KR20110039762A - Composition comprising isolated from cassia tora or cassia obtusifolia or for the prevention and treatment of cognitive dysfunction disorder - Google Patents

Abstract

PURPOSE: A composition for treating dementia and amnesia is provided to suppress beta-secretase and to be used as a pharmaceutical composition and health food. CONSTITUTION: A pharmaceutical composition for preventing and treating cognitive disorder contains a compound of general formula I as an active ingredient, isolated from Cassia obtusifolia or Cassia tora. The compound is gluco-obtusifolin, gluco-aurantio-obtusin, alaternin, or chrysophanoltriglucoside. The pharmaceutical composition for preventing and treating cognitive disorder contains a compound of general formula II isolated from Cassia obtusifolia or Cassia tora.

Description

Composition comprising isolated from Cassia tora or Cassia obtusifolia or for the prevention and treatment of cognitive dysfunction disorder}

The present invention relates to a pharmaceutical composition for preventing and treating cognitive dysfunction and a health functional food containing a compound isolated from Cassia tora or Cassia obtusifolia as an active ingredient.

JA Duke, Handbook of phytochemical constituents of GRAS herbs and other economic plants, Herbal Reference Library , CRC press, Boca Raton, Florida, pp. 143144, 1992

JS Choi, HJ Lee et al., In vitro antimutagenic effects of anthraquinone aglycones and naphthopyrone glycosides from Cassia tora, Planta Medica , 63 , pp. 1114, 1997

NJ Hao et al., Structure-activity relationships of anthraquinones as inhibitors of 7-ethoxycoumarin O-deethylase and mutagenicity of a-amino-3-methylimidazo [4,5-f] quinoline, Mutation Research , 328 , pp .183191, 1995

HS Choi et al., Effect of the ethanol extracts of Cassia tora L. of antioxidative compounds and lipid metabolism in hepatoxicity of rats-induced by ethanol, Journal of the Korean Society of Food Science and Nutrition , 30 (6) , pp.11771183, 2002

U. Patil et al., Hypolipidemic activity of seeds of Cassia tora Linn, Journal of Ethnopharmacology , 90 (2-3) , pp.249252, 2004

Ellman et al., Ellman GL, Courtney KD, Andres V Jr, Feather-stone RM, A new and rapid colorimetric determination of acetylcholinesterase activity. Biochem Pharmacol. , Jul; 7 , pp. 88-95, 1961

Ermoleff J, et al., Proteolytic activation of recombinant pro-memapsin 2 (pro-beta-secretase) studied with new fluorogenic substrates. Biochemistry. 10; 39 (40): 12450-6 , 2000 Oct

Ebert U, Kirch W. Scopolamine model of dementia: electroencephalogram findings and cognitive performance. Eur J Clin Invest ., 28, pp. 944-949, 1998

The present invention relates to a composition for the prevention and treatment of cognitive impairment containing the compound isolated from the defect or ultra-determination as an active ingredient.

The central nervous system, composed of the brain and spinal cord, is the central center for life phenomena, which is responsible for all functions of the human body, from sensory and (involuntary) movements to thinking, memory, emotions, and language. It is an essential organ. Therefore, in all cases such as rapid death of nerve cells caused by stroke, trauma, death of slow neurons that cause central nervous system degenerative diseases such as senile dementia and Parkinson's disease, which are represented by Alzheimer's disease This results in irreversible dysfunction of the network and eventually a permanent loss of the human body's function. Geriatric dementia, represented by Alzheimer's disease, has a characteristic of increasing proportionately with the extension of the average human life expectancy and the modernization of medical welfare facilities. According to the statistics of the Korea Institute of Health and Social Affairs, since the elderly population in Korea entered the aged society in 2000, the population reached 3.97 million in 2003, accounting for 8.3% of the elderly population, and 14.4% in 2019. It is expected to enter. Among the elderly population aged 65 years or older, the elderly had more than one chronic disease. In Western society, Alzheimer's disease occurs in about 10% of the population aged 65 and over, and about 40-50% of the population aged 80 and older.In the United States, more than 5 million people have this disease. It is estimated in dollars. In Korea, more than 200,000 people have dementia. In the United States, it is estimated to reach 14 million people by 2030, more than double the present, and more than 350% by 2050.

As the life expectancy increases due to the development of medical technology, the proportion of patients with degenerative neuropathy is increasing rapidly. Dementia is a disease that causes a significant drop in quality of life, both personally and socially, and makes you and those around you unhappy. It is the fourth leading cause of death for the elderly, following cancer, heart disease, and stroke. Alzheimer's disease is classified into Alzheimer's dementia and vascular dementia. Alzheimer's dementia is a type of dementia commonly observed in the West. Vascular dementia is a dementia caused by hypertension, stroke, and hyperlipidemia due to abnormal blood vessels leading to the brain, which kills hippocampus and limbic system nerves. Although these two types of dementia have different causes, they have the same mechanism in that they cause damage to memory by reducing the action of acetylcholine, also known as memory transmission neurons.

