WO2023286950A1 - Composition containing ginseng berry extract and lutein for preventing or treating macular degeneration - Google Patents

Abstract

The present invention provides a pharmaceutical composition and a food composition for preventing or treating macular degeneration, wherein the pharmaceutical composition and food composition contain ginseng berry extract and lutein. The composition for preventing or treating macular degeneration according to the present invention, wherein the composition contains ginseng berry extract and lutein, can reduce the migration of retinal cells from the cytoplasm of NF-κB to the nucleus, the decomposition of IκB, the expression of IL-1β, the expression of IL-6, the expression of TNFα, and the expression of VEGFα, and thereby reduce abnormal inflammatory reactions, apoptosis, and blood vessel formation to induce the prevention, relief, or treatment of macular degeneration. In addition, the composition can be effectively used as a pharmaceutical or food composition for preventing, relieving, and treating macular degeneration by suppressing light-induced death of retinal pigment epithelial cells, A2E photooxidation in retinal pigment epithelial cells, and damage to retinal cell layers.

Description

Composition for preventing or treating macular degeneration containing ginseng berry extract and lutein

The present invention relates to a composition for preventing or treating macular degeneration comprising a ginseng berry extract and lutein, and more particularly, to a pharmaceutical composition for preventing or treating macular degeneration comprising a ginseng berry extract obtained by extracting ginseng berry with water as a solvent and lutein; and It relates to food compositions.

The nerve tissue located in the center of the inner retina of the eye is called the macula. Most of the visual cells are gathered here, and the place where the image of the object is formed is also the center of the macula, so the macula plays a very important role in vision. Visual acuity refers to the ability to recognize the existence and shape of an object. It is most sensitive when the image of an object is focused on the fovea of the macula (central vision), and deteriorates as it moves toward the retina (peripheral vision). When you see an object, you see it through the central part of the retina, the macula, and it is generally referred to as central vision. The macula can cause visual impairment due to various causes such as aging, genetic factors, toxicity, and inflammation, which is called macular degeneration.

Macular degeneration affects central vision, making the center of the field of vision blurry, making it difficult to perform sophisticated activities such as reading and driving through central scotoma, metamorphosis, or local vision loss, and in severe cases, leading to blindness. It is a very serious disease.

There are two types of macular degeneration: non-exudative (dry) macular degeneration and exudative (wet) macular degeneration, and 90% of people with macular degeneration have symptoms of the dry form. In dry macular degeneration, waste products can build up in the tissue beneath the macula, forming yellow deposits called drusen. The presence of drusen obstructs blood flow to the retina, especially the macula, and when blood flow is reduced, the supply of nutrients to the macula is reduced, stopping or atrophying the efficient functioning of photosensitive cells and causing chronic inflammation. It's coming to light. In wet macular degeneration, new weak blood vessels can grow in or under the retina, allowing fluid and blood to leak into the space beneath the macula. Wet macular degeneration is sometimes described as choroidal neovascularization. The choroid is a subretinal blood vessel area and neovascularization refers to the growth of new blood vessels within the tissue. blood vessels are formed and grow.

The underlying mechanism of this macular degeneration is still unknown. Age-related macular degeneration, a type of macular degeneration, is characterized by loss of central vision due to the death of photoreceptor cells, and is believed to have a multifactorial etiological mechanism. Recently, it has been reported that various factors (e.g., smoking, obesity, eating habits, increased cholesterol in the blood, blue light irradiation, etc.) cause the acceleration of age-related macular degeneration, which means that damage to photoreceptor cells and subsequent cell death can lead to retinal damage. Causes degeneration (denaturation) of the pigment epithelium. On the other hand, it is known that the accumulation of fat brown pigment (lipofuscin) in the center of the retina of the eye is greatest in the retinal pigment epithelial cells below the center of the retina. This reflects the fact that this area has a high density of rod cell photoreceptors. N-retinyl-Nretinylidene ethanolamine (A2E) is the major chromophore of lipofuscin and induces the production of reactive oxygen species when exposed to blue light. Blue light is light with a relatively high energy amount of about 440 nm in wavelength, and when exposed to it for a short time, cell damage increases, and this increases more as the light of shorter wavelength increases. That is, continuous light exposure further promotes the death of retinal pigment epithelial cells in age-related macular degeneration, which eventually becomes one of the main causes of degeneration of photoreceptor cells. In addition, visual acuity due to retinal cell dysfunction is closely related to A2E oxidation, and thus may be a major cause of age-related macular degeneration.

To date, there are very few treatments available for macular degeneration. Moreover, there are some treatments such as anti-VEGF for wet macular degeneration, but there is no known treatment for dry macular degeneration.

On the other hand, lutein (lutein) is a carotenoid (carotenoid) that is naturally produced and does not have vitamin A activity and exists in a natural state, and is contained in a large amount in green and yellow vegetables. Lutein, one of the most abundant carotenoids in food and human blood, is known to perform antioxidant functions and protect plants from ultraviolet rays in plants. It is known as a component that plays an important role in improving eyesight.

However, recently, a research team at Moran Eye Hospital affiliated with Utah State University in the U.S. reported on the JAMA Ophthalmology, a journal of the American Medical Association, that round, yellow crystal-like glitter was found inside the central retinal fossa of women who had taken lutein excessively for a long time. reported that the substance was found. It was found that the patient took 20 mg of lutein supplements daily for 8 years and steadily consumed lutein-rich spinach, broccoli, kale, avocado, etc. Therefore, the research team assumed that lutein was precipitated in the eyeball, crystals were formed, and macular degeneration occurred.

Therefore, although lutein shows an excellent therapeutic effect in the treatment of macular degeneration, long-term or excessive intake causes side effects that cause macular degeneration. Therefore, it is necessary to develop a method that can prevent, alleviate or suppress them.

