KR100665504B1 - Composition comprising an extract of Aruncus dioicus. for the prevention and treatment of ischemic diseases and degenerative brain diseases - Google Patents

Abstract

The present invention is a snow horse riding horse having improved cell survival under hypoxic conditions ( Aruncus dioicus ( Walt. ) Fern var . The present invention relates to a composition containing the extract, and more particularly, the crude extract and the non-polar solvent soluble extract of the present invention inhibit apoptosis in ischemic state to prevent and treat ischemic disease and degenerative brain disease. And health functional foods.

Eyelid Riding, Nonpolar Solvent, Apoptosis, Ischemic Disease, Degenerative Brain Disease

Description

Composition for the prevention and treatment of ischemic diseases and degenerative brain diseases, including eyelid riding extract {Composition comprising an extract of Aruncus dioicus. for the prevention and treatment of ischemic diseases and degenerative brain diseases}             

1 is a diagram comparing the cell survival results under hypoxic and normal oxygen conditions of the cell line and the added cell line without the addition of eyelids.

Figure 2a is a diagram showing the results of DNA fragmentation analysis of the cell line without the addition of crude eye horse riding extract to the culture medium in hypoxic conditions,

Figure 2b is a diagram showing the results of DNA fragmentation analysis of the cell line added crude eye horse riding extract to the culture medium in hypoxic conditions,

Figure 3 is a diagram showing the effect of each fraction by performing an MTT assay (MTT assay) for each cell line to which the addition of crude horseback riding crude extract or non-polar solvent soluble extract in different culture medium under hypoxic conditions,

Figure 4a is a diagram comparing the results of the triphenyl tetrazolium chloride (TTC) staining of the cerebral infarction model and the injected cerebral infarction model without the injection of crude eyelids,

Figure 4b is a diagram showing the TTC staining results of the cerebral infarction model not injected with crude eyelids,

Figure 4c is a diagram showing the TTC staining results of the cerebral infarction model injecting the crude horseback riding extract,

5 is a diagram comparing the results of TTC staining of the cerebral infarction model and the injected cerebral infarction model that is not injected with the eyelid riding non-polar solvent soluble fraction.

The present invention relates to an eyelid equestrian extract having a therapeutic and prophylactic effect of ischemic diseases and degenerative brain diseases by inhibiting apoptosis.

Cerebral infarction and myocardial infarction, which are typical ischemic diseases, are blocked by atherosclerosis in which blood vessels are narrowed due to hypertension, hyperlipidemia, diabetes, and smoking, and the coronary arteries that supply blood to the brain by blood clots or coronary arteries that supply blood to the heart are blocked. It is caused by necrosis of surrounding tissues.

Cardiovascular disease accounts for 30% of all mortality in the world, leading the mortality rate, and 75% of these are caused by ischemic diseases such as myocardial infarction and cerebral infarction. It is coming true. The methods of reducing the mortality caused by myocardial infarction and cerebral infarction include the treatment of hypertension and hyperlipidemia to prevent the blockage of blood vessels and the reduction of necrosis of surrounding tissues during the occlusion.

The best way to reduce the necrotic area is to reperfusion the blood vessels by thrombolytically blocking the blood vessels in the earliest time, but within 3-6 hours of clogging the blood vessels as the heart and brain gain energy by breathing. Necrosis of these cells and subsequent reperfusion can not expect the therapeutic effect. However, it is practically difficult to get to the hospital within 3-6 hours of occlusion and reperfusion, and since the heart and brain are not able to regenerate once they have been damaged, it is possible to improve cell viability in ischemia until reperfusion at the hospital. The effect can be enhanced. At this time, the cause of tissue necrosis is apoptosis (Crow MT et al., Circ.Res ., 95 , pp957-970, 2004; Friedlander RM et al., N. Engl. J. Med. , 348 , pp1365- 1375, 2003), by inhibiting apoptosis can improve cell viability.