Acetylcholine with quaternary amine structure is a neurotransmitter and acetylcholinesterase acts to hydrolyze acetylcholine into choline. In patients with dementia, the concentration of acetylcholine, a neurotransmitter, is reduced, and the inhibition of acetylcholinesterase increases the concentration of acetylcholine in the brain, thereby improving symptoms of dementia patients. There are four drugs that have been developed and used as treatments for dementia by FDA approval: tacrine, donepezil, galanthamine, and rivastigmine, all of which are acetylcholinesterase inhibitors.

It is a herb that uses the seeds of Cassia tora L. and Cassia Obtusifolia L. belonging to the legumes, and it is chrysophanol, emodin, rhein, etc. It contains various anthraquinones that exhibit physiological activity (JA Duke, Handbook of phytochemical constituents of GRAS herbs and other economic plants, Herbal Reference Library , CRC press, Boca Raton, Florida, pp. 143144, 1992).

It has been reported that the deficiency has been used to antagonize aflatoxin B1 in the Ames test, which has been used for the purpose of nominal, blue liver, and stool. JS Choi, HJ Lee et al., In vitro antimutagenic effects of anthraquinone aglycones and naphthopyrone glycosides from Cassia tora, Planta Medica , 63 , pp. 1114, 1997). In addition, the deficiencies also have anti-mutant efficacy (NJ Hao et al., Structure-activity relationships of anthraquinones as inhibitors of 7-ethoxycoumarin O-deethylase and mutagenicity of a-amino-3methylimidazo [4,5-f] quinoline, Mutation Research , 328 , pp. 183 191, 1995), HS Choi et al., Effect of the ethanol extracts of Cassia tora L. of antioxidative compounds and lipid metabolism in hepatoxicity of rats-induced by ethanol, Journal of the Korean Society of Food Science and Nutrition , 30 (6) , pp. 11771183, 2002), hyperlipidemic action (UK Patil et al., Hypolipidemic activity of seeds of Cassia tora Linn, Journal of Ethnopharmacology , 90 (2-3) , pp.249252, 2004). However, there are no reports of amnesia improvement yet.

Therefore, the present inventors, in order to determine the effect of preventing and treating cognitive impairment of compounds isolated from Cassia tora or Cassia obtusifolia, beta secretase (β-secretase) activity inhibition experiment, acetylcholine Esterase inhibitory activity and passive avoidance experiments using the scoflavin-induced forgetfulness model inhibited β-secretase and confirmed the learning-promoting efficacy of the scopolamine-induced memory-depleted animals. The present invention has been completed.

In order to achieve the above object, the present invention is a cognitive impairment containing a compound represented by the following formula (I) or (II) isolated from Cassia tora or Cassia obtusifolia as an active ingredient It provides a pharmaceutical composition for the prevention and treatment of.

(I)

(I)

In this formula,

R ₁ to R ₈ are each independently selected from a hydrogen atom, a hydroxyl group, a C ₁ to C ₃ lower alkyl group, a C ₁ to C ₃ lower alkoxy group, O-Glu, O-Glu-Glu or O-Glu-Glu-Glu One or more substituents, where Glu means Glucose.

As a preferable compound of the said General formula (I) compound, R <_1> is a hydrogen atom or a compound group which is a methyl group; R ₂ is a hydrogen atom, a hydroxyl group, -O-Glu group of compounds; R ₃ is a hydrogen atom, a hydroxyl group, a methoxy group, -O-Glu-Glu-Glu; R ₄ is a group of compounds which are a hydrogen atom or a hydroxyl group; R ₅ is a hydrogen atom, a group of compounds which are a methoxy group; R ₆ is a hydrogen atom, a group of compounds which is a hydroxy group; R <_7> and R <_8> can mention the compound group which is a hydrogen atom.

Most preferred compounds of the general formula (I) are gluco-obtusifolin, gluco-aurantio-obtusin, alaternin, and chrysopanol. -Triglucoside (Chrysophanol-triglucoside), but the scope of the present invention is not limited thereto.

(II)

(II)

In this formula,

R ₁ to R ₈ are each independently selected from a hydrogen atom, a hydroxyl group, a C ₁ to C ₃ lower alkyl group, a C ₁ to C ₃ lower alkoxy group, O-Glu, O-Glu-Glu or O-Glu-Glu-Glu One or more substituents, where Glu means Glucose;

X, Y is a group selected from CH, O, S or N.

As a preferable compound of the said General formula (II) compound, R <_1> is a compound of a hydrogen atom or a methyl group; R ₂ is a group of compounds which are a hydrogen atom or a hydroxyl group; R ₃ is a hydrogen atom, O-Glu, O-Glu-Glu or O-Glu-Glu-Glu; R ₄ is a group of compounds which are hydrogen atoms; R ₅ is a compound group that is a hydrogen atom, a hydroxy group or a methoxy group; R ₆ or R ₇ is a compound group that is a hydrogen atom; X, Y is CH or O, a compound group is mentioned.

