KR20150001107A - A composition for improving memory and preventing or treating cognitive dysfunction comprising epigallocatechin-3-gallate and luteolin - Google Patents

Abstract

The present invention relates to a composition for promoting memory and preventing or treating diseases related to cognitive dysfunction including epigallocatechin-3-gallate and luteolin as active ingredients. More specifically, the mixture of epigallocatechin-3-gallate and luteolin of the present invention exhibits enhancement of memory in Morris water-maze test and passive avoidance test using a mouse in which memory loss is induced by beta-amyloid (A beta_1-42) which is one of factors causing dementia, thereby being used as pharmaceutical compositions and health food for preventing and treating diseases related to cognitive dysfunction and memory.

Description

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a composition for enhancing memory and cognitive dysfunction which comprises epigallocatechin gallate and luteolin as an active ingredient, and a composition for preventing or treating cognitive dysfunction, comprising epigallocatechin-3-gallate and luteolin,

The present invention relates to a composition for preventing or treating memory-related and cognitive dysfunction-related diseases containing epigallocatechin-3-gallate (EGCG) and luteolin (LUT) as active ingredients.

Dementia is a complex clinical syndrome characterized by a marked impairment in human cognitive function, intellectual ability, emotional and behavioral changes. It causes cortical dysfunction such as memory, attention, language function, and spatio-temporal ability to lead daily life and social life It is in a state of great difficulty. The term "dementia" refers to a condition in which one or more of the other cognitive functions, including memory, is present.

Alzheimer's Disease or Parkinson's Disease is the cause of dementia, which can be caused by cerebral hemorrhage, metabolic diseases such as hepatic encephalopathy, Wilson's Disease, infectious diseases such as neurosyphilis, acquired immunodeficiency, Such as drug addiction, brain trauma, etc. In some forms, diseases that may cause structural and functional abnormalities in the central nervous system may cause dementia. Among them, dementia caused by Alzheimer's disease is the most common with 50 ~ 60%, followed by dementia caused by cerebrovascular disease.

It is estimated that the number of dementia patients among the elderly people aged 65 or older is about 460,000 in Korea, and that it will reach 700,000 in 2020 and 2 million in 2040 (Seoul National University Hospital). p. 132, 2008). Because dementia requires long-term treatment and the nature of its symptoms, the degree of life inconvenience is worse than other severe diseases such as cancer, and the physical, psychological, and economic burden of patient care and protection is also great. With the rapid rate of increase in the elderly population and the prevalence of dementia, it is clear that dementia is a task that requires social and national solutions.

Memory cognitive impairment is the first and most common symptom of Alzheimer's disease. In the early stages of Alzheimer's disease, patients suffer from recent memory disorders that do not remember the details of recent conversations or work, due to the impaired ability of hippocampal neurons to damage recent memory . But at this time, the remote long-term memory of events in the distant past is relatively well preserved. However, as the disease progresses, memory of the past gradually becomes disturbed as the cerebral cortex associated with the storage of long - term memory is damaged.

These symptoms of Alzheimer's disease are related not only to cytotoxicity by β-amyloid deposition but also to synaptic dysfunction of the cholinergic system (PM et al., Interactions between the amyloid and cholinergic mechanisms in Alzheimer's disease. Neurochem Int., 53: pp. 103-111, 2008). Cholinergic dysfunction is known to contribute to memory and cognitive dysfunction in Alzheimer's disease patients. Meynert's basal nucleus of Meynert Cholinergic neurons are associated with memory and cognitive function in association with temporal lobe, hippocampus and amygdala. In brain of Alzheimer's disease, 78% in temporal lobe, 60% in hippocampus, , It is known that neurons decrease by 67%. When brain cells are damaged by cytotoxicity, there is a barrier to the transmission of certain information, ie, the metabolism of neurotransmitters, which causes memory impairment. Many researchers have been steadily reporting the selective reduction of acetylcholine and its associated enzyme (choline acetyltransferase) in Alzheimer's disease. Furthermore, in the brains of patients with Alzheimer's disease, the nicotinic acetylcholine receptor, the muscarinic acetylcholine receptor, as well as the cholinergic reabsorption and acetylcholine secretion The function is also degraded.