In addition, in the case of surgery to stop the heart, if the amount of oxygen required more than the amount of oxygen supplied by the cardiopulmonary system, myocardial damage or brain damage due to hypotension may occur. Heart failure due to ischemia and myocardial injury and brain injury, for example, by bypass graft surgery during coronary artery occlusion, and aneurysm surgery in the cerebral artery or aorta. Side effects such as involuntary return may occur. Indeed, surgical or interventional treatment of aortic aneurysms causes side effects such as myocardial ischemia, kidney failure, and lower extremity palsy in 3-16% of patients. Therefore, if a drug that inhibits apoptosis in ischemic conditions is used for pretreatment, the side effects can be reduced.

The cause of neuronal suicide is not yet clear, but when the cerebral ischemia is transiently induced, the supply of oxygen and glucose is blocked, resulting in a decrease in ATP and edema in neurons, leading to neuronal suicide. Known. Neuronal suicide mechanism by cerebral ischemia is an excitatory neuronal suicide mechanism in which excess glutamate accumulates outside the cell by ischemia, and this glutamic acid enters the cell, resulting in neuronal cell death due to excessive accumulation of intracellular calcium. Kang TC. Et al., J. Neurocytol . , 30 , pp945-955, 2001) and the increase of radicals in vivo due to sudden oxygen supply during ischemia-reperfusion damages DNA and cytoplasm, causing neuronal death. Oxidative neuronal suicide mechanism (Won MH. Et al., Brain Res. , 836, pp70-78, 1999). Based on these mechanisms, many researches have been conducted to search for substances that effectively inhibit neuronal cell death caused by cerebral ischemia or to reveal the mechanism of the substance. However, few substances have been found to effectively inhibit neuronal cell suicide.

However, in the case of minocycline, a tetracycline antibiotic that inhibits apoptosis under ischemic conditions, myocardial infarction (Yrjanheikki J. et al., Proc. Natl. Acad. Sci. USA , 96 (23) , pp13496-13500, 1999 .) And ischemic diseases such as cerebral infarction (Scarabelli TM et al., J. Am. Coll. Cardiol. , 43 (5) , pp865-874, 2004.) as well as Parkinson's disease (Wu DC et al., J. Neurosci ., 22 (5), pp1763-1771 , 2002), Alzheimer's disease, peak (Pick) disease, Creutzfeldt -... Jakob (Creutzfeldt-Jakob) disease and Huntington's disease (Chen M. et al, Nat Med, 6 (7) , pp797-801, 2000), and is known to be effective in the treatment of degenerative brain diseases caused by neuronal cell suicide. In addition, the present inventors have confirmed that such tetracycline-based antibiotics improve cell survival under ischemic conditions similar to this study (US Pat. No. 6716822). Therefore, natural products that improve cell survival by inhibiting apoptosis in ischemic conditions can be inferred to be effective in treating not only ischemic diseases but also degenerative brain diseases caused by neuronal suicide.

Snow riding horse ( Aruncus dioicus ( Walt. ) Fern var . kamtschaticus Hara or Korean goatsbeard) is a perennial herb that belongs to the rosacea rosaceae and is also called wild beast or wild horse. In oriental medicine and private sector, outposts are used as drugs for detoxification and hemostasis. The main components of the horseback riding with this effect are saponins, salicylic aldehydes, sparin, ash, lipids, vitamin A and the like.

There are Japanese Patent Nos. 3251188 and 5017768 using patents using components extracted from this plant. However, these patents use the extract as an antibiotic and have not been taught or described anywhere in the literature regarding uses related to ischemic and degenerative brain diseases. .

Therefore, the present inventors have recognized the problems of the prior art in the treatment of ischemic disease and degenerative brain disease, and through research and development to develop apoptosis inhibitors for improving cell viability in hypoxic conditions as a therapeutic agent for ischemic disease and degenerative brain disease. The present invention has been completed by confirming the effect of the eye horse riding extract and its active fraction.

It is an object of the present invention to provide an eyelid horse riding extract and composition containing the same that effectively inhibits apoptosis in ischemic conditions.

In order to carry out the above object, the present invention provides a pharmaceutical composition for the prevention and treatment of ischemic diseases containing the eyelid riding horse extract as an active ingredient.