Most preferred compounds of the general formula (II) include nor-rubrofusarin glucoside, Rubrofusarin-gentiobioside, torallactone-genthiobioside ( Toralactone-gentiobioside), but the scope of the present invention is not limited thereto.

The compounds of formula (I) or formula (II) inhibit beta secretase and exhibit efficacious learning effects in memory-induced memory groups induced by scopolamine, thereby preventing cognitive impairment. And it can be usefully used as a pharmaceutical composition and dietary supplement for the treatment.

Therefore, the present invention is to prevent and treat cognitive impairment containing the compound represented by the formula (I) or formula (II) isolated from Cassia tora or Cassia obtusifolia as an active ingredient To provide a pharmaceutical composition for.

The above cognitive dysfunction disorder includes general cognitive dysfunction disorder, preferably Alzheimer's dementia, cerebrovascular dementia, pick disease, Creutzfeldt-jakob disease, head injury Dementia or Parkinson's disease, more preferably Alzheimer's disease.

Compositions comprising a compound of the present invention may further comprise a suitable carrier, excipient or diluent according to conventional methods.

Carriers, excipients and diluents that may be included in the compositions 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.

The compositions comprising the compounds of the present invention are each formulated in the form of oral dosage forms, external preparations, suppositories, or sterile injectable solutions, such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., in accordance with conventional methods. Can be used.

More specifically, when formulating the composition, it can be prepared using a diluent or an excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, a surfactant, and the like. 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, sucrose in the extract. ), Lactose, gelatin and the like can be mixed. In addition to simple excipients, lubricants such as magnesium stearate and talc may also be used. Liquid preparations for oral use include suspensions, solvents, emulsions, and syrups, and include various excipients such as wetting agents, sweeteners, fragrances, and preservatives, in addition to commonly used simple diluents such as water and liquid paraffin. Can be. Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations and 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 a suppository base, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

Preferred dosages of the compounds of the present invention depend 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, but may be appropriately selected by those skilled in the art. However, for the desired effect, the compound of the present invention may be administered in an amount of 0.0001 to 100 mg / kg, preferably in an amount of 0.001 to 100 mg / kg once to several times daily. The compound of the present invention in the composition may be formulated in an amount of 0.0001 to 50% by weight based on the total weight of the total composition.

In addition, the pharmaceutical dosage forms of the compounds of the present invention may be used in the form of their pharmaceutically acceptable salts, and may be used alone or in combination with other pharmaceutically active compounds as well as in a suitable collection.

The pharmaceutical composition of the present invention can be administered to mammals such as mice, mice, livestock, humans, etc. by various routes. All modes of administration can be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intrauterine dural or intracerebroventricular injection.

The present invention is a health function for the prevention and improvement of cognitive impairment containing a compound isolated from the Cassia tora or Cassia obtusifolia represented by the formula (I) and formula (II) as an active ingredient Provide food.

The compounds of the present invention can be used in various ways, such as drugs, food and beverages for the prevention and improvement of cognitive dysfunction. Foods to which the compound of the present invention may be added include, for example, various foods, beverages, gums, teas, vitamin complexes, health supplements, and the like, and may be used in the form of powders, granules, tablets, capsules, or beverages. have.

Since the compound of the present invention has little toxicity and side effects, it is a drug that can be used safely even for long-term administration for the purpose of prevention.

The compound of the present invention may be added to food or beverage for the purpose of preventing and treating cognitive impairment. At this time, the amount of the compound in the food or beverage is generally added to the health food composition of the present invention to 0.01 to 15% by weight of the total food weight, the health beverage composition is 0.02 to 30 g based on 100 ml, preferably Can be added in a ratio of 0.3 to 10 g.

The health beverage composition of the present invention, in addition to containing the compound as an essential ingredient in the indicated proportions, has no particular limitation on the liquid component 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 disaccharides such as glucose and fructose such as maltose, sucrose and the like and polysaccharides such as dextrin, cyclodextrin and the like Sugar, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those mentioned above, natural flavoring agents (tauumatin, stevia extract (for example, 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 compounds of the present invention, the composition is a variety of nutrients, vitamins, minerals (electrolytes), synthetic and natural flavors, such as flavoring agents, coloring and neutralizing agents (such as cheese, 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 in the range of 0 to about 20 parts by weight per 100 parts by weight of the composition of the present invention.

Compounds represented by general formula (I) and general formula (II) isolated from the shortcomings of the present invention or the ultra-low definition of the present invention inhibit beta secretase (β-secretase) and suppress the memory loss induced by scopolamine. By showing the efficacy of learning, it can be usefully used as a pharmaceutical composition and health functional food for the prevention and treatment of cognitive impairment.

Hereinafter, the present invention will be described in detail with reference to Examples and Experimental Examples.

However, the following Examples and Experimental Examples are only illustrative of the present invention, and the content of the present invention is not limited to the following Examples and Experimental Examples.

Reference Example 1. Drugs and Reagents

Scopolamine and tacrine were purchased from Sigma-Aldrich Chemistry Co., and the other reagents were the highest commercially available.

