, wherein the polyphenol fraction is a pharmaceutical composition for the treatment, prevention or improvement of macular degeneration obtained by removing the carbohydrate / acid fraction from the blueberry extract with HCl solution and eluted with ethyl acetate. [6] The macular degeneration of claim 5, wherein the pigment fraction is a fraction obtained by removing the carbohydrate / acid fraction from the blueberry extract with HCl solution, eluting the polyphenol fraction with ethyl acetate, and then eluting with methanol / HCl solution. Pharmaceutical compositions for treatment, prophylaxis or improvement. The method of claim 5, wherein the pharmaceutical composition is a pharmaceutical composition for the treatment, prevention or improvement of macular degeneration, characterized in that the formulation as eye drops or eye ointment. Food composition for the treatment, prevention or improvement of macular degeneration containing blueberry extract or blueberry fraction as an active ingredient. 10. The food composition for treating, preventing or improving macular degeneration according to claim 9, wherein the macular degeneration is senile macular degeneration. The food composition of claim 9, wherein the blueberry fraction is a polyphenol fraction of a blueberry, a pigment fraction, or a mixture thereof. The pharmaceutical composition of claim 11, wherein the polyphenol fraction is obtained by removing the carbohydrate / acid fraction from the blueberry extract with an HCl solution and then eluting with ethyl acetate. 12. The macular degeneration of claim 11, wherein the pigment fraction is a fraction obtained by removing a carbohydrate / acid fraction from a blueberry extract with an HCl solution, eluting a polyphenol fraction with ethyl acetate, and then eluting with a methanol / HCl solution. Pharmaceutical compositions for treatment, prophylaxis or improvement.

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