The inventors of the present invention have previously revealed that a ginseng fruit extract having a different composition from that of a ginseng root extract can have preventive, ameliorative, or therapeutic effects on macular degeneration (Korean Application No. 10-2020-0187943). The inventors completed the present invention by confirming that, when the previously known lutein and ginseng fruit extract are combined, a more excellent eye protection effect can be exhibited even at a lower concentration than when each substance is used individually.

The technical problem to be achieved by the present invention is to provide a composition capable of inducing prevention, improvement or treatment of macular degeneration.

The technical problem to be achieved by the present invention is not limited to the above-mentioned technical problem, and other technical problems not mentioned can be clearly understood by those skilled in the art from the description below. There will be.

In order to achieve the above technical problem, one aspect of the present invention provides a pharmaceutical composition for preventing or treating macular degeneration comprising a ginseng berry extract and lutein.

In one embodiment, the ginseng fruit extract is Korean ginseng ( Panax ginseng ), Hoegi ginseng ( P. quiquefolius ), Jeonchilseng ( P. notoginseng ), bamboo shoots ( P. japonicus ), trifolium ginseng ( P. trifolium ), Himalayan ginseng ( P. pseudoginseng ), Vietnamese ginseng ( P. vietnamensis ) and American ginseng ( Panax quinquefolium ) It may be characterized in that the extract is extracted from the fruit of one or more species of ginseng selected from the group consisting of.

In one embodiment, the ginseng berry extract may be extracted using water as a solvent.

In one embodiment, the lutein may be characterized in that it is extracted from natural products by supercritical extraction.

In one embodiment, the composition may include the ginseng berry extract and the lutein in a weight ratio of 5:1 to 1:5.

In one embodiment, the macular degeneration may be characterized by light-induced death of retinal pigment epithelial cells, A2E photooxidation in retinal pigment epithelial cells, or retinal cell layer damage.

Another aspect of the present invention provides a food composition for preventing or treating macular degeneration comprising a ginseng berry extract and lutein.

The composition for preventing or treating macular degeneration comprising a ginseng berry extract and lutein according to an embodiment of the present invention is characterized by the movement of NF-κB from the cytoplasm to the nucleus, degradation of IκB, and expression of IL-1β in retinal cells. , IL-6 expression, TNFα expression and VEGFα expression can be reduced, and through this, abnormal inflammatory responses, apoptosis and angiogenesis can be reduced to induce prevention, improvement or treatment of macular degeneration. In addition, the composition can prevent, improve, and treat macular degeneration by inhibiting light-induced death of retinal pigment epithelial cells, A2E photooxidation in retinal pigment epithelial cells, and retinal cell layer damage.

In addition, the composition is known to have preventive, ameliorative and therapeutic effects on macular degeneration, but has lutein and macular degeneration prevention, improvement and treatment effects reported to have toxic effects such as causing macular degeneration when excessively consumed. As a result, it is possible to have a higher prevention, improvement, and treatment effect of macular degeneration while containing the ginseng berry extract, which the inventors have found, at a lower concentration.

The effects of the present invention are not limited to the above effects, and should be understood to include all effects that can be inferred from the description of the present invention or the configuration of the invention described in the claims.

1 is a result of analyzing the effect of a composition containing a ginseng berry extract and lutein according to Example 1 on cell death of retinal pigment epithelial cells (ARPE-19) by blue light.

2 is a result of analyzing the effect of the composition containing the ginseng berry extract and lutein according to Example 1 on the photooxidation of A2E by blue light in retinal pigment epithelial cells (ARPE-19).

Figure 3 is a representative image (outer nuclear layer (ONL), These are the Inner nuclear layer (INL), Photoreceptor segment layer (PL), and Whole retina).

4 is a result of analyzing the effect of the composition containing the ginseng berry extract and lutein according to Example 1 on retinal photoreceptor layer damage in an animal model of macular degeneration induced by blue light.

5 is an image and statistical results of Western blot analysis of the effect of the composition containing the ginseng berry extract and lutein according to Example 1 on the amount of NF-κB in the cytoplasm and nucleus in retinal pigment epithelial cells (ARPE-19). .

6 is an image and statistical results of Western blot analysis of the effect of the composition containing the ginseng berry extract and lutein according to Example 1 on the amount of IκB in the cytoplasm in retinal pigment epithelial cells (ARPE-19).

7 is a PCR analysis result of the effect of the composition containing the ginseng berry extract and lutein according to Example 1 on the expression level of IL-1β mRNA in retinal pigment epithelial cells (ARPE-19).

8 is a PCR analysis result of the effect of the composition containing the ginseng berry extract and lutein according to Example 1 on the expression level of IL-6 mRNA in retinal pigment epithelial cells (ARPE-19).

9 is a result of PCR analysis of the effect of the composition containing the ginseng berry extract and lutein according to Example 1 on the expression level of TNFα mRNA in retinal pigment epithelial cells (ARPE-19).

10 is a PCR analysis result of the effect of the composition containing the ginseng berry extract and lutein according to Example 1 on the expression level of VEGFα mRNA in retinal pigment epithelial cells (ARPE-19).

A pharmaceutical composition for preventing or treating macular degeneration according to an aspect of the present invention includes a ginseng berry extract and lutein.

Ginseng ( Panax ginseng ) is a plant belonging to the genus Panax of the Araliaceae family , and has been widely used for more than 2,000 years as a unique medicinal plant in Korea. The ginseng fruit extract is Korean ginseng ( Panax ginseng ), Hoegi ginseng ( P. quiquefolius ), Jeonchilseng ( P. notoginseng ), bamboo shoots ( P. japonicus ), trifolium ginseng ( P. trifolium ), Himalayan ginseng ( P. pseudoginseng ), It may be characterized in that it is an extract extracted from the fruit of one or more species of ginseng selected from the group consisting of Vietnamese ginseng ( P. vietnamensis ) and American ginseng ( Panax quinquefolium ). Preferably, the ginseng fruit used in the present invention may be the fruit of Korean ginseng (Panax ginseng).