Reference Example 2 Preparation of Experimental Animals

Six-week-old ICR mice (26-28 g) and 14-week-old SD rats were supplied from Orient Co., Ltd. (Seoul, Korea) and used for 5 days in a clean cage at Kyung Hee University College of Pharmacy. Water and feed were freely consumed, and the temperature (23 ± 2 ° C), humidity (55 ± 10%) and contrast cycle (12 hours) were automatically adjusted.

Reference Example 3. Statistics Processing

All experimental results were statistically analyzed using one way analysis of variance (ANOVA), and significance was tested at p <0.05 level using the Student-Newman-Keuls Test if significant. Was carried out.

Example 1 Preparation of Cassia tora Extract

After drying and pulverizing the clarifier (Gyeongdong Market, Seoul, Korea), 2,000 ml of methanol was added to 500 grams of the clarifier powder, reflux-cooled for 3 hours at 80 ° C., filtered and concentrated under reduced pressure to obtain 51.5 g of clarifier methanol extract. .

Example 2. Isolation and Identification of Compounds from Cassiae Extracts

2-1. Isolation and Identification of Gluco-obtusifolin

51.5 g of the methanol extract obtained in Example 1 was placed on a silica gel column and chromatographed using chloroform: methanol (20: 1 → 1: 1 (v / v)) as the elution solvent to give the same aspect in thin layer chromatography. The fractions were combined, concentrated, and divided into 10 fractions, and 8.4 g of the fifth fraction was placed on a silica gel column, and chloroform: methanol: water (15: 1: 0.1 → 1: 1 (v / v)) was used as an elution solvent. Chromatography was performed to combine and concentrate those showing the same behavior in thin layer chromatography to obtain 36 mg of Gluco-obtusifolin having the following physical properties.

¹ H-NMR (DMSO- d ₆ , 400 MHz) δ: 12.76 (OH), 7.87 (1H, s, H-4), 7.74 (1H, t, J = 7.9 Hz, H-6), 7.64 (1H , d, J = 7.0 Hz, H-5), 7.34 (1H, d, J = 8.3 Hz, H-7), 5.02 (1H, d, J = 7.5 Hz, H-1 '), 4.42-3.06 ( m, glucosyl-H), 3.89 (3H, s, OCH ₃ ), 2.43 (3H, s, CH ₃ );

¹³ C-NMR (DMSO- d ₆ , 100 MHz) δ: 188.0 (C-9), 181.4 (C-10), 161.4 (C-8), 154.7 (C-2), 153.3 (C-1), 141.5 (C-7), 136.5 (C-6), 132.6 (C-3), 129.7 (C-11), 125.2 (C-4), 124.6 (C-14), 124.1 (C-13), 118.4 (C-5), 116.9 (C-12), 103.8 (C-1 '), 77.4 (C-3'), 76.4 (C-5 '), 74.1 (C-2'), 69.8 (C-4 '), 61.5 (C-6'), 60.9 (OCH ₃ ), 17.6 (CH ₃ ).

2-2. Isolation and Identification of Gluco-aurantio obtusin

The same reaction process as in Example 2-1 was carried out to obtain 70 mg of gluco-aurantio obtusin having the following physical properties.

¹ H-NMR (DMSO- d ₆ , 400 MHz) δ: 13.08 (OH), 7.78 (1H, s, H-4), 7.43 (1H, s, H-5), 5.16 (1H, d, J = 7.3 Hz, H-1 '), 2.28 (3H, s, CH ₃ );

¹³ C-NMR (DMSO- d ₆ , 100 MHz) δ: 187.6 (C-9), 179.9 (C-10), 156.2 (C-8), 155.7 (C-2), 155.5 (C-6), 147.3 (C-1), 141.4 (C-7), 132.4 (C-3), 128.2 (C-11), 125.9 (C-4), 124.6 (C-14), 123.6 (C-13), 113.5 (C-12), 106.2 (C-5), 100.3 (C-1 '), 77.3 (C-3'), 76.5 (C-5 '), 73.2 (C-2'), 69.3 (C-4 '), 61.1 (CH ₃ ), 60.5 (CH ₃ ), 60.4 (C-6'), 16.5 (CH ₃ ).

2-3. Isolation and Identification of Alaternin

The same reaction process as in Example 2-1 was conducted to obtain 5 mg of alaternin having the following physical properties.

¹ H-NMR (DMSO- d ₆ , 400 MHz) δ: 12.02 (OH), 7.47 (1H, H-4), 7.08 (1H, H-5), 6.52 (1H, H-7), 2.34 (3H , s, CH ₃ );

_{^{13 C-NMR (DMSO- d 6}} , 100 MHz) δ: 190.0 (C-9), 179.8 (C-10), 164.3 (C-8), 165.6 (C-6), 150.1 (C-2), 149.1 (C-1), 135.5 (C-3), 131.2 (C-11), 122.9 (C-14), 122.8 (C-4), 113.9 (C-13), 108.9 (C-12), 108.5 (C-5), 107.1 (C-7), 16.2 (CH ₃ ).