The ischemic diseases include myocardial infarction, cerebral infarction, ischemic impaired nephropathy, diabetic foot ulcers and diseases caused by surgical side effects (heart failure, incomparable nephropathy, etc.).

The present invention provides a pharmaceutical composition for the prevention and treatment of degenerative brain diseases containing the eyelid riding horse extract as an active ingredient.

The degenerative brain diseases include Parkinson's disease, Huntington's disease, Alzheimer's disease, Pick's disease, and Creutzfeldt-Jakob's disease.

The extract is a crude extract soluble in a polar solvent of C ₁ to C ₄ lower alcohols such as water, ethanol and methanol, and a non-polar solvent soluble extract soluble in an organic solvent such as a non-polar solvent of hexane, ethyl acetate and dichloromethane. Preferably, crude extracts soluble in water or methanol and nonpolar solvent soluble extracts soluble in ethyl acetate are included.

Hereinafter, the present invention will be described in detail.

The crude horseback riding extract of the present invention, after drying, has a volume of about 1 to 15 times the weight of the fine eyelid stem or leaf, preferably about 5 to 10 times the amount of C ₁ to C ₄ such as water, ethanol, methanol, and the like. of Lower alcohol or a mixed solvent having a mixing ratio of about 1: 0.1 to 1:10, preferably methanol, is added at an extraction temperature of 20 to 100 ° C, preferably 50 to 100 ° C, for about 0.5 to 2 days, preferably Cold sediment extraction, hot water extraction, ultrasonic extraction, reflux cooling extraction, herbal bath extraction, etc. for 1 hour to 1 day is an extraction method, preferably extracted by repeating 1 to 12 times, preferably 3 to 4 times with a water bath extraction method. After filtration, the filtered extract was concentrated under reduced pressure at 20 to 100 ° C., preferably at 40 to 70 ° C. with a vacuum rotary concentrator, and the extracted residue was dried with a vacuum freeze dryer to be soluble in water, lower alcohol or a mixed solvent thereof. A one-eye horse riding extract can be obtained.

In addition, the non-polar solvent soluble extract of the present invention is about 1 to 10 times, preferably about 1 to 10 times of these suspensions after suspending the water-soluble extract, the lower alcohol soluble extract, and the soluble extract by the mixed solvent thereof in water. A non-polar solvent such as hexane, ethyl acetate, chloroform, dichloromethane, and the like, can be obtained by fractionation of 1 to 5 times, preferably 2 to 4 times, by addition of ethyl acetate. In addition, conventional fractionation processes well known in the art may also be performed (Harborne JB, Phytochemical methods: A guide to modern techniques of plant analysis. 3rd Ed ., Pp 6-7, 1998).

The present invention provides a pharmaceutical composition effective for the prevention and treatment of ischemic diseases and degenerative brain diseases, containing the crude horse riding horse extract or non-polar solvent soluble extract of the present invention obtained as the active ingredient.

Pharmaceutical compositions comprising the extract of the present invention may further comprise suitable carriers, excipients and diluents commonly used in the manufacture of pharmaceutical compositions.

Pharmaceutical compositions comprising extracts according to the invention, in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like, oral preparations, suppositories and sterile injectable solutions, respectively, according to conventional methods. Can be formulated and used. Carriers, excipients and diluents that may be included in the composition comprising the extract include 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. When formulated, diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, and surfactants are usually used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules and the like, and such solid preparations may include at least one excipient such as starch, calcium carbonate and sucrose in the extract. Or lactose, gelatin and the like. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid preparations for oral use may include various excipients, such as wetting agents, sweeteners, fragrances, preservatives, etc., in addition to water and liquid paraffin, which are commonly used to include suspensions, solutions, emulsions, and syrups. have. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, suppositories. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate and the like can be used. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

The compositions of the present invention can be administered via several routes including oral, transdermal, subcutaneous, intravenous or intramuscular.

Preferred dosages of the extracts of the present invention vary depending on the condition and weight of the patient, the extent of the disease, the form of the drug, the route of administration and the duration, and may be appropriately selected by those skilled in the art. Administration may be administered once a day or may be divided several times.