The ginseng fruit is a part of ginseng and is different from the commonly used ginseng root (root) in the type and composition of ingredients contained in it. Specifically, ginseng berries contain more vitamins and minerals than ginseng roots. In addition, ginseng berries contain more ginsenosides than ginseng roots, and the composition of the ginsenosides contained is different. Therefore, in one embodiment of the present invention, the ginseng fruit extract is a ginsenoside PT (Protopanaxatriol) system containing ginsenosides Re, Rg1, Rg2, etc., ginsenosides containing ginsenosides Rb1, Rb2, Rc, Rd, etc It can contain more than PD (Protopanaxadiol) system, and thus can show different effects. For example, ginseng fruit has been reported to exhibit superior antidiabetic efficacy than ginseng root (Dey et al., Phytomedicine , 2003, 10; 600-605). In addition, as ginseng berries become overripe in green, such as leaves, the flesh changes to red or yellow depending on the variety. Anthocyanins include, for example, cyanidin chloride, delphinidin chloride, malvidin chloride, pelargonin chloride, cyanin chloride, ideain chloride ), keracyanin chloride, kuromanin chloride, pelagonidin chloride, and petunidin chloride (Crozier et al., Nat. Prod. REP. , 2009, 26, 1001-1043). In addition, anthocyanins are effective in antioxidant, anti-allergic, anti-inflammatory, antiviral, antiproliferative, antimutagenic, antibacterial, anticarcinogenic, protection from cardiovascular damage and allergy, protection of microcirculation, protection of peripheral capillary resistance, and improvement of diabetes. (Ghosh & Konishi, Asia Pac J Clin Nutr. , 2007, 16(2):200-8).

"Extract" referred to in the present invention means a material extracted from raw materials by any method, and is used in the sense of including all of the extracted extract, the concentrate obtained therefrom, and the dried product and powder of the concentrate without limitation.

The extract may be obtained by extracting from a raw material or a dried product thereof, and the raw material of the extract may be used without limitation, such as cultivated or commercially available.

When the extract is obtained by extraction from raw materials, all known conventional extraction methods such as solvent extraction, ultrasonic extraction, high pressure extraction, ultra-high pressure extraction, filtration and reflux extraction may be used as the extraction method, preferably solvent extraction or It can be produced by using a reflux extraction method. The extraction process may be repeated several times, and then additional steps such as concentration or lyophilization may be performed. Specifically, the obtained extract may be concentrated under reduced pressure to obtain a concentrate, and the concentrate may be lyophilized and then a high-concentration extract powder may be prepared using a grinder. The extract also includes fractions obtained by further fractionating the extract.

The ginseng berry extract may preferably be extracted by a water-soluble fractionation method, and may be prepared by a manufacturing method including the following steps: 1) preparing an extract by adding an extraction solvent to ginseng berry; 2) filtering the extract of step 1); and 3) drying after concentrating the filtered filtrate of step 2) under reduced pressure.

In one embodiment, the ginseng berry may be one selected from the group consisting of ginseng seeds, fruits, immature fruits, mature fruits, rinds, and combinations thereof, and ginseng berries (all other ginseng seeds except seeds) Or it may include a seed, preferably a rind of a ginseng fruit.

In one embodiment, the ginseng berry extract may be extracted using an aqueous solvent, preferably water, alcohol, or a mixture thereof as a solvent, and more preferably extracted using water as a solvent. The solvent may be added in an amount of 1 to 10 ml per 1 g of the weight of the ginseng berry used for extraction, preferably in an amount of 1 to 5 ml.

The ginseng fruit extract may contain ginsenoside Re. According to the inventors of the present invention, the ginseng fruit extract obtained by extracting ginseng fruit by a water-soluble fractionation method has a higher content of ginsenosides, including ginsenoside Re, than the ginseng root extract, and polycenol (syringa Lecinol, anthocyanin, etc.) were found to contain various active ingredients (Korean Patent Application No. 10-2020-0187943). The inventors have confirmed that the water extract of ginseng fruit extracted by the water-soluble fractionation method as described above contains the largest amount of ginsenoside Re among the active ingredients.

The extraction method of step 1) may be shaking extraction, Soxhlet extraction, reflux extraction, or ultrasonic extraction. In one embodiment of the present invention, the extraction method may be ultrasonic extraction. At this time, the extraction temperature may be 15 to 45 °C, specifically 15 to 35 °C, more specifically 20 to 30 °C. In addition, the extraction pressure may be 200 to 1,000 MPa, specifically 300 to 900 MPa, and more specifically 400 to 800 MPa. The extraction time may be 10 seconds to 1 hour, specifically 20 seconds to 30 minutes, and more specifically 30 seconds to 5 minutes. In addition, the number of extractions may be 1 to 5 times, specifically 1 to 3 times. In a range outside the conditions of the extraction temperature, extraction pressure, extraction time, or number of times of extraction, the content of the active ingredient in the ginseng berry extract may be insignificant because extraction is not sufficiently performed. Alternatively, the efficiency of the extraction operation may decrease because the extraction yield is no longer increased.

The vacuum concentration in step 3) may use a vacuum concentrator or a vacuum rotary evaporator. In addition, the drying may be reduced pressure drying, vacuum drying, boiling drying, spray drying or freeze drying.

Lutein is contained in green leaves along with chlorophyll, and is contained in various flowers as an ester. It is also contained in egg yolk along with zeaxanthin and is the main pigment component of alfalfa extract pigment. Lutein is a type of xanthophylls among carotenoids and is concentrated in the macula, lens, brain, skin, heart, and spinal tissues of the eye. Protects the eyes and reduces free radicals. It cannot be synthesized in the body and must be supplied through food.

Foods that contain a large amount of lutein include spinach, kale, broccoli, lettuce, lettuce, Brussels sprouts, pumpkin, peas, yellow carrots, and some seaweeds. In addition, it is known that flowers such as marigold contain a large amount.