2-4. Isolation and Identification of Chrysophanol-triglucoside

The same reaction process as in Example 2-1 was conducted to obtain 10 mg of chrysophanol-triglucoside having the following physical properties.

¹ H-NMR (DMSO- d ₆ , 360 MHz) δ: 12.90 (OH), 7.76 (1H, t, J = 7.6 Hz, H-6), 7.72 (1H, brs, H-4), 7.66 (1H , dd, J = 2.0, 7.6 Hz, H-5), 7.59 (1H, brs, H-2), 7.35 (1H, dd, J = 2.0, 7.6 Hz, H-7), 5.28 (1H, brd, J = 6.0 Hz, H-1 '), 4.43 (2H, brd, J = 6.0 Hz, H-1'',H-1'''), 5.25-3.0 (m, glucosyl-H), 2.53 (3H , brs, CH ₃ );

¹³ C-NMR (DMSO- d ₆ , 90 MHz) δ: 187.7 (C-9), 182.1 (C-10), 161.4 (C-1), 158.3 (C-8), 147.6 (C-3), 136.2 (C-11), 134.4 (C-6), 132.4 (C-14), 124.4 (C-4), 122.8 (C-5), 121.4 (C-7), 118.3 (C-12), 118.3 (C-13), 116.8 (C-2), 104.2 (C-1 '''), 103.1 (C-1'), 100.5 (C-1 ''), 88.2 (C-3 ''), 77.0 (C-3 '''), 76.5 (C-3'), 76.2 (C-5 '''), 76.1 (C-5''), 75.8 (C-5'), 73.9 (C-2 '''), 73.5 (C-2 '), 72.4 (C-2''), 70.2 (C-4'''), 69.8 (C-4 '), 69.0 (C-4''), 68.5 ( C-6 '), 61.2 (C-6'''), 60.8 (C-6 ''), 22.0 (CH ₃ ).

2-5. Isolation and Identification of Nor-rubrofusarin glucoside

The same reaction process as in Example 2-1 was carried out to obtain 100 mg of nor-rubrofusarin glucoside having the following physical properties.

¹ H-NMR (DMSO- d ₆ , 400 MHz) δ: 14.94 (OH), 10.33 (OH), 7.06 (1H, s, H-10), 6.72 (1H, d, J = 1.6 Hz, H-9 ), 6.68 (1H, d, J = 1.9 Hz, H-7), 6.16 (1H, s, H-3), 4.97 (1H, d, J = 7.5 Hz, H-1 '), 5.10-3.10 ( m, glucosyl-H), 2.37 (3H, s, CH ₃ );

¹³ C-NMR (DMSO- d ₆ , 100 MHz) δ: 183.7 (C-4), 168.6 (C-2), 162.1 (C-5), 159.7 (C-8), 158.3 (C-6), 152.3 (C-11), 140.4 (C-14), 106.9 (C-3), 106.5 (C-13), 103.0 (C-12), 102.5 (C-9), 101.6 (C-7), 101.3 (C-1 '), 99.9 (C-10), 77.2 (C-5'), 76.4 (C-3 '), 73.5 (C-2'), 69.5 (C-4 '), 60.6 (C- 6 '), 20.1 (CH ₃ ).

2-6. Isolation and Purification of Rubrofusarin-gentiobioside

The same reaction process as in Example 2-1 was carried out to obtain 10 mg of Rubrofusarin-gentiobioside having the following physical properties.

¹ H-NMR (DMSO- d ₆ , 300 MHz) δ: 14.87 (OH), 7.18 (1H, s, H-10), 6.93 (1H, s, H-9), 6.81 (1H, s, H- 7), 6.18 (1H, s, H-3), 5.06 (1H, d, J = 7.5 Hz, anomeric H), 5.05 (1H, d, J = 7.5 Hz, anomeric H), 5.00-3.00 (glucosyl- H), 3.88 (3H, s, OCH ₃ ), 2.38 (3H, s, CH ₃ );

¹³ C-NMR (DMSO- d ₆ , 75.5 MHz) δ: 183.6 (C-4), 168.7 (C-5), 161.8 (C-2), 161.0 (C-8), 157.5 (C-6), 152.3 (C-11), 140.2 (C-14), 107.6 (C-13), 106.6 (C-3), 103.5 (C-1 '& C-10), 101.1 (C-12), 100.8 (C -7), 100.7 (C-9), 99.7 (C-1``), 76.8 (C-3 '), 76.6 (C-5''), 76.3 (C-3''), 75.5 (C- 5 '), 73.4 (C-2'& C-2 ''), 70.1 (C-4 '), 69.7 (C-4''), 68.8 (C-6'), 61.6 (C-6 '' ), 55.4 (OCH ₃ ), 20.0 (CH ₃ ).

2-7. Isolation and Identification of Toralactone-gentiobioside

The same reaction process as in Example 2-1 was carried out to obtain 15 mg of toralactone-gentiobioside having the following physical properties.

