KR102406452B1 - Composition for Preventing, Improving or Treating Macular Degeneration Comprising Compound K and Lutein - Google Patents

Abstract

The present invention relates to a composition for preventing, improving or treating macular degeneration comprising compound K and lutein as active ingredients. When the compound K and lutein are administered in combination, there is an effect of inhibiting the occurrence of macular degeneration, and thus it can be usefully utilized for the prevention, improvement and treatment of macular degeneration.

Description

Composition for preventing, improving or treating macular degeneration comprising compound K and lutein {Composition for Preventing, Improving or Treating Macular Degeneration Comprising Compound K and Lutein}

The present invention relates to a composition for preventing, improving or treating macular degeneration comprising compound K and lutein.

The nerve tissue located in the center of the retina is called the macula. This area has a fairly dense concentration of photoreceptors and is responsible for central vision as an object image is formed and contributes to the sharpness and accuracy of the field of view. The macula can be degenerated by causes such as aging, damage caused by ultraviolet or blue light, genetic factors, underlying diseases (diabetes, dyslipidemia, high cholesterol, etc.), toxicity and inflammation, which can cause visual impairment. It is said Macular degeneration is one of the main causes of blindness along with cataracts, glaucoma, and diabetic retinopathy. Degeneration occurs in the macula, and the central vision is damaged in the early stages, causing the central vision to appear blurred or distorted. In severe cases, it leads to blindness It is a serious disease.

There are two main types of macular degeneration: dry (non-exudative) macular degeneration and wet (exudative) macular degeneration. Dry macular degeneration accounts for about 90% of patients, and dry macular degeneration is characterized by atrophy or thinning of the macular tissue due to the accumulation of a waste product called drusen between the retinal pigment epithelium and Bruch's membrane. do. On the other hand, wet macular degeneration accounts for about 10% of all patients, and normal blood and nutrients are not supplied to the macula, abnormally new blood vessels are formed, rupture, and blood or mucus leaks into the macula, causing serious vision loss. say The initial symptoms of dry macular degeneration are asymptomatic or even if there are symptoms, they are considered presbyopia, so it is easy to neglect. However, if the progression of dry macular degeneration is not suppressed or treated, vision loss due to persistent retinal damage can come, and it can progress to wet macular degeneration and even lead to blindness.

Lutein is one of the xanthophylls, one of 600 naturally occurring carotenoids known to date. Lutein is found in large quantities in leafy vegetables such as spinach, kale, and yellow carrots, and is known to be concentrated in the macula, a small area of the retina responsible for three-dimensional vision.

Therefore, although lutein is taken for the prevention and treatment of macular degeneration, long-term overdose may increase the probability of developing eye diseases such as macular degeneration.

One object of the present invention is to provide a pharmaceutical composition for preventing or treating macular degeneration comprising compound K and lutein.

Another object of the present invention is to provide a food composition for preventing or improving macular degeneration comprising compound K (compound K) and lutein.

In order to achieve the above object, one aspect of the present invention is to provide a pharmaceutical composition for preventing or treating macular degeneration comprising compound K and lutein.

In the present invention, compound K refers to the state in which the macromolecular compound ginsenoside, a saponin of ginseng, is broken down into small pieces for easy eating by cells in the intestine, that is, the final metabolite, and can be separated from the ginseng extract, and can be purchased on the market. However, the present invention is not limited thereto.

The ginseng may be at least one selected from the group consisting of wild ginseng, wild ginseng, water ginseng, whole ginseng, hwagi ginseng, cultivar root, and ginseng root. However, the present invention is not limited thereto.

The ginseng extract can be obtained by extracting ginseng with one or more solvents selected from the group consisting of water, C1 to C4 lower alcohol, acetone, hexane, tichloromethane and ethyl acetate, and the C1 to C4 lower alcohol may be methanol, ethanol, propanol, isopropanol, butanol, or the like. Preferably it is ethanol.

The compound K can be obtained by fractionation of the ginseng extract by chromatography, the chromatography is silica gel column chromatography (silicagel vacuum liquid column chromatography), RP-18 column chromatography (RP-18 column chromatography), HP-20 column Chromatography (HP-20 column chromatography), preparative reversed-phase high performance chromatography, medium pressure liquid chromatography, high-performance liquid chromatography You can choose to use it, etc.

In the present invention, lutein is a pigment present in the largest amount among xanthophylls of carotenoids, and may exist in a large amount together with β-carotene in the chloroplast of plants, and may be purchased commercially, but is not limited thereto. In addition, as an important component of the retina, it has an effect of inhibiting macular degeneration, but excessive intake for a long period of time may cause macular degeneration.

In the present invention, the term "prevention" refers to any action that inhibits the onset of macular degeneration by administering the composition to an individual.