Several cholinergic drugs have been developed to reverse these cholinergic system disorders, but the most effective and most widely used cholinesterase inhibitors are acetylcholinesterase inhibitors (AchEI). Currently marketed under FDA approval are donepezil, rivastigmine, and galanthamine, which inhibit the degradation of acetylcholine in synaptic clefts and increase the concentration, . These drugs show improvement in cognitive function and daily life in the early stages but they do not fundamentally prevent disease progression in that they return to the pre-administration state after 9 months to 1 year. The effect can be seen only if it is used relatively early, and it is not effective in the case of severe dementia. Common side effects include nausea, diarrhea, loss of appetite, dizziness, muscle cramps and sleep disturbances due to increased acetylcholine, and serious side effects include syncope. In addition, memantine (NMDA receptor blocker), ginkgo biloba extract (ginko biloba), huperzine A (reversible AChEI) extracted from Chinese medicine, Various therapeutic agents have been developed in connection with the pathology of Alzheimer's disease, including protein, beta amyloid protein formation, inhibition of deposition, antioxidant, and anti-inflammatory effects.

At present, in the case of Alzheimer's disease treatment in Korea, foreign affiliated companies are highly dependent on imports so that they occupy a share of 98%. In addition, one company accounts for 80% of the market, which is virtually monopolistic (Korea Institute of Science and Technology Information, dementia treatment p.116, 2002). This shows that the growth of dementia treatment sector is urgent in terms of national competitiveness. Therefore, the development of effective anti-cancer drugs and memory-improving agents with fewer side effects can significantly improve the quality of life of Alzheimer's disease patients and improve and cure early-onset disease, as well as for amenorrhea and dementia patients suffering from memory cognitive impairment. It is thought that it can give.

On the other hand, Korean Patent Application Nos. 10-1148623 and 10-1141355 disclose compositions for treating and preventing memory impairment and cognitive dysfunction containing alpha-tunon or alpha-pinene as an active ingredient, There is no mention of the epigallocatechin-3-gallate and luteolin of the invention.

Therefore, the inventors of the present invention conducted a systematic review of the treatment and prevention of epigallocatechin-3-gallate and luteolin memory and cognitive dysfunction, and found that β-amyloid, Aβ _{1- 42} ) were used to conduct a water-maze test and a passive avoidance test to measure memory performance using an experimental animal. As a result, it has been found that the improved activity of a combination composition containing both epigallocatechin-3-gallate and luteolin leads to excellent memory enhancement and cognitive dysfunction for beta-amyloid-induced memory impairment The present invention has been completed.

It is an object of the present invention to provide a pharmaceutical composition containing epigallocatechin-3-gallate (EGCG) and luteolin (LUT) as active ingredients for the prevention or treatment of memory- To provide.

Another object of the present invention is to provide a health functional food containing epigallocatechin-3-gallate and luteolin as an active ingredient for preventing or ameliorating memory-related and cognitive dysfunction-related diseases To provide.

In order to achieve the above object, the present invention provides a pharmaceutical composition comprising epigallocatechin-3-gallate (EGCG) represented by the following general formula (1) and luteolin (LUT) A pharmaceutical composition for preventing or treating memory-related and cognitive dysfunction-related diseases is provided.

≪ Formula 1 >

(2)

Specifically, the cognitive dysfunction-related diseases include, but are not limited to, forgetfulness, Alzheimer's disease, cerebrovascular dementia, dementia caused by head injury, or Parkinson's disease.

Epigallocatechin-3-gallate (EGCG) used as an active ingredient in the present invention is a kind of polyphenol present in green tea leaves, and luteolin (LUT) is a kind of polyvinylpyrrolidone such as rooibos leaf, celery, Which is a kind of flavonoid which is widely found in green vegetables. Both of these substances are emerging as a neuroprotective substance having antioxidant and anti-inflammatory action.