¹ H-NMR (DMSO- d ₆ , 400 MHz) δ: 12.60 (OH), 7.13 (1H, s, H-5), 6.93 (1H, d, J = 2.1 Hz, H-6), 6.51 (1H , s, H-4), 5.09 (1H, d, J = 7.8 Hz, anomeric H), 4.20 (1H, d, J = 7.8 Hz, anomeric H), 3.89 (3H, s, OCH ₃ ), 2.23 ( 3H, s, CH ₃ );

¹³ C-NMR (DMSO- d ₆ , 100 MHz) δ: 166.8 (C-3), 162.6 (C-1), 161.3 (C-9), 157.4 (C-7), 152.5 (C-10), 141.5 (C-14), 132.3 (C-12), 111.6 (C-5), 109.1 (C-13), 104.1 (C-4), 103.6 (C-8), 101.8 (C-1 '), 100.9 (C-1``), 100.3 (C-6), 98.4 (C-11), 76.8 (C-3 '), 76.6 (C-5''), 76.4 (C-3``), 75.5 (C-5 '), 73.5 (C-2''), 73.5 (C-2'), 70.1 (C-4 '), 69.7 (C-4''), 68.8 (C-6'), 61.6 (C-6``), 55.4 (OCH ₃ ), 18.8 (CH ₃ ).

Experimental Example 1. Measurement of inhibition of acetylcholinesterase activity

Acetylcholinesterase inhibitory activity was measured by Elman's method (Ellman et al., Ellman GL, Courtney KD, Andres V Jr, Feather-stone RM, A new and rapid colorimetric determination of acetylcholinesterase activity.Biochem Pharmacol. , Jul; 7 , pp .88-95, 1961).

The brains of the mice were extracted and placed in a 10-fold volume of PBS1 (12.5M sodium phosphate buffer pH 7.0, 400mM NaCl), and ground at 500 rpm using a Teflon glass tube and centrifuged at 1000 X g for 10 minutes. PBS1 and Triton X-100 were added to the supernatant, stirred for 30 minutes, centrifuged at 10000 X g for 10 minutes, and the supernatant was used as an enzyme. 1.5 ml of the dissolved drug, 2.6 ml of buffer, 75 mM acetylthioholine iodide solution (20 µl), and 0.1 ml of Elman's reagent are all incubated at 25 ° C for 30 minutes, followed by enzyme solution. Was added 0.4 ml and the absorbance was measured at 410 nm for 5 minutes at 30 second intervals. Acetylcholinesterase inhibitory activity was calculated using Equation 1 below, and the results are shown in Table 1.

ingredient Inhibition Rate (%) Formula (I) Gluco Optosipoline 51.2 ± 4.1 Gluco-Auranthio-Optucin 2.1 ± 0.3 Alaternin 10.24 ± 1.5 Chrysophanol Triglucoside 45.50 ± 4.2 General formula (II) NOR-Ruburofusarin Glucoside 33.2 ± 3.2 Luburofusarine-Genthiobioside 12.22 ± 1.8 Torallactone-genthiobioside 7.41 ± 2.2

As a result of the experiment, as shown in Table 1, the inhibitory activity of acetylcholinesterase activity of each component at the concentration of 100 μg / kg gluco-obtusifolin, Nor-rubrofusarin glucoside (nor-rubrofusarin glucoside) It was confirmed that this excellent inhibitory activity was exhibited.

In addition, the inhibitory activity of each concentration of gluco-obtusifolin having the most excellent inhibitory activity is shown in FIG. 1.

Experimental Example 2. Measurement of β-secretase Activity Inhibition

Inhibition activity against β-secretase, an enzyme associated with Alzheimer's dementia in the present invention was performed using the method described in the literature as follows (Ermolieff J, et al., Proteolytic activation of recombinant pro -memapsin 2 (pro-beta-secretase) studied with new fluorogenic substrates.Biochemistry. 10; 39 (40): 12450-6 , 2000 Oct).

Substrate solution was made by dissolving a certain amount of synthetic substrate (MCA) Ser-Glu-Val-Asn-Leu-Asp-Ala -Glu-Phe-Lys (DNP) in dimethylsulfoxide (DMSO). (β-secretase) was used by dissolving in 0.1 M sodium acetate buffer (pH 4.0). Inhibition reaction was carried out by taking a certain amount of enzyme solution and sample solution into 0.5 mL cell and maintained in a fluorescent spectrophotometer controlled at 37 ° C for 10 minutes, and then added a certain amount of substrate solution and reacted with the excitation wavelength 328 nm over time. And fluorescence was measured at an emission wavelength of 393 nm. The control was measured for fluorescence in the same manner as above without adding the sample solution. Inhibitory activity was calculated using Equation 2 below.