The composition comprising the extract of the present invention provides a dietary supplement for the prevention and treatment of ischemic diseases and degenerative brain diseases. Examples of the food to which the extract can be added include various foods, beverages, gums, teas, vitamin complexes, and health supplements.

In addition, the extract of the present invention may be added to food or beverages for the purpose of preventing the effects of ischemic diseases and degenerative brain diseases. At this time, the amount of the extract in the food or beverage may be added in 0.01 to 15% by weight of the total food weight, the health beverage composition may be added in a ratio of 0.02 to 5 g, preferably 0.3 to 1g based on 100 ml. have.

Health functional food of the present invention includes the form of tablets, capsules, pills, liquids and the like.

The health functional beverage composition of the present invention is not particularly limited to other ingredients except for having the extract as an essential ingredient in the indicated ratio, and may contain various flavors or natural carbohydrates, etc. as additional ingredients, as in general beverages. Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And conventional sugars such as polysaccharides such as dextrin, cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those described above, natural flavoring agents (tauumatin, stevia extract (e.g., Rebaudioside A, glycyrrhizin, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. The proportion of said natural carbohydrate is generally about 1-20 g, preferably about 5-12 g per 100 ml of the composition of the present invention.

In addition to the above, the composition of the present invention includes various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, coloring and neutralizing agents (such as cheese and chocolate), pectic acid and salts thereof, alginic acid and its Salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated beverages, and the like. The compositions of the present invention may also contain pulp for the production of natural fruit juices and fruit juice beverages and vegetable beverages. These components can be used independently or in combination. The proportion of such additives is not so critical but is generally selected from the range of 0 to about 20 parts by weight per 100 parts by weight of the composition of the present invention.

The present invention will be described in more detail based on the following examples and experimental examples, but the present invention is not limited thereto.

Example 1 Preparation of Crude Horse Riding

1-1. Preparation of Snow Horse Riding Water Extract

After washing and washing with snow-capped horses and roots collected from Hwacheon-gun, Gangwon-do, dried, and adding 2 liters of water to 200 g of snow-capped horses according to the conventional extractor extraction method. 2 times), and then filtered. 2.5 l of the extract obtained by filtration was lyophilized to obtain 16 g of a horseback riding extract.

1-2. Preparation of Snow Horse Riding Methanol Extract

5 g of methanol was added to 500 g of the collected eye horse riding sample, extracted three times in an ultrasonic extractor for 24 hours, filtered, and concentrated under reduced pressure with a reduced pressure concentrator (NP-1, eyella). Obtained.

Example 2 Preparation of Snow Horse Riding Nonpolar Solvent Soluble Extracts

In the case of the water extract of Example 1-1 as it is, in the case of the methanol extract of Example 1-2, 30 g of the concentrated extract was suspended in 1 L of distilled water, and 1 L of hexane was added thereto, followed by strong shaking. The supernatant was recovered when the two layers were completely separated. 1 L of hexane was added to the remaining water layer, and the same process was repeated two more times to obtain 3 L. The resulting hexane layer was concentrated under reduced pressure to give 2 g of hexane fraction. The same amount of ethyl acetate was added to the remaining water layer after hexane extraction, and the above process was repeated three times, and then concentrated under reduced pressure to prepare 10 g of ethyl acetate fractions. In the same manner, 10 g butanol fractions were prepared. Finally, the remaining water layer was concentrated as it is.

Reference Example 1. Preparation of Laboratory Animals

The experimental animals were Sprague-Dawley male rats (Hyochang Science Co., Ltd.) weighing 280-310g, and were subjected to constant conditions (temperature: 21 ± 2, contrast: 12 hours) in Daegu Catholic University animal breeding room. Cycle), free feeding of food and water is allowed, and sufficient water and food are provided until the start of the experiment, and the experimental animals are pretreated for 10 minutes before the start of the experiment.

Experimental Example 1. Measurement of cell viability improvement ability of crude extract (in vitro)

In order to investigate the effect of the extract on the survival of cells under hypoxic and normal oxygen conditions, after adding a medium containing extracts of different concentrations to the cells, the cell survival by performing the experiment as follows under hypoxic and normal oxygen conditions Trypan blue staining exclusion method (trypan) described in Sambrook J. and Russel DW, Molecular Cloning 3rd ed., Vol. 3, A8.6-8,8, Cold Spring Harbor Laboratory Press, 2001 blue dye exclusion).

