KR20030055678A - Composition containing an extract of rubiae radix for protecting brain cells and treating brain stroke - Google Patents

Abstract

PURPOSE: A pharmaceutical composition containing a Rubiae Radix extract is provided. The composition has preventing and treating effects on ischemic stroke caused by damage of cerebral nerve cells and thus can be effectively used as a brain-protecting agent for treatment of brain damage. CONSTITUTION: A pharmaceutical composition contains a water extract and a methanol extract of Rubiae Radix and a fraction thereof as an active ingredient and a conventional pharmaceutically acceptable salt. The water extract of Rubiae Radix is a dried powder that is obtained by concentrating a Rubiae Radix extract prepared by any one method of a cold water precipitation, ultrasonic wave and refluxing, under reduced pressure, and then freeze drying. The composition is formulated into powders, granules, tablets, capsules, liquids and injections.

Description

COMPOSITION CONTAINING AN EXTRACT OF RUBIAE RADIX FOR PROTECTING BRAIN CELLS AND TREATING BRAIN STROKE}

The present invention relates to a composition for protecting brain cells and treating stroke comprising cheoncho root extract.

Stroke is divided into two main categories: ischemic stroke, which is caused by an ischemic stroke of the brain tissue due to a decrease or blocking of blood supply to the brain tissue, and a hemorrhagic stroke that causes blood vessels to bleed into the brain tissue. Ischemic stroke accounts for about 80% of all stroke patients, so ischemic stroke is a serious disease. Unlike the existing theories of the immune privileged organs, the nervous system has an inflammatory response in the central nervous system in the case of neurological diseases such as multiple sclerosis, trauma, Alzheimer's disease, cerebral ischemia and various brain injuries. It is reported that this is one of the major causes of neurodegeneration.

Cerebral nerve cell damage caused by stroke may be explained by the release of excessive excitatory neurotransmitters, the production of free radicals, inhibition of protein synthesis, abnormal gene expression, and activation of immune responses. Due to the complexity of cell damage mechanisms, there are no therapeutics that can protect the damage of neurons caused by stroke. Therefore, the development of a substance that can protect the damage of the nervous system caused by cerebral ischemia has a variety of objectives, and the evaluation method is also studied in an in vitro test and in vivo test. Because of the peculiarity of herbal medicines, they have been effective in in vitro tests, but are ineffective in in vivo tests.

For this reason, the development of a substance having a protective effect on brain damage, such as cerebral ischemia using natural products, must be performed simultaneously with in vitro and in vivo tests.

Inhibition of cyclooxygenase-2 (COX-2) in cerebral ischemia has been reported to have neuroprotective effects on the inhibition of production of PGE ₂ and consequent inhibition of glutamate, and many patients with arthritis or pain have already reported COX- Because of the use of inhibitors, it has been reported that epidemiological investigations of stroke incidence in these patients could be new targets for the prevention and treatment of stroke in the future [Iadecola et al. See Iadecola, C. et al., PNAS. , 30 p1294-1299, 2001.

Inflammatory reactions occur in the central nervous system, which has been reported to be one of the leading causes of neurodegeneration. It is thought to consist of drugs that inhibit enzymes or inhibit the deposition of neutrophils (Dirnagl, U. et al; TINS ., 22 , pp391-397, 1999).

In addition, inducible nitric oxide synthase (iNOS), which can release NO from neutrophils penetrated into brain tissue, is induced, and NO is released from induced iNOS, which is known to cause brain cell damage. Neurotoxicity has also been reported to be via NO, and development of neuroprotective agents related to iNOS expression inhibition is expected (Vaughan CJ, Delanty N., Stroke 30: 1969-73, 1999).

Cloth chogeun (Rubiae radix) in the present invention is the dry root and rhizome same genus belonging to the Rubiaceae (Rubiaceae), known as the rake madder (Rubia cordifolia L), madder (Rubia akane Nakai) or greater madder (Rubia chinensis REGEL), etc. The herbal medicines referred to in the present invention include various types of extracts extracted from water or lower alcohols and mixed solvents of the herbal medicines and / or extractable therefrom.

Cheonchogeun is a lump in shape, the remainder of the stem, there are several or dozens of roots. Conical curved, brown or reddish brown, with longitudinal wrinkles. The bark is easily detached and the reddish yellow wood is exposed. The quality is solid, the cross section is yellow and light red and the taste is bitter. X levels include vasodilation, shortening of blood clotting time, increased uterine contractility, excitability of the bladder, lowering blood pressure, dissolving stones, or antibacterial effects. In oriental medicine, bleeding, hemostasis, and bowing are known. It is known to be applied to blood, carcinoma, etc. (Medicinal Pharmacology, Korean Herbal Medicine Reorganization Committee, Dongmyeongsa, Seoul, 2000. See Choon-Geun) and its ingredients include purpurin, alizarin, and pseudo-fur. Ingredients such as pseudopurpurin, munjistin, etc. are known (see Chung-Sup Shin, Shin Min-kyo, Medicinal Metabolism, Yeonglimsa, p929-930 1998). However, there are no reports of the use of this herb in ischemic stroke.