The epigallocatechin-3-gallate and luteolin according to the present invention are preferably contained in an amount of 0.1 to 50% by weight based on the total weight of the composition. However, the composition is not limited thereto, and may vary depending on the condition of the patient, the type of disease, and the progress of the disease.

In addition, the composition of the present invention may be provided in various forms selected from a pharmaceutical composition or a health food composition.

When the composition of the present invention is a pharmaceutical composition, the method of administration is determined depending on the severity of symptoms. The extract of the present invention can be administered to mammals such as rats, mice, livestock, and humans in various routes. All modes of administration may be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intra-uterine or intracerebroventricular injections. The dosage of the active ingredient in the pharmaceutical composition may vary depending on the route of administration, the severity of the disease, the age, sex, and weight of the patient, and may be administered once to several times per day. However, in order to obtain the desired effect, it is preferable to administer it at 0.01 mg / kg to 10 g / kg, preferably 1 mg / kg to 1 g / kg per day. The administration may be carried out once a day or divided into several doses. Thus, the dosage amounts are not intended to limit the scope of the invention in any manner.

The epigallocatechin-3-gallate and luteolin according to the present invention can be used in the form of a general pharmaceutical preparation, and can be administered in various formulations, oral or parenteral, have. In addition, in the case of formulation, in addition to the active ingredients epigallocatechin-3-gallate and luteolin, one or more additional pharmaceutically acceptable carriers may be prepared. The pharmaceutically acceptable carrier may be a mixture of saline, sterilized water, Ringer's solution, buffered saline, dextrose solution, maltodextrin solution, glycerol, ethanol and one or more of these components. If necessary, an antioxidant, , And other conventional additives such as a bacteriostatic agent may be added.

Solid formulations for oral administration may include tablets, pills, powders, granules, capsules and the like, which may contain at least one of epigallocatechin-3-gallate and luteolin The above excipients such as starch, calcium carbonate, sucrose or lactose, gelatin and the like are mixed. In addition to simple excipients, lubricants such as magnesium stearate talc are also used. The liquid preparations for oral administration include suspensions, solutions, emulsions and syrups. In addition to water and liquid paraffin which are commonly used diluents, various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like May be included.

Further, the pharmaceutical composition of the present invention can be administered parenterally, and parenteral administration is by subcutaneous injection, intravenous injection, or intramuscular injection. In order to formulate parenteral administration formulations, epigallocatechin-3-gallate and luteolin are mixed with water and a stabilizer or buffer to prepare a solution or suspension, which is then mixed with an ampoule or vial It is formulated in unit dosage form.

In addition, when the composition of the present invention is a health food composition, there is no particular limitation on the type of food to which the compound of the present invention can be added, but various foods such as beverage, gum, tea, , Health supplement foods, etc., and can be used in the form of pills, powders, granules, infusions, tablets, capsules or beverages.

The health beverage composition of the present invention is not particularly limited to liquid ingredients other than the above-mentioned compounds as essential ingredients in the indicated ratios, and may contain various flavors or natural carbohydrates as additional ingredients such as ordinary 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; Polysaccharides such as dextrin, cyclodextrins; And sugar alcohols such as xylitol, sorbitol, and erythritol. As natural flavors other than those described above, natural flavors (such as tau martin, stevia extract (e.g., rebaudioside A, glycyrrhizin)) and synthetic flavors (saccharin, aspartame, etc.) have. The ratio 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-mentioned composition, the composition of the present invention can be used as a flavoring agent such as various nutrients, vitamins, minerals (electrolytes), synthetic flavors and natural flavors, coloring agents and intermediates (cheese, chocolate etc.), pectic acid and its salts, Salts, organic acids, protective colloid thickening agents, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonating agents used in carbonated beverages and the like. In addition, the compositions of the present invention may contain flesh for the production of natural fruit juices and fruit juice drinks and vegetable drinks. These components may be used independently or in combination. The proportion of such additives is not so critical, but is generally selected in 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 relates to a composition for preventing or treating memory-related and cognitive dysfunction-related diseases, which comprises epigallocatechin-3-gallate and luteolin as an active ingredient, Morris water-maze test and passive avoidance test in animal models where epigallocatechin-3-gallate and luteolin induced memory impairment by beta-amyloid It is expected that the compound can be usefully used as a pharmaceutical composition and health functional food useful for prevention and treatment of amnesia and memory disorder related diseases.