HepG2 (1 × 10 ⁶ cells / 4 ml), a liver cancer cell line, was incubated in a 60 mm dish using Eagle minimal media. The cells were incubated for 48 hours at 37 ° C. and 5% CO ₂ -95% atmosphere, and then replaced with new culture medium, and then the crude extract was dissolved in DMSO and added to the culture medium at a concentration of 100 μg / ml, and Negative control with nothing added was incubated for 2 days at low oxygen (3% oxygen) and normal oxygen conditions, and then cell number was measured using trypan blue dye exclusion. It was.

As shown in Figure 1, the extract was confirmed to improve the cell survival at a concentration of 100㎍ / ㎖ under hypoxic conditions (Fig. 1 above), it did not inhibit the growth of cells under normal oxygen conditions (Fig. Below picture 1). Accordingly, it can be confirmed that the extract effectively improves cell survival under hypoxic conditions and should be administered at a concentration of 100 µg / ml or more in order to have an effect under hypoxic conditions without inhibiting cell growth under normal conditions. there was.

Experimental Example 2 Test of apoptosis inhibitory effect in hypoxic condition of crude extract (in vitro)

In order to confirm the mechanism by which the extract helps cell survival under hypoxic conditions, the DNA fragmentation assay (DNA fragmentation assay-GP Studzinski, Apoptosis , pp47-48, 1999) described in the literature was modified.

HepG2 (1 × 10 ⁶ cells / 4 ml), a liver cancer cell line, was incubated in a 60 mm dish using Eagle minimal media. Cells were incubated for 48 hours at 37 ° C. and 5% CO ₂ -95% atmosphere under the same conditions as in Experimental Example 1, and then replaced with new culture medium. Then, the extract of Example 1 was dissolved in DMSO 100 The DNA ladder appears by performing DNA fragmentation assay while culturing the experimental group added to the culture medium at the concentration of ㎍ / ㎖ and the negative control group (no negative control) for 2 days at low oxygen conditions. Was investigated. As a result, the time when the DNA ladder appeared in the experimental group to which the extract was added was delayed (see FIGS. 2A and 2B). Therefore, it was confirmed that the crude extract has an effect of suppressing apoptosis and improving cell survival.

Experimental Example 3. Measurement of cell viability improvement of fractions (in vitro)

To confirm the activity of the fractions isolated from this crude extract, the MTT assay described in the literature (Julio E. Celis (Ed.), Cell Biology , 1, pp369-370, Academic Press, 1994) was modified to HepG2 cell line. After the addition of the medium containing the fractions of different concentrations, the experiment was performed in the hypoxic condition as follows to investigate the degree of improving cell survival.

HepG2 (2 × 10 ⁵ cells / 800 μl), a human hepatoma cell line, was incubated in a 12 well-plate using Eagle Minimal Media. After incubating the cells for 48 hours at 37 ° C. and 5% CO ₂ -95% atmosphere, the crude extract of Example 1 as well as the fractions of Example 2 were dissolved in DMSO to a concentration of 1-1000 μg / ml. The MTT assay was performed while the experimental group added to the culture medium and the negative control group (negative control) to which nothing was added were incubated under hypoxic and normal oxygen conditions for 2 days, respectively.

As a result of the experiment, as shown in FIG. 3, the hexane fractionation layer (Hx) hardly contributed to the increase of cell viability, and the crude extract (Wh) and butanol fractionation layer (Bu) at a concentration of 100-1,000 µg / ml. The survival rate of the cells can be increased, whereas the ethyl acetate fractionation layer (EA) has been shown to increase the survival rate of the cells at a concentration of 10-100 µg / ml. Therefore, it was confirmed that a lot of substances to improve cell survival exist in the ethyl acetate fractionation layer.

Experimental Example 4. Effect test on cerebral infarction during intraperitoneal injection (in vivo)

In order to confirm whether the extract is effective in the treatment of cerebral infarction, the experiment described in the literature (Han HS et al., Journal of Neuroscience , 22 , pp3921-3928, 2002) was modified to use the rat infarction model of Reference Example 1 above. Animal experiments were performed as follows.