Inhibitory activity (%) = (initial reaction rate of sample / initial reaction rate of control) × 100

As a result, as shown in Figure 2, in the beta cryptase (β-secretase) inhibitory activity test, a substance that produces neurotoxic Aβ in Alzheimer's dementia patients, crysophanol triglucoside, glucoside Aurantio optocin (gluco-aurantio-obtusin), nor-rubrofusarin glucoside, gluco-obtusifolin, and alaternin exhibited inhibitory activity in a concentration-dependent manner. The degree of activity is gluco-aurantio-obtusin,>gluco-obtusifolin> nor-rubrofusarin glucoside = alaternin = chrysopanol triglucoside ( crysophanol triglucoside).

Experimental Example 3. Passive avoidance experiment using the forgetfulness model induced by scopolamine

Each component was dissolved in 10% Tween 80 (Polyoxyethyl enesorbitan monooleate: Sigma, U.S.A.) and administered orally at various doses. The positive control group was administered 10 mg / kg of tacrine and 10% Tween.

Was administered intraperitoneally to the scopolamine was dissolved in distilled water and 30 minutes later the drug administration by using a method disclosed in a dose of 1 mg / kg (Ebert U, Kirch W. Scopolamine model of dementia:. Electroencephalogram findings and cognitive performance Eur J Clin Invest ., 28, pp. 944-949, 1998), 30 minutes after scopolamine administration, the rats were placed in the illuminated bright compartment and the guillotine door opened into the dark compartment after 20 seconds of search time. It was. Mice that did not enter the dark side within 40 seconds after the guillotine door was opened were excluded from the experiment. After the guillotine door was opened, the time until the rat entered the dark side was measured. Once the rat entered the dark side, the guillotine door was closed and a 0.5 mA electric shock flowed through the bottom of the grid for three seconds and the mouse remembered this electrical action.

The experiment was conducted 24 hours after the study test. After the 20 second search time, the time required for the guillotine door to open and the 4 foots to enter the dark side (latency time) was measured up to 300 seconds. The longer the time taken, the better the learning and memory of passive avoidance.

Nor-rubrofusarin glucoside, gluco-obtusifolin, chrysophanol triglucoside having excellent activity in the inhibitory activity of acetylcholinesterase activity of Experimental Example 1 Figure 3 shows the results of passive avoidance experiments using the scoflavin-induced forgetfulness model. As a result of comparative experiments using 0.1 mg / kg, respectively, excellent effects were observed in NOR ruburofusarin glucoside and gluco optupoline, and the results of concentration-dependent experiments are shown in FIGS. 4 and 5. Knorr ruburofusarin glucoside showed similar effects to tacrine, which was used as a positive control, whereas gluco optofolipoline showed better efficacy than tacrine, which was used as a positive control.

Experimental Example 4. Acute Toxicity Test

In order to test the acute toxicity of the compounds of the present invention, according to the regulations of the Korea Food and Drug Administration (KFDA), male ICR mice were purchased from Orient, Seoul, Korea, and placed in a clean cage at Kyung Hee University College of Pharmacy. The experiment was carried out after adaptation for about 5 days.

Ten mice in each group were orally administered with a compound of the present invention at a dose of 0.625-2 g / kg and observed for two weeks.

After administration of the test substance, mortality, clinical symptoms, and changes in body weight were observed, blood biochemical tests were performed, and autopsy was performed to observe abnormalities of organs and thoracic organs.

As a result, there were no clinical symptoms or deaths in all animals treated with the test substance, and no toxicity change was observed in weight change, blood test, blood biochemistry test and autopsy findings.

The compound of the present invention did not show a toxic change when administered orally to a dose of 2g / kg in mice, 1-1000 mg / kg orally administered to humans, 0.2-100 mg / cm ₂ when applied to the skin effect Was determined to be a substance.

Examples of the formulation of the composition are described below, but are not intended to limit the present invention but to explain in detail only.

Formulation Example 1 Preparation of Powder

Gluco Optosipoline 20 mg

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

Knorr loufufusarin glucoside 10 mg

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

Gluco-Optupoline 10 mg

3 mg of crystalline cellulose

Lactose 14.8 mg

Magnesium Stearate 0.2 mg

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

Knorr loufufusarin glucoside 10 mg

Mannitol 180 mg

Sterile distilled water for injection 2974 mg

Na ₂ HPO ₄ 12H ₂ O 26 mg

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

Gluco Optosipoline 20 mg

10 g of isomerized sugar

5 g of mannitol

Purified water

According to the conventional method for preparing a liquid, each component is added and dissolved in purified water, lemon flavor is added, the above components are mixed, purified water is added, the whole is adjusted to 100 ml by adding purified water, and then filled in a brown bottle. The solution is prepared by sterilization.