In the present invention, the term "treatment" refers to any action to improve or benefit symptoms of macular degeneration by administering the composition to an individual.

In the present invention, the pharmaceutical composition can be formulated and used in the form of oral dosage forms such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, external preparations, suppositories, or sterile injection solutions, respectively, according to a conventional method. have. In detail, when formulating, it may be prepared using a diluent or excipient such as a filler, a weight agent, a binder, a wetting agent, a disintegrant, and a surfactant commonly used. Solid preparations for oral administration include, but are not limited to, tablets, pills, powders, granules, capsules, and the like. Such a solid preparation may be prepared by mixing at least one excipient, for example, starch, calcium carbonate, sucrose, lactose, gelatin, etc., with the composition or pharmaceutically acceptable salt thereof comprising the compound K and lutein. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized formulations and tasks. Non-aqueous solvents and suspending agents include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate. As the base of the suppository, Witepsol, Macrosol, Tween 61, cacao butter, laurin fat, glycerogelatin, etc. can be used.

A suitable dosage of the composition of the present invention varies depending on the condition and weight of the patient, the severity of the disease, the drug form, and time, but may be appropriately selected by those skilled in the art.

According to one embodiment of the present invention, the compound K may be a pharmaceutical composition for preventing or treating macular degeneration that reduces the side effects that appear when using lutein.

The side effect of using lutein may be that when lutein is treated, cell death or macular degeneration is accelerated. The treatment of lutein may be treatment or continuous administration of lutein at a high concentration, but is not limited thereto.

Reducing the side effects that appear when using the lutein may be to delay or prevent cell death or accelerated macular degeneration.

According to one embodiment of the present invention, the composition may be a pharmaceutical composition for preventing or treating macular degeneration in which the weight ratio of compound K and lutein is 1:5 to 1:50. More preferably, the weight ratio of compound K and lutein may be 1:5 to 1:10.

Another aspect of the present invention is to provide a food composition for preventing or improving macular degeneration comprising compound K and lutein.

In the present invention, the term "improvement" refers to any action to improve or benefit symptoms of macular degeneration by administering the composition to an individual.

In addition to compound K and lutein as active ingredients, the food composition of the present invention may include ingredients commonly added during food production, for example, proteins, carbohydrates, fats, nutrients, seasonings and flavoring agents. can do. Examples of carbohydrates include monosaccharides such as glucose, fructose, and the like; disaccharides such as maltose, sucrose, oligosaccharides and the like; and polysaccharides, for example, conventional sugars such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As the flavoring agent, natural flavoring agents [taumatine, stevia extract (eg, rebaudioside A, glycyrrhizin, etc.)] and synthetic flavoring agents (saccharin, aspartame, etc.) can be used.

For example, when the food composition of the present invention is prepared as a drink, in addition to the compound K and lutein of the present invention, citric acid, high fructose, sugar, glucose, acetic acid, malic acid, fruit juice, cephalopod extract, jujube extract and/or licorice extract, etc. This may be further included.

In addition, the food composition of the present invention includes various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and thickeners (cheese, chocolate, etc.), pectic acid and its salts, alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated beverages, and the like.

These components may be used independently or in combination, and the ratio of these additives may be selected from 0 to about 20 parts by weight per 100 parts by weight of the food composition of the present invention, but is not limited thereto.

According to one embodiment of the present invention, the compound K may be a food composition for preventing or improving macular degeneration that reduces the side effects that appear when using lutein.

According to one embodiment of the present invention, the composition may be a food composition for preventing or improving macular degeneration in which the weight ratio of compound K and lutein is 1:5 to 1:50. More preferably, the weight ratio of compound K and lutein may be 1:5 to 1:10.

The present invention relates to a composition for preventing, improving, or treating macular degeneration, comprising compound K and lutein as active ingredients, wherein when the compound K and lutein are administered in combination, the occurrence of macular degeneration and the occurrence of side effects causing macular degeneration through lutein Since it has an inhibitory effect, it can be usefully used for the prevention, improvement and treatment of macular degeneration.