4-1. Effect of Crude Extracts on Cerebral Infarction Model

After inhalation anesthesia of the rat of Reference Example 1 with enflurane, the blood pressure measurement and blood sampling route were secured in the lower limb artery, and the cervical incision was exposed to expose the carotid artery. A cerebral infarction model was created by inserting 3-0 nylon sutures to block the middle cerebral artery.

The crude extract of Example 1 was injected into the cerebral infarction model to investigate the area of necrotic sites. At this time, the dose should be solved at the same time that it is a measurement of the safe nontoxic dose and the determination of the effective dose. Therefore, based on the result of the in vitro experiment, the method is to raise the step by step to the minimum possible starting point. Was used. To this end, the dose increase was adopted by the fold increase method, and starting at 50 mg / kg increased to 100 and 200 mg / kg.

20 hours and 1 hour before blocking the middle cerebral artery, 1 ml of crude extract of 200 mg / kg was injected into the abdominal cavity, respectively, and left for 2 hours to remove the suture and the animals were recovered. After 22 hours, the animals were euthanized, brain tissues were extracted, stained in TTC solution, and the damage was measured using an image analysis system to calculate the ratio of damage to the brain hemispheres. Ischemic index was measured to compare the effects of drugs.

Ischemia index (%) = A / B × 100

A: the volume of the injury site in the hemisphere of the brain (mm ³ ),

B: total volume of brain hemispheres (mm ³ ).

As a result of the experiment, rats not injected with crude extracts showed an ischemic index of 93% compared to rats injected with 78%, and the volume of necrotic area was reduced by 15% (p <0.05). It was confirmed that there is a significant effect in this cerebral infarction model (see Figs. 4a, 4b, 4c).

4-2. Effect of Ethyl Acetate Fraction on Cerebral Infarction Model

The ethyl acetate fraction layer, which was found to contain a large amount of active substance in Experimental Example 3, was injected into a cerebral infarction model made by the same method as Experimental Example 4-1, and the area of necrotic sites was examined.

1 ml of 10 mg / kg ethyl acetate fractionation layer was injected into the abdominal cavity 20 hours and 1 hour before blocking the middle cerebral artery and causing ischemia, and left perfusion for 1 hour and 30 minutes. After 3 hours, TTC staining was performed, and an ischemic index was measured in the same manner as in Experimental Example 4-1.

As a result of the experiment, rats not injected with the ethyl acetate fractionation layer had an ischemic index of 55%, compared with only 24% of the rats injected. The necrotic site volume was reduced by about 57% (p <0.05), and it was confirmed that the eyelid horse riding ethyl acetate fraction had an excellent effect in the cerebral infarction model (see FIG. 5).

Experimental Example 5. Toxicity Test

The crude extract of Example 1 was injected into 320 ± 20 g of Sprague-Dawley rats to observe changes.

The body weight was measured 24 hours after the injection of 500 mg / kg of eyelid horseshoe crude extract into the abdominal cavity. There was no significant weight change. There was also no change in behavioral behavior and no revenge when incised.

It describes an example of the formulation of the pharmaceutical composition containing the eyelid riding horse extract of the present invention, the present invention is not intended to limit this but only to explain in detail.

Formulation Example 1 Preparation of Powder

300 mg of the eyelid horse riding non-polar solvent soluble extract of Example 1

Lactose 100 mg

Talc 10 mg

The above ingredients are mixed and filled in an airtight cloth to prepare a powder.

Formulation Example 2 Preparation of Tablet

50 mg of eyelid horse riding non-polar solvent soluble extract of Example 1

Corn starch 100 mg

Lactose 100 mg

2 mg magnesium stearate

After mixing the above components, tablets are prepared by tableting according to a conventional method for preparing tablets.

Formulation Example 3 Preparation of Capsule

50 mg of horseshoe-dried crude extract of Example 1

Corn starch 100 mg

Lactose 100 mg

2 mg magnesium stearate

According to a conventional capsule preparation method, the above ingredients are mixed and filled into gelatin capsules to prepare capsules.