Figure 1 shows the effect of epigallocatechin-3-gallate (EGCG) and luteolin (LUT) on SD rats in which memory impairment was induced by beta-amyloid (A? _1-42 ) Morris water-maze test is a result of the combination administration, which is a remarkable memory enhancing effect.
2 is a graph showing the effect of epigallocatechin-3-gallate (EGCG) and luteolin (LUT) on SD rats in which memory impairment was induced by β-amyloid (Aβ _1-42 ) The results of the passive avoidance test show that the combined administration is a remarkable memory enhancing effect.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the present invention is not limited by these examples.

Example  1. Beta-amyloid-induced inhibition of memory damage

1. Epigallocatechin-3-gallate (EGCG) and luteolin (LUT)

Epigallocatechin-3-gallate and luteolin represented by Chemical Formulas 1 and 2 were obtained from Sigma-Aldrich (St. Louis, USA) and compared with the physical properties disclosed in the literature, And dissolved in corn oil at a dose of 5 mg / kg and 15 mg / kg, respectively.

≪ Formula 1 >

(2)

2. Experimental animals

Male SD (Sprague-Dawley) rats, 7 weeks old, were used as experimental animals for one week. The test group and the control group were divided into five groups, respectively. Epigallocatechin-3-gallate and luteolin in concentrations of 5 mg / kg and 15 mg / kg were dissolved in corn oil and administered intraperitoneally during the experimental period (14 days after surgery). The animals were kept at 21-26 ° C and 40-60% humidity in the animal breeding room.

3. Morris water-maze test

In order to confirm that the epigallocatechin-3-gallate and luteolin obtained from the above exhibit memory enhancing effect in an animal model in which memory amelioration is induced by beta amyloid, water-maze test).

A beta-amyloid (Aβ _1-42 ) was infused at 600 pmoles / day (1 μl / min rate, 5 μl total volume) for 14 days in the lateral ventricle of 7-week-old male SD rats, (5 mg / kg and 15 mg / kg) of epigallocatechin gallate and luteolin (15 mg / kg) for the same period of time were measured by the Morris water-maze test Respectively.

Experiments were carried out to investigate whether epigallocatechin gallate and luteolin complexes contribute to the spatial perception of experimental animals and to the recovery of short- and long-term memory (short and long-term memory recovery). Experimental equipment consists of a circular tank (180 cm in diameter, 60 cm in height) and a platform (12 cm in diameter, 38 cm in height). The height of the water in the water tank (temperature 22 ± 2 ° C) was up to 2 cm above the platform so that when the mouse sat on the escape platform, the body could come out of the water. In the experiment with water, the test animals were kept at the same level during the experiment period so that the environment was not changed because the animals were looking for the escape pods using the markers around the water tanks. The time taken for the animals to visit when they visited the escape pod for more than 20 seconds was defined as the escape latency, and the mean value obtained by performing this three times a day was the mean escape latency. The incubation latency was recorded by observing the computer program (Ethovision 3.1, Noduls, The Netherlands) by installing a camera on the ceiling above the tank. The experiment was carried out three times daily for 4 days. In this case, the possibility of visiting the esophagus by chance was minimized by sequentially changing the position of placing the experimental animals in the water tank every time. If the experimental animal can not find the escape pod within 90 seconds, the escape latency is set to 90 seconds, and if the experimental animal is sitting on the escape pod, it is placed for 20 seconds to remind the surrounding cues.