1 is a result of confirming the cell viability after the concentration-dependent treatment of lutein in RPE cells. (2: H ₂ O ₂ , 3: H ₂ O ₂ with lutein (100 μM), 4: H ₂ O ₂ with lutein (200 μM), 5: H ₂ O ₂ with lutein (500 μM))
2 is a result of confirming the activity of caspase-3 after RPE cells are treated with lutein in a concentration-dependent manner. (2: H ₂ O ₂ , 3: H ₂ O ₂ with lutein (100 μM), 4: H ₂ O ₂ with lutein (200 μM), 5: H ₂ O ₂ with lutein (500 μM))
Figure 3 shows that RPE cells were mixed with high concentration lutein (500 μM), high concentration lutein (500 μM) and concentration-dependently compound K (1 μM, 2 μM, 10 μM), respectively, and then the viability of the cells was checked and compound K showed lutein side effects. It is the result of confirming suppression. (2: H ₂ O ₂ , 3: H ₂ O ₂ with lutein (500 μM), 4: H ₂ O ₂ , lutein (500 μM), compound K (1 μM), 5: H ₂ O ₂ , lutein (500 μM), Compound K (2 μM), 6: H ₂ O ₂ , Lutein (500 μM), Compound K (10 μM))
Figure 4 shows RPE cells mixed with high concentration lutein (500 μM), high concentration lutein (500 μM) and concentration-dependently compound K (1 μM, 2 μM, 10 μM), respectively, after confirming the activity of caspase-3, compound K is lutein It is the result of confirming the suppression of side effects. (2: H ₂ O ₂ , 3: H ₂ O ₂ with lutein (500 μM), 4: H ₂ O ₂ , lutein (500 μM), compound K (1 μM), 5: H ₂ O ₂ , lutein (500 μM), Compound K (2 μM), 6: H ₂ O ₂ , Lutein (500 μM), Compound K (10 μM))
Figure 5 shows the results of treatment with lutein and compound K alone in RPE cells in a concentration-dependent manner, and after the concentration-dependent treatment of compound K with lutein (100 μM), the cell viability was confirmed, and the mixture of lutein and compound K was better This is the result of confirming the effect. (2: H ₂ O ₂ , 3: H ₂ O ₂ with lutein (1 μM), 4: H ₂ O ₂ with lutein (10 μM), 5: H ₂ O ₂ with lutein (50 μM), 6: H ₂ O ₂ with lutein (100 μM), 7: H ₂ O ₂ with compound K (1 μM), 8: H ₂ O ₂ with compound K (2 μM), 9: H ₂ O ₂ with compound K (10 μM), 10: H ₂ O ₂ , lutein (100 μM), compound K (1 μM), 11: H ₂ O ₂ , lutein (100 μM), compound K (2 μM), 12: H ₂ O ₂ , lutein (100 μM), compound K (10 μM))
6 shows that RPE cells were treated with lutein and compound K alone, respectively, and after treatment with lutein (100 μM) with compound K in a concentration-dependent manner, caspase-3 activity was confirmed, and the mixture of lutein and compound K was better This is the result of confirming the effect. (2: H ₂ O ₂ , 3: H ₂ O ₂ with lutein (1 μM), 4: H ₂ O ₂ with lutein (10 μM), 5: H ₂ O ₂ with lutein (50 μM), 6: H ₂ O ₂ with lutein (100 μM), 7: H ₂ O ₂ with compound K (1 μM), 8: H ₂ O ₂ with compound K (2 μM), 9: H ₂ O ₂ with compound K (10 μM), 10: H ₂ O ₂ , lutein (100 μM), compound K (1 μM), 11: H ₂ O ₂ , lutein (100 μM), compound K (2 μM), 12: H ₂ O ₂ , lutein (100 μM), compound K (10 μM))
7 is a result of confirming the optimal ratio showing the effect by checking the cell viability after the concentration-dependent treatment of compound K in lutein (100 μM) in RPE cells. (2: H ₂ O ₂ , 3: H ₂ O ₂ with lutein (100 μM), 4: H ₂ O ₂ , lutein (100 μM), compound K (2 μM), 5: H ₂ O ₂ , lutein (100 μM), Compound K (10 μM), 6: H ₂ O ₂ , Lutein (100 μM), Compound K (20 μM), 7: H ₂ O ₂ , Lutein (100 μM), Compound K (50 μM))
8 is a result of confirming the optimal ratio showing the effect by confirming the activity of caspase-3 after the concentration-dependent treatment of compound K in lutein (100 μM) in RPE cells. (2: H ₂ O ₂ , 3: H ₂ O ₂ with lutein (100 μM), 4: H ₂ O ₂ , lutein (100 μM), compound K (2 μM), 5: H ₂ O ₂ , lutein (100 μM), Compound K (10 μM), 6: H ₂ O ₂ , Lutein (100 μM), Compound K (20 μM), 7: H ₂ O ₂ , Lutein (100 μM), Compound K (50 μM))

Hereinafter, one or more specific examples will be described in more detail through examples. However, these examples are for illustrative purposes of one or more embodiments, and the scope of the present invention is not limited to these examples.

Experimental Method 1: Cell Preparation and Sample Processing

Retinal pigment epithelium (RPE) cells purchased from ATCC were used. The cells were cultured in a CO ₂ incubator maintained at 37 ° C. using DMEM (Dulbecco's modified Eagle's medium) medium containing 10% fetal bovine serum (FBS) and 1% penicillin streptomycin. It was used in this experiment. The prepared cells were treated by dissolving H ₂ O ₂ prepared above, lutein, compound K or a mixture of lutein and compound K at an appropriate concentration in PBS or medium.