In one embodiment, the lutein may be extracted from natural products including the above-mentioned plants, and may be commercially available lutein for food. The lutein may be a powder or oil suspension, and may be selected from lutein (alone), lutein-zeaxanthin complex extract, lutein ester, and combinations thereof.

When the lutein is extracted from a natural product, hexane, carbon dioxide, and alcohol (fermented alcohol) may be used alone or in combination as solvents. At this time, the extraction temperature may be, for example, 10 to 80 ° C, and, for example, it may be extracted for 1 to 10 hours. The lutein may be characterized in that it is preferably extracted by a supercritical extraction method.

In one embodiment, the composition comprises the ginseng berry extract and the lutein in an amount of 50:1 to 1:1, preferably 10:1 to 1:1, more preferably 5:1 to 1:5, most preferably 2.5: It may be characterized in that it is included in a weight ratio of 1 to 1: 2.5.

'Macular degeneration' of the present invention means that degeneration occurs in the macula, which is a nerve tissue located at the center of the inner retina of the eye, resulting in visual impairment such as visual acuity reduction, central scotoma, metamorphosis (a symptom in which objects appear distorted). Macular degeneration is largely divided into non-exudative (dry) and exudative (wet). There is no special treatment for non-exudative cases, and most of them do not significantly affect vision, whereas exudative cases have a very poor visual prognosis. Among the structures that make up the outside of the eyeball, which exist in order from the outside to the sclera, choroid, and retina, the middle membrane is called the choroid. causes Age-related macular degeneration is the leading cause of blindness in the elderly. In one embodiment, the macular degeneration may be characterized by light-induced death of retinal pigment epithelial cells, A2E photooxidation in retinal pigment epithelial cells, or retinal cell layer damage.

The pharmaceutical composition of the present invention may contain a pharmaceutically acceptable carrier or diluent, and may be prepared according to conventional methods, such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, oral formulations, external preparations, It can be formulated in the form of suppositories and sterile injection solutions. The pharmaceutically acceptable carrier is lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil; and the like. In addition, diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrants, and surfactants are included. Oral solid preparations include tablets, pills, powders, granules, capsules, etc., and these solid preparations contain at least one or more excipients, for example, starch, calcium carbonate, sucrose or lactose. ), gelatin, and the like, and may include lubricants such as magnesium stearate and talc. Oral liquid preparations include suspensions, internal solutions, emulsions, syrups, and the like, and may include diluents such as water and liquid paraffin, wetting agents, sweeteners, aromatics, and preservatives. Parenteral preparations include sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, creams, lyophilized preparations, and suppositories, and non-aqueous solvents and suspensions include vegetable oils, injectable esters such as ethyl oleate, and the like. As a base for the suppository, witepsol, macrogol, tween 61, cacao butter, laurin paper, glycerogelatin, and the like may be used.

The dose of the active ingredient contained in the pharmaceutical composition of the present invention varies depending on the condition and weight of the patient, the severity of the disease, the type of active ingredient, the route and period of administration, and may be appropriately adjusted according to the patient. For example, the pharmaceutical composition may be administered at a dose of 0.0001 to 1000 mg/kg, preferably 0.01 to 100 mg/kg, of the ginseng berry extract and the lutein per day, and the administration is performed once a day. It may be administered once or in several divided doses. In addition, the pharmaceutical composition of the present invention may include the ginseng berry extract and the lutein in a weight percentage of 0.001 to 90% with respect to the total weight of the composition.

The pharmaceutical composition of the present invention is administered to mammals such as rats, mice, livestock, and humans through various routes, for example, oral, dermal, abdominal, rectal or intravenous, intramuscular, subcutaneous, intrauterine intrathecal or intracerebral vascular (intracerebroventricular) It can be administered by injection, preferably orally.

Another aspect of the present invention provides a food composition for preventing or treating macular degeneration comprising a ginseng berry extract and lutein.

The ginseng berry extract, the lutein, the weight ratio of the ginseng berry extract and lutein, macular degeneration, etc. have been described above in the section on the pharmaceutical composition for preventing or treating macular degeneration containing the ginseng berry extract and lutein, and thus will be omitted.

In the food composition according to an embodiment of the present invention, there is no particular limitation on the type of food, for example meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, gum, dairy products, various soups, It may be beverages, tea, drinks, alcoholic beverages, vitamin complexes, and the like, and may include all foods in a conventional sense.

In addition, it can be added to food or beverages for the purpose of preventing, improving or treating macular degeneration. At this time, the amount of the ginseng berry extract and lutein in the food or beverage may be added at 0.01 to 15% by weight of the total food weight, respectively, and the health drink composition is 0.02 to 5 g, preferably 0.3 to 1 g, based on 100 ml. It can be added at a rate of g.

The food composition of the present invention is not particularly limited in other ingredients except for containing the ginseng fruit extract and lutein as essential ingredients in the indicated proportions, and may contain additional ingredients such as various flavoring agents or natural carbohydrates like conventional beverages. can Examples of the aforementioned natural carbohydrates include monosaccharides such as glucose, fructose, and the like; disaccharides such as maltose, sucrose and the like; and polysaccharides such as conventional sugars such as dextrins, cyclodextrins, and the like, and sugar alcohols such as xylitol, sorbitol, and erythritol. Examples of flavoring agents other than those described above include natural flavoring agents such as thaumatin and stevia extracts such as rebaudioside A and glycyrrhizin; and synthetic flavoring agents such as saccharin, aspartame, and the like can advantageously be used. The proportion of the natural carbohydrate is generally about 1 to 20 g, preferably about 5 to 12 g per 100 ml of the composition of the present invention.

In addition to the above, the food composition containing the ginseng fruit extract and lutein of the present invention contains various nutrients, vitamins, minerals (electrolytes), synthetic and natural flavors, colorants and enhancers (cheese, chocolate, etc.), pectic acid and its salts, Alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, and the like may be contained. In addition, the composition containing the ginseng berry extract and the lutein extract of the present invention may contain natural fruit juice, fruit juice beverages, and fruit flesh for preparing vegetable beverages. These components may be used independently or in combination. At this time, the ratio of the additive is not very important, but it is generally selected from 0 to about 20 parts by weight per 100 parts by weight of the composition containing the ginseng berry extract and lutein of the present invention.