Experimental results also go beta amyloid and Teller tekin go EPI as shown in Fig. 1 Rate 5 mg / kg, Luteolin _{5 mg / kg (Aβ 1-42 +} EGCG + LUT) administered at the same time a group is beta amyloid (Aβ ₁ mean as compared to _-42), only the singly administered control group, the experimental group administered alone to beta-amyloid and a single material with a higher dose (15 mg / kg) (Aβ 1-42 + EGCG and Aβ _1-42 + LUT) The latency to escape was significantly shortened from the 2nd to the 4th day and decreased to a level similar to that of the normal group (NOR). The fourth day, the control group (Aβ _1-42), as shown in FIG. 1 75.71 ± 1.49 cho, Aβ _1-42 + LUT is 59.07 ± 3.01 cho, Aβ _1-42 + EGCG was compared with 58.47 ± 3.03 cho, Aβ _{1 -42} + EGCG + LUT was 28.13 ± 1.79 sec, which was statistically significant in comparison with the single dose group. (Figure 1, * p < 0.05 and ** p < 0.01: Aβ _1-42 + EGCG + LUT vs. other groups).

4. Passive avoidance test

In order to confirm the effects of epigallocatechin gallate and luteolin on memory performance, the following experiment was performed by applying the passive avoidance experiment.

Experiments were carried out in a manual evasion experiment in a device in which bright compartments and dark compartments of the same structure (26 cm by 28 cm, 15 cm by height) were connected by guillotine doors. The light compartment is equipped with a light bulb and the dark compartment is provided with a 2 mm thick stainless steel rods at 1 cm intervals on the floor to provide electrical stimulation. The condition avoidance reaction proceeded for a total of 3 days. On the first day, the rats were placed in a bright compartment, adapted for 20 seconds, turned on, and opened the compartment door. In general, the rats are placed in a dark compartment without lighting, which automatically closes the compartment door. This training was repeated until the rats entered the room without illumination in 20 seconds. On the second day, the rats were placed in a bright compartment, acclimated for 20 seconds, illuminated, and then the compartment door was opened in the same manner. When the rats entered the darkened compartment without lighting, the partition door was automatically closed and the fire was turned off, and a current of 0.5 mA was applied to the bottom of the floor for 5 seconds to receive an electrical shock (training trial). On the third day of the experiment (after 24 hours), if the rats are placed back into the light compartment, the rats are reluctant to enter the dark compartment even when lit, and the time required for the rats to enter the dark compartment, ie, step-through latency (Retention trial). On the other hand, when the rat did not enter the dark compartment, the cut-off time was set to 300 seconds.

As shown in FIG. 2, in the step-through latency time, 281.59 ± 18.41 seconds were recorded in the normal group (NOR) and 109.12 ± 27.16 seconds in the control group (Aβ _1-42 ) (P <0.01), respectively. This suggests that the control group definitely induced memory impairment. In the complex-administered group (Aβ _1-42 + EGCG + LUT), 263.76 ± 34.82 sec was recorded in the step-through latency time, and Aβ _1-42 + LUT 110.56 ± 14.73 sec and Aβ _1-42 + EGCG 137.56 ± Compared with 19.17 sec. (Figure 2, * p < 0.05 and ** p < 0.01: Aβ _25-35 + EGCG + LUT vs. other groups).

Hereinafter, examples of formulations using the compounds will be illustrated, but the present invention is not intended to be limited thereto, but is only specifically described.

Formulation example  One. Sanje  Produce

Epigallocatechin gallate 300 mg

Luteolin 300 mg

Lactose 100 mg

Talc 10 mg

The above components are mixed and filled in airtight bags to prepare powders.

Formulation example  2. Preparation of tablets

Epigallocatechin gallate 300 mg

Luteolin 300 mg

Corn starch 100 mg

Lactose 100 mg

Magnesium stearate 2 mg

After mixing the above components, tablets are prepared by tableting according to the usual preparation method of tablets.

Formulation example  3. Preparation of capsules

Epigallocatechin gallate 150 mg

150 mg luteolin

Crystalline cellulose 3 mg

Lactose 14.8 mg

Magnesium stearate 0.2 mg

The above components are mixed according to a conventional capsule preparation method and filled in gelatin capsules to prepare capsules.