Experimental method 2: Confirmation of cell viability

Cell viability was confirmed using flow cytometry analysis. To this end, 100 μl of PBS was added to the cells for resuspension, and 200 μl of 95% ethanol was added to vortex the cells, followed by incubation at 4° C. for 1 hour. Thereafter, the cells were washed with PBS, resuspended by adding 250 μl of a 1.12% sodium citrate solution of pH 8.4 containing 12.5 μg ribonuclease (RNase), and incubated at 37° C. for an additional 30 minutes. did

After that, 100 μl of MTT solution with a concentration of 35.4 μg/ml was added to the cells, and the cells were stained for 1 hour at room temperature. did

Experimental method 3: Confirmation of Caspase-3 activity

After dissolving the cells treated with H ₂ O ₂ , lutein, compound K, or a mixture of lutein and compound K, caspase-3 activity was assayed using a caspase-3 colorimetric assay kit (Abcam). Confirmed. First, retinal pigment epithelial cells were treated with a lysis buffer containing protease inhibitor cocktail (Sigma-Aldrich) (50 mM Tris (pH 7.4), 150 mM NaCl, 1 mM EDTA, 1% nonyl phenoxypolyethoxylethanol-40) -40, NP-40))), and then color development was induced using the color buffer included in the kit, and the fluorescence intensity was measured at 540 nm to evaluate the degree of caspase-3 activity.

Example 1. Lutein toxicity confirmation

In order to check the effect on the cells when lutein is administered in excess, lutein was administered to the cells in a concentration-dependent manner, and the cell viability and the activity of caspase-3, a marker of apoptosis, were checked.

Cell viability and caspase-3 activity are shown in FIGS. 1 and 2, respectively.

In the case of lutein, the cytoprotective effect was maintained with respect to H ₂ O ₂ up to 200 μM, but when treated with 500 μM, cell viability decreased and caspase-3 activity was increased. Therefore, it was shown that a high concentration of lutein causes apoptosis rather than administration.

Example 2. Confirmation of Lutein Side Effects Alleviation Effect of Compound K

In the side effects caused by the high concentration of lutein, in order to confirm the effect of compound K, after the concentration-dependent treatment of compound K with high concentration of lutein (500 μM), the viability of cells and the activity of caspase-3 were confirmed.

Cell viability and caspase-3 activity are shown in FIGS. 3 and 4, respectively.

As a result, it was confirmed that the cell viability was increased and the activity of caspase-3 was inhibited when compound K was simultaneously treated than when treated with high concentration of lutein. Therefore, it was confirmed that compound K could significantly reduce the side effects of lutein in a concentration-dependent manner.

Example 3. Confirmation of synergistic effect between Lutein and Compound K

In order to confirm the synergistic effect of lutein and compound K, a mixture of lutein, compound K, lutein and compound K was administered to each cell in a concentration-dependent manner, and the cell viability and the activity of caspase-3, a marker of apoptosis, were checked. did

Cell viability and caspase-3 activity are shown in FIGS. 5 and 6, respectively.

Both lutein and compound K showed cytoprotective effects in a concentration-dependent manner. On the other hand, when compound K was treated concentration-dependently on the basis of 100 μM lutein, a higher concentration showed a significant effect compared to the case of conventional lutein or compound K treatment.

Example 4. Confirmation of Optimal Capacity of Lutein and Compound K

After mixing lutein and compound K to confirm the optimal treatment concentration during treatment, compound K (Compound K) was treated in a concentration-dependent manner based on lutein 100 μM, and cell viability and caspase-3 activity were checked.

Cell viability and caspase-3 activity are shown in FIGS. 7 and 8, respectively.

When lutein 100 μM and compound K were mixed and treated by concentration, compound K showed a concentration-dependent effect up to 20 μM, but when compound K was 50 μM, there was no significant change in cell survival and caspase-3 activity.

Claims (6)

A pharmaceutical composition for preventing or treating macular degeneration comprising compound K and lutein,
The compound K (compound K) is to reduce the side effects that appear when using lutein,
A pharmaceutical composition for preventing or treating macular degeneration.
delete According to claim 1, wherein the composition is a pharmaceutical composition for preventing or treating macular degeneration in which the weight ratio of compound K and lutein is 1:5 to 1:50.
As a food composition for preventing or improving macular degeneration comprising compound K and lutein,
The compound K (compound K) is to reduce the side effects that appear when using lutein,
A food composition for preventing or improving macular degeneration.
delete In claim 4, wherein the composition is a food composition for preventing or improving macular degeneration in which the weight ratio of compound K and lutein is 1:5 to 1:50.

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