Formulation Example 6 Preparation of Healthy Food

Knorr loufufusarin glucoside 1000 mg

Vitamin mixture proper amount

70 μg of Vitamin A Acetate

Vitamin E 1.0 mg

Vitamin B1 0.13 mg

Vitamin B2 0.15 mg

Vitamin B6 0.5 mg

0.2 μg of vitamin B12

Vitamin C 10 mg

10 μg biotin

Nicotinic Acid 1.7 mg

50 μg folic acid

Calcium Pantothenate 0.5mg

Mineral mixture

Ferrous Sulfate 1.75 mg

Zinc Oxide 0.82 mg

Magnesium carbonate 25.3 mg

Potassium monophosphate 15 mg

Dibasic calcium phosphate 55 mg

Potassium Citrate 90 mg

Calcium Carbonate 100 mg

Magnesium chloride 24.8 mg

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 Preparation of Healthy Drink

Gluco Optosipoline 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 according to a conventional healthy beverage production method, and then stirred and heated at 85 ℃ for about 1 hour, the resulting solution is filtered and obtained by sterilization in a sterilized 2 L container, sealed sterilized and then stored in the present invention For the preparation of healthy beverage compositions.

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

[ National R & D project supporting this invention ]

[ Assigned unique number ] no. 10524

[ Department Name ] Industry-University Support Center affiliated with Seoul Development Institute

[ Research Project Name ] Aging Disease System

[ Project Name ] Commercialization of KHUR-4X, an antidepressant made from natural products

[ Organizer ] Kyung Hee University Industry-Academic Cooperation Group

[ Research Period ] December 01, 2008 ~ November 30, 2009

1 is a diagram showing the effect of inhibiting the acetylcholine esterase activity according to the concentration of gluco optofolipoline,

Figure 2 is a diagram showing the effect of inhibiting beta cyclase activity of each component,

3 is a view showing the effect on the passive avoidance experiment of each component,

Figure 4 is a diagram showing the efficacy of each concentration for the passive avoidance experiment of gluco optofolipoline,

Figure 5 is a diagram showing the effect of the concentration for the passive avoidance experiments of the NOR Lubulofusarin glucoside.

Claims (9)

A pharmaceutical composition for the prevention and treatment of cognitive impairment containing a compound represented by the following general formula (I) isolated from a defect or ultra-determination as an active ingredient:

(I) In this formula, R ₁ to R ₈ are each independently selected from a hydrogen atom, a hydroxyl group, a C ₁ to C ₃ lower alkyl group, a C ₁ to C ₃ lower alkoxy group, O-Glu, O-Glu-Glu or O-Glu-Glu-Glu One or more substituents, where Glu means Glucose. The compound of claim 1, wherein the compound of Formula (I) is R ₁ is a hydrogen atom or a methyl group; R ₂ is a hydrogen atom, a hydroxyl group, -O-Glu group of compounds; R ₃ is a hydrogen atom, a hydroxyl group, a methoxy group, -O-Glu-Glu-Glu; R ₄ is a compound group that is a hydrogen atom or a hydroxyl group; R ₅ is a hydrogen atom, a group of compounds which are a methoxy group; R ₆ is a hydrogen atom, a group of compounds which is a hydroxy group; R ₇ and R ₈ is a pharmaceutical composition for the prevention and treatment of cognitive impairment containing a compound which is a hydrogen atom as an active ingredient. The method of claim 2, wherein the general formula (I) compound is Gluco-obtusifolin, Gluco-aurantio-obtusin, Alaternin, or Cree Pharmaceutical composition that is Crysophanol-triglucoside. A pharmaceutical composition for the prevention and treatment of cognitive impairment containing a compound represented by the following general formula (II) isolated from a defect or ultra-determination as an active ingredient:

(II) In this formula, R ₁ to R ₈ are each independently selected from a hydrogen atom, a hydroxyl group, a C ₁ to C ₃ lower alkyl group, a C ₁ to C ₃ lower alkoxy group, O-Glu, O-Glu-Glu or O-Glu-Glu-Glu One or more substituents, where Glu means Glucose; X, Y is a group selected from CH, O, S or N. The compound of claim 4, wherein the compound of Formula (II) is R ₁ is a hydrogen atom or a methyl group; R ₂ is a group of compounds which are a hydrogen atom or a hydroxyl group; R ₃ is a hydrogen atom, O-Glu, O-Glu-Glu or O-Glu-Glu-Glu; R ₄ is a group of compounds which are hydrogen atoms; R ₅ is a compound group that is a hydrogen atom, a hydroxy group or a methoxy group; R ₆ or R ₇ is a compound group that is a hydrogen atom; X, Y is a pharmaceutical composition for the prevention and treatment of cognitive impairment containing a compound which is CH or O as an active ingredient. The compound according to claim 5, wherein the general formula (II) compound is Nor-rubrofusarin glucoside, Rubrofusarin-gentiobioside, or torallactone-genthio Pharmaceutical composition that is a bioseed (Toralactone-gentiobioside). The method of claim 6, wherein the cognitive dysfunction disorder is Alzheimer's dementia, cerebrovascular dementia, pick disease, Creutzfeldt-jakob disease, dementia due to head injury and Parkinson's disease Composition. The health functional food for preventing and improving cognitive impairment according to claim 1 or 4, wherein the functional compound contains a compound represented by the above general formula (I) or (II) isolated from the defector or the ultra-clarity as an active ingredient. The dietary supplement of claim 8 in the form of a powder, granule, tablet, capsule or beverage.

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