Tezuka et al. ( J. Ethnopharmacology , 77, 209-217, 2001) reported that various herbal drugs inhibit NO release in in vitro experiments, while Rubia cordifolia L also released NO release. Is reported to be suppressed. However, macrophages, the cells used in this study, are cells isolated by stimulation with LPS / IFN-γ in mice intraperitoneally, and it is reported that they inhibit simple NO release. It is not.

Meanwhile, U.S. Patent Nos. 5,821,237 (published on October 13, 1998) and 5,709,847 (published on January 1, 1998) reported that the extract of Cheoncho-geun has anti-inflammatory effects, and also U.S. Patent No. 5,700,451 (published on December 23, 1997) Examples thereof include sunscreens using cheonchokgeun extract. US Patent No. 6,139,854 (published on Oct. 31, 2000) and the like have illustrated the antioxidant activity of the extract of Cheoncho Geun, all these inventions are mainly related to the prevention of skin diseases using the anti-inflammatory activity of Cheoncho Geun extract, The protective effect of cerebellar ischemia is not exemplified anywhere.

Accordingly, the present inventors have completed the present invention as a result of earnestly researching on the assumption that Chunchogeuneun will also be effective in cerebral ischemia.

It is an object of the present invention to provide a pharmaceutical composition and a dietary supplement showing a brain cell protective effect and a stroke prevention and treatment effect.

In accordance with the above object, the present invention provides a composition for preventing and treating brain cells and stroke and brain cells containing cheoncho root extract.

Brain cell protection and memory enhancing composition of the present invention, the cheoncho root extract contains 0.5 to 50% by weight based on the total weight of the composition.

The cheoncho root extract of the present invention is about 1 to 15 times, preferably about 5 to 10 times, water or lower alcohol or about 1: 0.1 to 1:10, preferably 1: 1 of the dried cheoncho root volume. Ultrasonic extraction and reflux extraction in a mixed solvent having a mixing ratio of 1 to 1: 3 at 20 to 100 ° C, preferably 70 to 100 ° C at an extraction temperature of about 1 hour to 2 days, preferably about 2 hours to 1 day It can obtain by extraction methods, such as these.

The present invention provides a composition for protecting and preventing brain cells and stroke, comprising a water extract obtained in the extraction process, a lower alcohol extract, and a soluble extract according to a mixed solvent thereof.

In addition, the cheoncho root extract of the present invention may be performed an additional fractionation process by a conventional fractionation method (Harborne JB Phytochemical methods: A guide to modern techniques of plant analysis. 3rd Ed. Pp 6-7, 1998).

The composition comprising the myelin extract of the present invention may further comprise suitable carriers, excipients and diluents commonly used in the preparation of pharmaceutical compositions.

Carriers, excipients, and diluents that may be included in the composition comprising the herbaceous extract of the present invention 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.

Compositions containing the myelin extract according to the present invention, respectively, in the form of powders, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like oral formulations, external preparations, suppositories and sterile injectable solutions in accordance with conventional methods Can be used.

The amount of Cheoncho Geun extract may vary depending on the age, sex, and weight of the patient, but may be administered once or several times in an amount of 0.1 to 500 mg / kg body weight per day. The dosage of the natural extracts and fractions may be increased or decreased depending on the route of administration, the severity of the disease, sex, weight, age, and the like. Therefore, the above dosage does not limit the scope of the present invention in any aspect.

The composition containing the extract of Cheoncho Geun of the present invention can be used in various forms, such as drugs, foods and beverages for the protection of brain cells and memory enhancement in the above formulation. Examples of the food to which the cheoncho root extract can be added include various foods, beverages, gums, teas, vitamin complexes, and health supplements.

Cheonchogeun extract itself of the present invention is a drug that can be used with confidence even for long-term administration for the purpose of prevention because there is little toxicity and side effects.

The herbaceous extract of the present invention may be added to food or beverage for the purpose of protecting brain cells and preventing stroke. At this time, the amount of the myelin extract in the food or beverage is generally added to the health food composition of the present invention 0.1 to 15% by weight, preferably 1 to 10% by weight of the total food weight, the health beverage composition is 100 The amount may be added in a proportion of 1 to 30 g, preferably 3 to 10 g, based on ml.