The food composition according to one embodiment of the present invention can be applied to humans as well as animals.

Hereinafter, the present invention will be described in more detail through examples. However, the following examples are intended to illustrate the present invention, and the scope of the present invention is not limited thereto.

<Example 1. Preparation of composition for preventing or treating macular degeneration containing ginseng berry extract and lutein>

100 g of ginseng berries harvested from 4-year-old or older ginseng were washed, and the washed ginseng berries were put into a seed separator with about 3 times as much purified water and subjected to a separation process to obtain the skin of ginseng berries excluding seeds. The obtained product, including the peel, was put in water and extracted under reflux at room temperature (around 25℃) for 2 to 5 hours, then filtered, water was evaporated, and powdered ginseng berry extract was obtained through spray drying, which was used until use. Stored at -20°C. The production yield of the ginseng berry extract obtained through the above process was about 2.5%, and the ginseng berry extract was standardized to contain ginsenoside Re at 6%. The ginseng berry extract and lutein (LU) were mixed at a weight ratio of 2:1 and dissolved in 0.5% carboxymethyl cellulose (CMC) at 20% (w/w) to obtain macular degeneration containing the ginseng berry extract and lutein. A composition for sexual prophylaxis or treatment was obtained.

A2E accumulated in human retinal pigment epithelial cells (ARPE-19 cells) is known to induce cytotoxicity when irradiated with blue light (BL), and the cytoprotective ability and cell viability against blue light-induced photooxidation were analyzed as follows <Example 2 > and <Example 3>.

At this time, ARPE-19 cells, cells without any A2E in the cells, were used as the starting material, and the cells were treated with DMEM containing 10% FBS, 100 U/ml penicillin, and 100 mg/ml streptomycin. medium was prepared and cultured at 37°C under humid conditions of 5% CO2.

<Example 2. Effect of a composition for preventing or treating macular degeneration containing pretreated ginseng berry extract and lutein on ARPE-19 cell death by blue light>

Human retinal pigment epithelial cells (ARPE-19 cells) were seeded in a 96-well tissue culture plate at a density of 1 x 10 ⁴ cells/well and cultured for 24 hours. Then, as an experimental group, each test substance was treated prior to A2E accumulation in cultured ARPE-19 cells, and 24 hours later, A2E dissolved in DMEM at 20 μM was treated to accumulate A2E in ARPE-19 cells. After culturing for 24 hours after A2E treatment, blue light (BL, 430 nm, 6000 lux) was irradiated for 20 minutes, and cell viability was measured using a cell count kit after 24 hours. At this time, the group with only A2E accumulation (normal group, Normal) and the group treated with A2E accumulation and blue light (BL) were used as negative and positive control groups, respectively, and the group treated with A2E accumulation, test substance, and blue light was used as the experimental group. Test substances were ginseng berry extract ( GBE 25, 50, or 100 μg/ml) alone, lutein (LU 17) alone, or ginseng berry extract (25 μg/ml) and lutein (10 μg/ml) at different concentrations depending on the group. μM) mixture (GBE 25 + LU 10) was used in the experiment.

As shown in FIG. 1, the cell death caused by blue light irradiation was reduced by pretreatment with the test substance. More specifically, in the case of the control group with only A2E accumulation without pretreatment of the test substance, the cell viability decreased to 68.9% in the blue light treated group (BL) compared to the blue light untreated group (normal group, Normal). On the other hand, in the group treated with ginseng berry extract at 25, 50 and 100 μg/ml, it was 78.9, 82.6, and 84.1%, respectively, compared to the normal group, and 1.15, 1.20, and 1.22 times higher than the blue light treatment group (BL), respectively. It was confirmed that the fruit extract could inhibit apoptosis caused by blue light in a concentration-dependent manner, and in the group treated with lutein at 17 μM, the cell viability was 76.7% compared to the normal group and 1.16 times higher than that of the blue light treated group. It was confirmed that cell death induced by A2E photooxidation according to blue light irradiation could be inhibited. In addition, in the group (GBE 25+ LU 10) treated with 25 μg/ml of ginseng berry extract, the lowest concentration of ginseng berry extract tested, and 10 μM lutein, which is lower than the concentration of lutein tested, the cell viability was higher than that of the normal group. 85%, which is 1.23 times higher than that of the blue light treatment group, higher than that of the four groups treated with the single substance, and cell viability was higher despite mixing each substance at a lower concentration compared to the treatment with the single substance. appeared to be high. Therefore, when the composition of the present invention containing the ginseng berry extract and lutein is pre-treated, cell death induced by A2E photooxidation by blue light can be suppressed, and synergistic effect compared to the case of treatment with the ginseng berry extract or lutein alone. It was confirmed that it has

<Example 3. Effect of post-processed ginseng berry extract and composition for preventing or treating macular degeneration containing lutein on ARPE-19 cell death by blue light>

Human retinal pigment epithelial cells (ARPE-19 cells) were seeded in a 96-well tissue culture plate at a density of 1 x 10 ⁴ cells/well and cultured for 24 hours. A2E was treated at a concentration of 20 μM and cultured for 24 hours, then the experimental group was post-treated with the test substance, and after 24 hours, blue light (6000 lux, 20 min) was irradiated, and then, after 24 hours, cell viability was measured through a cell count kit. . At this time, the group with only A2E accumulation (normal group, Normal) and the group treated with A2E accumulation and blue light (BL) were used as negative and positive control groups, respectively, and the group treated with A2E accumulation, test substance, and blue light was used as the experimental group. Test substances were ginseng berry extract ( GBE 25, 50, or 100 μg/ml) alone, lutein (LU 17) alone, or ginseng berry extract (25 μg/ml) and lutein (10 μg/ml) at different concentrations depending on the group. μM) mixture (GBE 25 + LU 10) was used in the experiment.