Formulation example  4. Preparation of injections

Epigallocatechin gallate 150 mg

150 mg luteolin

180 mg mannitol

Sterile sterilized water for injection 2974 mg

Na ₂ HPO ₄ .12H ₂ O 26 mg

(2 ml) per 1 ampoule in accordance with the usual injection preparation method.

Formulation example  5. Liquid  Produce

Epigallocatechin gallate 150 mg

150 mg luteolin

10 g per isomer

5 g mannitol

Purified water quantity

Each component was added and dissolved in purified water according to the usual liquid preparation method, and the lemon flavor was added in an appropriate amount. Then, the above components were mixed, and purified water was added thereto. The whole was added with purified water to adjust the total volume to 100 ml, And sterilized to prepare a liquid preparation.

Formulation example  6. Manufacture of health food

Epigallocatechin gallate 1000 mg

Luteolin 1000 mg

Vitamin mixture quantity

70 [mu] g of vitamin A acetate

Vitamin E 1.0 mg

0.13 mg vitamin B1

0.15 mg of vitamin B2

0.5 mg vitamin B6

0.2 [mu] g vitamin B12

10 mg vitamin C

Biotin 10 μg

Nicotinic acid amide 1.7 mg

50 ㎍ of folic acid

Calcium pantothenate 0.5 mg

Mineral mixture quantity

1.75 mg of ferrous sulfate

0.82 mg of zinc oxide

Magnesium carbonate 25.3 mg

15 mg of potassium phosphate monobasic

Secondary calcium phosphate 55 mg

Potassium citrate 90 mg

100 mg of calcium carbonate

24.8 mg of magnesium chloride

Although the composition ratio of the above-mentioned vitamin and mineral mixture is comparatively mixed with a composition suitable for health food as a preferred embodiment, the compounding ratio may be arbitrarily modified, and the above ingredients are mixed according to a conventional method for producing healthy foods , Granules can be prepared and used in the manufacture of health food compositions according to conventional methods.

Formulation example  7. Manufacture of health drinks

Epigallocatechin gallate 150 mg

150 mg luteolin

Vitamin C 15 g

Vitamin E (powder) 100 g

19.75 g of ferrous lactate

3.5 g of zinc oxide

Nicotinic acid amide 3.5 g

Vitamin A 0.2 g

Vitamin B1 0.25 g

Vitamin B2 0.3g

Water quantification

The above ingredients were mixed according to a conventional health drink manufacturing method, and the mixture was stirred and heated at 85 ° C for about 1 hour. The resulting solution was filtered, sterilized in a sterilized 2 L container, It is used in the production of the health beverage composition of the present invention.

Although the compositional ratio is relatively mixed with a component suitable for a favorite drink, it is also possible to arbitrarily modify the compounding ratio according to the regional or national preference such as the demand class, the demanding country, and the use purpose.

Claims (5)

(EGCG) represented by the following general formula (1) and luteolin (LUT) represented by the following general formula (2) as an active ingredient. &Lt; / RTI >
&Lt; Formula 1 >

(2)

The cognitive function disorder according to claim 1, wherein the cognitive dysfunction-related disorder is any one selected from the group consisting of forgetfulness, Alzheimer's disease, cerebrovascular dementia, dementia caused by head injury, and Parkinson's disease. A pharmaceutical composition for preventing or treating diseases associated with promotion and cognitive dysfunction. [3] The composition according to claim 1, wherein the epigallocatechin-3-gallate and luteolin are contained in an amount of 0.1 to 50% by weight based on the total weight of the composition. A pharmaceutical composition for preventing or treating a disease. A method for preventing or ameliorating memory-related and cognitive dysfunction-related diseases comprising, as an active ingredient, epigallocatechin-3-gallate represented by the following formula (1) and luteolin represented by the following formula Health functional foods.
&Lt; Formula 1 >

(2)

5. The method according to claim 4, wherein the cognitive dysfunction-related disorder is any one selected from the group consisting of forgetfulness, Alzheimer's disease, cerebrovascular dementia, dementia caused by head injury, and Parkinson's disease. A health functional food for preventing or improving diseases associated with promotion and cognitive dysfunction.

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