The health beverage composition of the present invention has no particular limitations on the liquid component except for containing the cheoncho root extract as an essential ingredient in the indicated ratio, and may contain various flavors or natural carbohydrates as additional ingredients, such as ordinary drinks. 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 mentioned 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 natural carbohydrates 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 salts thereof. , 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 effect on the prevention and treatment of stroke of the composition of the present invention is to confirm the effect of inhibiting ischemic stroke of the myeloid root extract in vivo using the midbrain aortic occlusion model in which the test method has already been established. Is a method of reperfusion by inserting nylon filament into the internal carotid artery to block the midbrain aorta and removing the filament again 120 minutes later. At this time, the cheoncho muscle and methanol extract orally administered to confirm the effect on ischemic stroke.

The present invention is described in more detail based on the following examples, although the present invention is not limited thereto.

Example 1. Preparation of Water Extract of Cheoncho Geun

10 kg of water was added to 1 kg of dried Cheoncho root purchased from Gyeongdong Market, and the extract was heated at reflux for 2 hours at a temperature of 80-100 ° C. The extract was freeze-dried in a freeze dryer (Elela, model FD-5N). This dried material was used as a sample (yield: 8.31%).

Example 2. Methanol Extract Preparation

10 kg of methanol was added to 1 kg of dried Cheoncho root purchased from Gyeongdong Market and heated for 2 hours at a temperature of 50-70 ℃. The extract was lyophilized (Elela, model FD-5N). The dried product was used as a sample (yield: 6.6%).

Example 3 Preparation of Mixed Solvent Extract

A freeze-drier (Elela, model FD-5N) was added to 1 kg of dried Cheoncho-Keun purchased at Gyeongdong Market, and the mixture was heated under reflux for 2 hours at 70-100 ° C. Freeze-drying in) was used as a sample (yield: 7.2%).

Experimental Example 1. Ischemic stroke protection effect using water and methanol extract of Cheoncho-geun

The water and methanol extracts of Chuncho-Keun were administered at 150 mg / kg about 5 minutes before the start of reperfusion in the rat ischemic animal model using rats (Biogenomeics, Korea), and then again after 150 hours About 24 hours after the first operation after administration of the dose, the animal was lethal and brains were extracted, and a brain slice was made using a brain matrix to a thickness of 2 mm, then 2, 3, 5- After staining using triphenyltetrazolinum chloride (TTC) staining, cerebral infarction sites were analyzed using an image analysis system. On the other hand, a dose-dependent experiment was conducted by administering twice at a dose of 75 mg / kg in the same manner as described above. As a result, as shown in Table 1, the cheonchomuscle extract was effectively known as cerebral ischemia and used as a positive control. It was confirmed that the effect is superior to Ebsel (Ebselen) and Baikalein (baicalein).

The protective effect of the following Table 1 is converted in% of the degree of protection for the vehicle (physiological saline or 5% carboxymethylcellulose) administration group.

Cerebral Ischemia Protective Effect Treatment group Protective effect (%) Ebceline (30 mg / kg x 1) 45.2% Bicalane (50 mg / kg x 2 times) 41.1% Cheoncho Geun Water Extract (150 mg / kg x 2 times) 54.7% Cheoncho Geun Water Extract (75 mg / kg x 2 times) 14.4% Cheoncho Geun methanol extract (150 mg / kg x 2 times) 37.1%

Experimental Example 2. Free radical scavenging effect of the extract

DPPH (1,1-diphenyl-2-picrylhydrazyl radical) scavenging function was used as an index of free radical scavenging activity. Since DPPH itself is a free radical, it is known to have a function of removing purple color of DPPH in the case of a compound having antioxidant activity. Experimental methods have already used a known method described in several documents (Vivot, E. et al., J. Ethnopharmacology, 76 pp65-71, 2001), the results are shown in Table 2 below. The DPPH scavenging activity of Chuncho Geun was found to have a moderate effect.

DPPH Scavenging Activity Experiment Results Sample group (1 mg / ml) effect (%) Evceline 5.4% Baikallein 92.2% Cheoncho Geun (Water Extract) 12.1%

Experimental Example 3. Scavenging function of peroxynitrite of extract

It is intended to confirm the mechanism of action of the extract of Cheoncho-Geun extract by checking how much peroxynitrite produced by SIN-1 (3-morpholinosydnonimine) can be eliminated. First, 200 μM of SIN-1 and extracts of various concentrations (0.05 to 300 μg / ml) were extracted in PBS and incubated at 37 ° C. for 20 minutes. After incubation, 100 μM of dihydrorhodamine-123 (DHR123), which is oxidized by peroxynitrite and generates fluorescence, was added to the culture medium and incubated again at 37 ° C. for 30 minutes to give 536 nm (emission), The fluorescence was measured at 500 nm (excitation) to measure the degree of scavenging action of the peroxynitrite of the extract of Cheonchogeum and the results are shown in Table 3 below. As a result, it was confirmed that Chuncho-Keun was very excellent in the scavenging activity of the peroxynitrite produced by SIN-1 (IC ₅₀ : 60 µg / ml).