As shown in FIG. 2, the cell death caused by blue light irradiation was reduced by the post-treatment of the test substance. More specifically, in the case of the control group with only A2E accumulation without pretreatment of the test substance, the cell viability decreased to 69.94% in the blue light treated group (BL) compared to the blue light untreated group (normal group, Normal). On the other hand, in the group treated with ginseng berry extract at 25, 50 and 100 μg/ml, it was 80.17, 83.70, and 88.18%, respectively, compared to the normal group, and 1.15, 1.20, and 1.27 times higher than the blue light treatment group (BL), respectively. It was confirmed that the fruit extract could inhibit apoptosis caused by blue light in a concentration-dependent manner. In the group treated with 17 μM of lutein, the cell viability was 84.08% compared to the normal group and 1.21 times higher than that of the blue light treated group. It was confirmed that cell death induced by A2E photooxidation according to blue light irradiation could be inhibited. In addition, in the group (GBE 25+ LU 10) treated with 25 μg/ml of ginseng berry extract, the lowest concentration of ginseng berry extract tested, and 10 μM lutein, which is lower than the concentration of lutein tested, the cell viability was higher than that of the normal group. 90%, which is 1.23 times higher than the blue light treatment group, which is higher than the four groups treated with the single substance, and the cell viability is higher even though each substance was mixed at a lower concentration compared to the single substance treatment. appeared to be high. Therefore, in the case of post-treatment with the composition of the present invention containing ginseng berry extract and lutein, cell death induced by A2E photooxidation by blue light can be inhibited, and the increase compared to the case of treatment with ginseng berry extract or lutein alone It was confirmed that it had an effect.

<Example 4. Confirmation of the therapeutic effect of a composition for preventing or treating macular degeneration containing ginseng berry extract and lutein in an animal model of blue light-induced macular degeneration>

Balb/C mice (purchase: DBL) were acclimatized for a week, and then the test substance was administered to the experimental group once a day for 5 days. Irradiation and test substances were administered once a day for 1 hour a day, and then autopsy was performed 24 hours later, and the eyeballs including the optic nerve were excised. The excised ocular tissue was fixed in Davidson solution for 10 days, then fixed in formalin for 1-2 days, and then paraffin blocks were prepared and H&E staining was performed. At this time, the group not treated with blue light and test substance (normal group, Normal) and the group treated with blue light without treatment with test substance (BL) were used as negative and positive control groups, respectively, and the group treated with test substance and blue light was used as the experimental group. . Test substances were ginseng berry extract (GBE100, 100 mg per kg of test animal weight) alone, lutein (LU50, 50 mg per kg of test animal weight) alone, or ginseng berry extract (50 mg per kg of test animal weight), respectively, depending on the group. ) and lutein (25 mg per kg of animal weight) mixture (GBE50+LU25) was used for the experiment.

The stained ocular tissue was observed to confirm the photoreceptor outer nuclear layer (ONL), inner nuclear layer (INL), photoreceptor layer (PL), and whole retina thickness (FIG. 3). As shown in Figure 4 and Table 1, the decrease in retinal layer thickness according to blue light irradiation was suppressed by the treatment of the test substance.

		Normal	BL	BL+ GBE100	BL+ LU50	BL+ GBE50+LU25
outer nuclear layer (ONL)	Thickness (μm)	36.6	18.1	23	25.5	29.5
	% of normal	-	49.5	62.8	69.7	80.6
	% compared to BL	-	-	127.1	140.9	163.0
inner nuclear layer (INL)	Thickness (μm)	26.4	18.3	21	20	24.2
	% of Normal	-	69.3	79.5	75.8	91.7
	% compared to BL	-	-	114.8	109.3	132.2
photoreceptor layer (PL)	Thickness (μm)	23.1	14.4	16	16	16.5
	% of normal	-	62.3	69.3	69.3	71.4
	% compared to BL	-	-	111.1	111.1	114.6
entire retinal layer (whole retina)	Thickness (μm)	139.9	87.7	95	100	114.1
	% of Normal	-	62.7	67.9	71.5	81.6
	% compared to BL	-	-	108.3	114.0	130.1

More specifically, in the case of the control group not treated with the test substance, the photoreceptor outer nuclear layer (ONL), inner nuclear layer (INL), and photoreceptor in the blue light treated group (BL) compared to the blue light untreated group (Normal group, Normal). Layer (PL) and whole retina thickness decreased to 45.5, 69.3, 62.3 and 62.7%, respectively. On the other hand, in the group treated with 100 mg/kg of ginseng berry extract (GBE100), ONL, INL, PL, and whole retina thickness were 62.8, 79.5, 69.3, and 67.9%, respectively, compared to the normal group. In the group (LU50), ONL, INL, PL, and whole retina thicknesses were 69.7, 75.8, 69.3, and 71.5%, respectively, compared to the normal group, and about 8 to 40% higher than the blue light treatment group. In addition, ONL, INL, PL and whole retina thickness were 80.6, 91.7, 71.4, and 81.6% compared to the normal group, which was about 9 to 60% higher than the blue light treatment group, and not only higher than each group treated with the single substance, but also treated with the single substance. Even though each substance was mixed at a lower concentration than when compared to the time, the effect of suppressing the decrease in retinal layer thickness was higher. Therefore, it was confirmed that the composition of the present invention containing the ginseng berry extract and lutein can inhibit damage to the retinal layer caused by blue light, and has a higher effect than the treatment with the ginseng berry extract or lutein alone.