Formulation Example 4 Preparation of Injection

50 mg of horseshoe-dried crude extract of Example 2

Appropriate sterile distilled water for injection

pH adjuster

According to the conventional method for preparing an injection, the amount of the above ingredient is prepared per ampoule (2 ml).

Formulation Example 5 Preparation of Liquid

100 mg crude horseback riding extract of Example 2

10 g of isomerized sugar

5 g of mannitol

Purified water

According to the conventional method of preparing a liquid solution, each component is added and dissolved in purified water, lemon flavor is added to the mixture, and then the above ingredients are mixed, purified water is added to adjust the total amount to 100 ml, and then filled in a brown bottle. The solution is prepared by sterilization.

Formulation Example 6 Preparation of Healthy Food

Crude Horse Equine Extract of Example 1

Vitamin mixture proper amount

70 μg of Vitamin A Acetate

Vitamin E 1.0 mg

Vitamin B ₁ 0.13 mg

Vitamin B ₂ 0.15 mg

Vitamin B ₆ 0.5 mg

0.2 μg of vitamin B ₁₂

Vitamin C 10 mg

10 μg biotin

Nicotinic Acid 1.7 mg

Folate 50 ㎍

Calcium Pantothenate 0.5mg

Mineral mixture

Ferrous Sulfate 1.75 mg

Zinc Oxide 0.82 mg

Magnesium carbonate 25.3 mg

Potassium phosphate monobasic 55 mg

Potassium Citrate 90 mg

Calcium Carbonate 100 mg

Magnesium chloride 24.8 mg

Although the composition ratio of the vitamin and mineral mixture is a composition suitable for a relatively healthy food in a preferred embodiment, the composition ratio may be arbitrarily modified, and the above ingredients are mixed according to a conventional health food manufacturing method. The granules may be prepared and used for preparing a health food composition according to a conventional method.

Formulation Example 7 Preparation of Healthy Drink

Crude Horse Equine Extract of Example 1 1000 mg

Citric acid 1000 mg

100 g oligosaccharides

Plum concentrate 2 g

1 g of taurine

Add 900 ml of purified water

After mixing the above components in accordance with a conventional healthy beverage production method, and stirred and heated at 85 ℃ for about 1 hour, the resulting solution is filtered and obtained in a sterilized 2 L container, sealed sterilization and then refrigerated and stored in the present invention For the preparation of healthy beverage compositions.

Although the composition ratio is mixed with a component suitable for a favorite beverage in a preferred embodiment, the composition ratio may be arbitrarily modified according to regional and ethnic preferences such as demand hierarchy, demand country, and usage.

The composition containing the eyelid equestrian extract of the present invention inhibits apoptosis in the ischemic state, not only reduces the infarct of the tissue, but also has no side effects even for long-term use, so the medicine and health functional food for the treatment of ischemic disease and degenerative brain disease. Can be used for

Claims (8)

A pharmaceutical composition for the prevention and treatment of cerebral infarction and myocardial infarction, which contains an eyelid horseback extract as an active ingredient. delete A pharmaceutical composition for the prevention and treatment of degenerative brain diseases, which contains an eyelid horseback extract as an active ingredient. The pharmaceutical composition according to claim 3, wherein the degenerative brain disease is Parkinson's disease, Huntington's disease, Alzheimer's disease, Peak disease and Creutzfeldt-Jakob disease. 4. The pharmaceutical composition according to claim 1 or 3, wherein the extract is a crude extract or ethyl acetate soluble extract extracted with water, lower alcohol or a mixed solvent thereof. A dietary supplement for the prevention and improvement of cerebral infarction and myocardial infarction, including eyelid horseback extract and food acceptable additives. The dietary supplement for preventing and improving cerebral infarction and myocardial infarction according to claim 6, which is a tablet, a pill, a capsule or a liquid. The health functional food for preventing and improving cerebral infarction and myocardial infarction according to claim 6, wherein the extract is a crude extract or ethyl acetate soluble extract extracted with water, lower alcohol or a mixed solvent thereof.

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