Peroxynitrite Scavenging Activity of Water Extracts Concentration (µg / ml) % Oxidation of DHR123 10 minutes 20 minutes 40 mins 60 mins 0.05 106.93 96.87 100.65 91.30 0.5 101.94 100.60 97.56 97.74 5 101.05 102.23 97.03 102.07 10 102.38 99.75 99.41 98.71 20 94.71 95.30 91.49 91.95 50 67.55 62.11 56.38 57.82 100 32.42 37.06 33.43 36.21 300 11.26 14.19 11.75 13.77

Experimental Example 4. Acute Toxicity Test of Cheoncho-Geun Extract Using Mouse

Mice were used for the acute toxicity test of various cheoncho root extracts. As a result of the acute toxicity test, no water death was observed at the dose of 2000 mg / kg, and no significant abnormality was found in weight gain and feed intake. Therefore, it was found that the extract of Chuncho Geun is a safe drug.

Hereinafter, the preparation examples of the pharmaceutical composition will be described, but the present invention is not intended to be limited thereto, but is intended to be described in detail.

Formulation Example 1 Preparation of Powder

According to the preparation method of powder in the pharmacopeia formulation, it is prepared in the following ingredient content per one packet.

Cheoncho Geun Water Extract (Dry Powder) ㆍ ······· 300 mg

Lactose · ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ 100 mg

Talc 10 mg

Formulation Example 2 Preparation of Tablet

According to the preparation method of tablets in the pharmacopeia formulation, it is prepared in the following component content per tablet.

Cheoncho Geun Water Extract (Dry Powder) ㆍ ······ 300 mg

Corn starch ㆍ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ 100 mg

Lactose ····················· 100 mg

Magnesium Stearate 2 mg

Formulation Example 3 Preparation of Capsule

According to the preparation method of capsules in the pharmacopeia formulation, it is prepared in the following component content per capsule.

Cheoncho Geun Water Extract (Dry Powder) ······· 300 mg

Corn starch ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ ・ 100 mg

Lactose ····················· 100 mg

Magnesium stearate 2 mg

Formulation Example 4 Preparation of Injection

According to the preparation method of injection in the pharmacopeia formulation, it is prepared in the following component content per ampoules (2 ml).

Cheoncho Geun Water Extract (Dry Powder) ㆍ ······ 300 mg

Sterile distilled water for injection ㆍ ··········

pH regulator ㆍ ·················

Formulation Example 5 Preparation of Liquid

According to the preparation method of the liquid formulation in the Pharmacopoeia General Formulation, it is prepared in the following component content per 100 ml of the liquid formulation.

Cheoncho Geun Water Extract (Dry Powder) ㆍ ······· 1 g

Heterosaccharides ··············· 10 g

Mannitol ················· 5 g

Purified water ㆍ ·················

In addition, the health beverage is prepared as follows.

5-10% of sugar, 0.05-0.3% citric acid, 0.005-0.02% caramel, 0.1-1% of vitamin C are mixed and 79-94% of purified water is mixed to make syrup, and the syrup is 85-98 ° C. Sterilized in 20 ~ 180 seconds in a mixture of cooling water and a ratio of 1: 4 and then prepared carbonated beverage containing Cheoncho root dry water extract by injecting carbon dioxide gas 0.5 ~ 0.82%.

Instant sterilization by homogeneously mixing the subsidiary ingredients such as liquid fructose (0.5%), oligosaccharide (2%), sugar (2%), salt (0.5%), water (75%) and tasteless dry extract. , Packed in a small packaging container such as plastic bottles to prepare a healthy beverage.

Although the composition ratio is a composition suitable for a preferred beverage in a preferred embodiment, the compounding ratio may be arbitrarily modified according to regional and ethnic preferences such as demand hierarchy, demand country, and use purpose.

Since the composition containing the cheonchogeun extract, it has a prophylactic and therapeutic effect against ischemic stroke due to damage of the brain neurons, it can be usefully used as a brain protection agent for treating brain injury.

Claims (4)

A pharmaceutical composition for treating ischemic stroke, comprising water extract of Rubiae radix, methanol extract or fractions fractionated thereof as an active ingredient and mixed with a conventional pharmaceutically acceptable carrier. The method of claim 1, Water extract is a pharmaceutical composition for treating ischemic stroke, characterized in that the dried powder obtained by freeze-drying and then concentrated under reduced pressure of the extract obtained by any one method of cold needle, ultrasonic, reflux. The method of claim 1, A pharmaceutical composition for the treatment of ischemic stroke, wherein the pharmaceutical form is pharmaceutically selected from powders, granules, tablets, capsules, solutions, and injections. A dietary supplement comprising Cheoncho-geun extract and food-acceptable food supplements that exhibit brain cell protection and stroke prevention effects.