<Example 5. Effect of composition for preventing or treating macular degeneration containing ginseng berry extract and lutein on NF-κB signaling in retinal pigment epithelial cells (ARPE-19)>

NF-κB is a protein complex, and the NF-κB/Rel heterodimer exists in the cytoplasm in a state bound to IκB, and when IκB is decomposed, the heterodimer enters the nucleus and induces gene transcription. Genes regulated by NF-κB are known to be related to inflammation/immune response, angiogenesis, cell survival or cell proliferation. In order to investigate how the composition containing the ginseng berry extract and lutein of the present invention affects NF-κB signaling, the amounts of NF-κB and IκB in retinal pigment epithelial cells (ARPE-19) were examined.

Human retinal pigment epithelial cells (ARPE-19 cells) were seeded in a 96-well tissue culture plate at a density of 1 x 10 ⁴ cells/well and cultured for 24 hours. A2E was treated at a concentration of 20 μM and cultured for 24 hours, then the experimental group was post-treated with the test substance, and after 24 hours, blue light (6000 lux, 20 min) was irradiated, and then, after 24 hours, cell viability was measured through a cell count kit. . At this time, the group with only A2E accumulation (normal group, Normal) and the group treated with A2E accumulation and blue light (BL) were used as negative and positive control groups, respectively, and the group treated with A2E accumulation, test substance, and blue light was used as the experimental group. Test substances were ginseng berry extract (GBE 50 μg/ml) alone, lutein (LU 30 μM) alone, or ginseng berry extract (25 μg/ml) and lutein (15 μM) mixture at different concentrations depending on the group. (GBE 25 + LU 15).

Cells were subjected to cell lysis and nuclear/cytoplasmic fractionation according to Rosner & Hengstschlaeger ( Hum. Mol. Genet. , 2008), and proteins were extracted for each fraction. In this regard, Western blot was performed to confirm the amount of NF-κB in the nucleus and the cytoplasm and the expression of IκB-α in the cytoplasm, respectively. At this time, the group not treated with blue light and test substance (normal group, Normal) and the group treated with blue light without treatment with test substance (control) were used as negative and positive control groups, respectively, and the group treated with test substance and blue light was used as the experimental group. .

As a result, the amount of NF-κB in the nucleus increased 3.48-fold in the blue light treated group (control) compared to the normal control group (FIG. 5B), and the expression in the cytoplasm was decreased correspondingly (FIG. 5A). In the case of the blue light treatment group administered with the test substance, NF-κB in the nucleus increased compared to the normal group, but compared to the blue light treatment group (control) not administered with 50 μg/ml ginseng berry extract and 30 μM lutein, In each group, the levels were low at 59% and 67%, respectively, and in the group administered with the composition of the present invention containing 25 μg/ml of ginseng berry extract and 15 μM of lutein, which are half of the concentrations of each single test substance, the nucleus It was confirmed that my NF-κB was lower than that of the blue light treatment group (control) at a level of 50% (FIG. 5B).

As a result of confirming the amount of IκB in the cytoplasm (FIG. 6), the amount of IκB decreased to 58% in the blue light treatment group (control) compared to the normal control group, but was higher than the control group in the blue light treatment group administered with the test substance. . That is, compared to the blue light treatment group (control), the amount of IκB in the cytoplasm was higher in the group administered with 50 μg/ml ginseng berry extract and 30 μM lutein by 150% and 129%, respectively. In particular, 25 μg/ml ginseng berry extract In the group administered with the composition of the present invention containing the extract and 15 μM of lutein, the amount of IκB in the cytoplasm was higher than that of the blue light-treated group (control) by 160%.

Taken together, the composition containing the ginseng berry extract and lutein of the present invention inhibits the degradation of IκB, thereby further enhancing the effect of the ginseng berry extract and lutein alone, which allows NF-κB to move less from the cytoplasm to the nucleus. It was confirmed that it could be raised.

<Example 6. Effect of composition for preventing or treating macular degeneration containing ginseng berry extract and lutein on NF-κB target gene expression in retinal pigment epithelial cells (ARPE-19)>

As described above, it has been reported that NF-κB can induce the expression of target genes when it moves from the cytoplasm to the nucleus. In order to confirm whether the inhibition of NF-κB migration by the composition of the present invention actually led to an effect on the expression of target genes, IL-1β and IL-6 among the target genes in retinal pigment epithelial cells (ARPE-19) , mRNA expression levels of TNFα and VEGFα were confirmed.

Human retinal pigment epithelial cells (ARPE-19 cells) were seeded in a 96-well tissue culture plate at a density of 1 x 10 ⁴ cells/well and cultured for 24 hours. A2E was treated at a concentration of 20 μM and cultured for 24 hours, then the experimental group was post-treated with the test substance, and after 24 hours, blue light (6000 lux, 20 min) was irradiated, and then, after 24 hours, cell viability was measured through a cell count kit. . At this time, the group with only A2E accumulation (normal group, Normal) and the group treated with A2E accumulation and blue light (control) were used as negative and positive control groups, respectively, and the group treated with A2E accumulation, test substance, and blue light was used as the experimental group. Test substances were ginseng berry extract (GBE 50 μg/ml) alone, lutein (LU 30 μM) alone, or ginseng berry extract (25 μg/ml) and lutein (15 μM) mixture at different concentrations depending on the group. (GBE 25 + LU 15). RNA was extracted from cells for each group using TRIzol agent (Invitrogen), and cDNA was synthesized using quantified 1 μg/μl RNA and a reverse transcription system (Promega). The expression patterns of each gene were measured using the synthesized cDNA and pre-designed primers and probes (Applied biosystems) for each gene. PCR reaction and analysis were performed using a Rotor-Gene 3000 system (Corbett Research, Sydney, Australia).

As a result, in the case of IL-1β, whose expression is known to be induced through NF-κB by tissue/cell damage, the mRNA expression level is nearly 4 times higher in the blue light treated group than in the normal control group. However, in the blue light treatment group administered with the test substance, it decreased compared to the control group (FIG. 7). That is, in the group administered with 50 μg/ml ginseng berry extract and 30 μM lutein, the expression level of IL-1β mRNA was 44% and 60%, respectively, lower than that of the blue light treatment group (control), and 25 μg/ml In the group administered with the composition of the present invention containing the ginseng berry extract and 15 µM of lutein, the IL-1β mRNA expression level was as low as 44% of the blue light treated group (control).

IL-6 is secreted by T cells and macrophages to stimulate immune responses, and is known to mediate, in particular, fever and rapid immune responses. In RPE cells, IκB degradation is induced by oxidative stress, and it is known that NF-κB moves into the nucleus to induce IL-6 expression. 6 mRNA expression level was as high as nearly three times (FIG. 8). Compared to the blue light treatment group (control), the IL-1β mRNA expression level was lowered by 41% and 79%, respectively, in the group administered with 50 μg/ml ginseng berry extract and 30 μM lutein, respectively. In particular, 25 μg/ml ginseng In the group administered with the composition of the present invention containing fruit extract and 15 μM lutein, the expression level of IL-1β mRNA was 34% of that of the blue light-treated group (control), which was the normal control group not treated with blue light. is at a similar level to

TNFα is mainly secreted by macrophages, but also secreted by other cell types such as CD4+ lymphocytes, NK cells, neutrophils, mast cells, eosinophils, and neurons, and activates NF-κB, MAPK signaling, and death signaling. It is known to act as an inducer. TNFα was higher in the blue light treatment group (control) than in the normal control group, so that the mRNA expression level was nearly 4 times higher, but it was lower than the control group in the blue light treatment group administered with the test substance (FIG. 9). That is, in the group administered with 50 μg/ml ginseng berry extract and 30 μM lutein, the IL-1β mRNA expression levels were lower at 36% and 48%, respectively, compared to the blue light treatment group (control), and the 25 μg/ml In the group administered with the composition of the present invention containing ginseng berry extract and 15 μM of lutein, the expression level of IL-1β mRNA was as low as 31% of the blue light treated group (control), which was compared to the normal control group not treated with blue light ( It was confirmed that the level was similar to normal).

VEGFα, which mediates angiogenesis, was found to be nearly 4 times higher in the blue light-treated group compared to the normal control group, whereas in the blue light-treated group treated with the test substance, the expression level was lower than that in the control group. level was shown (FIG. 10). That is, in the group administered with 50 μg/ml ginseng berry extract and 30 μM lutein compared to the blue light treatment group (control), the VEGFα mRNA expression level was lowered by 36% and 41%, respectively. In particular, 25 μg/ml ginseng In the group administered with the composition of the present invention containing fruit extract and 15 μM lutein, the expression level of VEGFα mRNA was as low as 31% of the blue light treatment group (control).

Taken together, the composition for preventing or treating macular degeneration comprising a ginseng berry extract and lutein according to an embodiment of the present invention is effective for the movement of NF-κB from the cytoplasm to the nucleus, degradation of IκB, and expression of IL-1β in retinal cells. , IL-6 expression, TNFα expression and VEGFα expression can be reduced, and through this, abnormal inflammatory responses, apoptosis and angiogenesis can be reduced to induce prevention, improvement or treatment of macular degeneration. In addition, the composition can be expected to prevent, improve, and treat macular degeneration by inhibiting light-induced death of retinal pigment epithelial cells, A2E photooxidation in retinal pigment epithelial cells, and retinal cell layer damage.

The above description of the present invention is for illustrative purposes, and those skilled in the art can understand that it can be easily modified into other specific forms without changing the technical spirit or essential features of the present invention. will be. Therefore, the embodiments described above should be understood as illustrative in all respects and not limiting. For example, each component described as a single type may be implemented in a distributed manner, and similarly, components described as distributed may be implemented in a combined form.

The scope of the present invention is indicated by the following claims, and all changes or modifications derived from the meaning and scope of the claims and equivalent concepts should be interpreted as being included in the scope of the present invention.

Modes for carrying out the invention have been described together in the best mode for carrying out the invention.

The composition containing the ginseng berry extract and lutein according to an embodiment of the present invention is effective for the movement of NF-κB from the cytoplasm to the nucleus of retinal cells, the degradation of IκB, IL-1β expression, IL-6 expression, TNFα expression and VEGFα expression It can reduce abnormal inflammatory response, apoptosis, and angiogenesis through which it can induce prevention, improvement, or treatment of macular degeneration, which can be used in macular degeneration disease treatment and health functional food industry.

Claims (7)

1. A pharmaceutical composition for preventing or treating macular degeneration comprising a ginseng berry extract and lutein.
2. According to claim 1,

   The ginseng fruit extract is Korean ginseng ( Panax ginseng ), Hoegi ginseng ( P. quiquefolius ), Jeonchilseng ( P. notoginseng ), bamboo shoots ( P. japonicus ), trifolium ginseng ( P. trifolium ), Himalayan ginseng ( P. pseudoginseng ), A pharmaceutical composition for preventing or treating macular degeneration, characterized in that it is an extract extracted from the fruit of one or more species of ginseng selected from the group consisting of Vietnamese ginseng ( P. vietnamensis ) and American ginseng ( Panax quinquefolium ).
3. According to claim 1,

   The ginseng berry extract is a pharmaceutical composition for preventing or treating macular degeneration, characterized in that extracted with water as a solvent.
4. According to claim 1,

   The lutein is a pharmaceutical composition for preventing or treating macular degeneration, characterized in that extracted from natural products by supercritical extraction.
5. According to claim 1,

   The composition is a pharmaceutical composition for preventing or treating macular degeneration, characterized in that it comprises the ginseng berry extract and the lutein in a weight ratio of 5: 1 to 1: 5.
6. According to claim 1,

   The macular degeneration is a pharmaceutical composition for preventing or treating macular degeneration, characterized in that by death of retinal pigment epithelial cells induced by light, A2E photooxidation or retinal cell layer damage in retinal pigment epithelial cells.
7. A food composition for preventing or treating macular degeneration comprising a ginseng berry extract and lutein.

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