KR101124621B1 - Pharmaceutical Composition for Angiogenesis Stimulation Comprising Sesamin As an Active Ingredient - Google Patents

Abstract

The present invention relates to an angiogenesis promoter, and specifically provides an angiogenesis promoter containing sesamine or a pharmaceutically acceptable salt thereof as an active ingredient. Sesamin or a pharmaceutically acceptable salt thereof of the present invention can induce angiogenesis and thus can be used for the treatment of angiogenic dependent diseases such as ischemic disease and delayed wound healing.

Ag hair, sesamin, angiogenesis promoter, vascular endothelial cell, eNOS, ischemic disease, delayed wound healing

Description

Pharmaceutical Composition for Angiogenesis Stimulation Comprising Sesamin As an Active Ingredient}

The present invention relates to angiogenesis promoters.

Angiogenesis is a process of forming new blood vessels and is known to be regulated by the balance between several types of promoters and inhibitors (J Folkman, Annu . Rev. Med . 57: 1-18, 2006). The angiogenesis process requires a series of different steps, including vascular endothelial cell proliferation, migration, and tube formation. Neovascularization is known to functionally improve ischemia-related tissue damage and injury by providing blood vessels that supply nutrients, growth factors, and oxygen required for metabolism to hypoxia or ischemic tissues and organs due to blood circulation disorders. Accordingly, attempts have been actively made to apply drugs and techniques for promoting angiogenesis to methods for treating angiogenesis-dependent diseases such as ischemic diseases.

The vascular endotherlial growth factor (VEGF), a well-known angiogenesis promoter, is known as myocardial infarction, cerebral ischemic injury, limb ischemia, and wound healing. It has been used for the treatment of various human diseases such as (J Folkman, Annu Rev Med 57 :..... 1-18, 2006; JM Isner & T Asahara, J. Clin Invest 103: 1231-1236, 1999). In addition, angiogenesis promoters such as fibroblast growth factor, epidermal growth factor and platelet-derived epidermal growth factor have been studied for clinical treatment. However, these factors are difficult to separate and purify as proteins and are expensive and have difficulty in clinical application. In recent years, interest in research for the development and establishment of new neovascular inducers has increased.

Sesamin, a potent antioxidant, is the most lignan contained in sesame seed oil (D. Nakano et. al . , J Pharmacol Exp Ther 318: 328-335, 2006), Acanthopanax It is also found in several herbs, such as senticosus ) (J Ryu et. al ., Arch Pharm Res 27: 912-914, 2004). Sesamin is well known to have various pharmacological effects such as chemical cancer prevention, anti-inflammatory, anti-hypertension, and protection against oxidative damage of the liver. However, the effects of sesamin on angiogenesis have not been studied.

Accordingly, the present inventors have tried to develop angiogenesis promoters, and as a result, sesamin directly activates eNOS to induce angiogenesis and promotes angiogenesis by activating vascular endothelial cells associated with angiogenesis, invasion and vascular formation. It was confirmed that the present invention was completed.

It is an object of the present invention to provide an angiogenesis promoter containing sesamine or a pharmaceutically acceptable salt thereof as an active ingredient.

Another object of the present invention to provide a pharmaceutical composition for the prevention or treatment of angiogenic dependent disease containing sesamin or a pharmaceutically acceptable salt thereof as an active ingredient, and a method for preventing or treating the same.

Another object of the present invention to provide a health food for the prevention or improvement of angiogenic dependent diseases containing sesamin or a pharmaceutically acceptable salt thereof as an active ingredient.

In order to achieve the above object, the present invention provides an angiogenesis promoter containing sesamine (sesamin) or a pharmaceutically acceptable salt thereof as an active ingredient.

The present invention also provides a pharmaceutical composition for the prevention or treatment of angiogenic dependent diseases containing sesamine or a pharmaceutically acceptable salt thereof as an active ingredient.

The present invention also provides a health food for preventing or improving angiogenic dependent diseases containing sesamin or a pharmaceutically acceptable salt thereof as an active ingredient.

In addition, the present invention provides a method for preventing or treating angiogenic dependent disease in a subject, comprising administering to the subject a pharmaceutically effective amount of the pharmaceutical composition.

As described above, sesamine or a pharmaceutically acceptable salt thereof of the present invention can induce angiogenesis and thus can be used for the treatment of angiogenic dependent diseases such as ischemic disease and delayed wound healing.

Hereinafter, the term used by this invention is demonstrated.

As used herein, the term "prevention" means any action that promotes angiogenesis by administration of a composition of the invention.

As used herein, the terms "treatment" and "improvement" refer to any action in which angiogenic dependent disease is improved or beneficially altered by administration of a composition of the present invention.

As used herein, the term "administration" means providing a subject with a composition of the present invention in any suitable manner.

The term "individual" as used in the present invention means any animal, such as a human, monkey, dog, goat, pig or rat, having a disease in which angiogenic dependent disease can be improved by administering the composition of the present invention.

As used herein, the term “pharmaceutically effective amount” means an amount sufficient to treat a disease at a reasonable benefit or risk ratio applicable to medical treatment, which means the type of disease, the severity, the activity of the drug, the drug Sensitivity to, time of administration, route of administration and rate of administration, duration of treatment, factors including drug used concurrently, and other factors well known in the medical arts.

Hereinafter, the present invention will be described in detail.

The present invention provides an angiogenesis promoter containing sesamine or a pharmaceutically acceptable salt thereof as an active ingredient.

The sesamin or a pharmaceutically acceptable salt thereof may be isolated from other plants containing the present compound, such as Acanthopnax divaricatus var. Albeofructus, Acanthopanax senticosus, sesame seeds, sesame oil, sesame gourd, etc. Possible, but not limited to.

In a specific embodiment of the present invention, the effect on angiogenesis in sesamin (Formula 1) isolated from albino and sesame gourd was confirmed in vitro using HUVEC (human umbilical vein endothelial cells) cell line. As a result, sesamin induced cell proliferation in a concentration-dependent manner for the proliferation and migration of vascular endothelial cells, which is the most basic step in angiogenesis, and showed higher cell migration effects than the positive control group VEGF (FIGS. 1A and FIG. 1b). In addition, the sesamin was confirmed to increase concentration-dependently the high tubulation effect compared to the control (see Fig. 1c and 1d).

In addition, in the specific example of the present invention, the sesamin was confirmed to have an angiogenic effect compared to the control even in the results of Intravital microscopy assay, and in particular, the case of sesamin was higher than that of the positive control group VEGF. (See FIGS. 1E and 1F).

In addition, in the specific embodiment of the present invention, by the sesamine treatment, the activity of eNOS included in the angiogenesis signaling pathway was increased in a time and concentration-dependent manner, NO production was increased in a concentration-dependent manner, which is NOS (Nitric oxide synthase) It was inhibited to baseline level by the inhibitor 1 mM NMA (N-methyl-L-arginine) (see FIG. 2).

This is a result showing that the hairy white hair extract of the present invention and sesamin or a pharmaceutically acceptable salt thereof can be used as angiogenesis promoter.

The present invention also provides a pharmaceutical composition for the prevention or treatment of angiogenic dependent diseases containing sesamine or a pharmaceutically acceptable salt thereof as an active ingredient.

In a specific embodiment of the present invention, white hair extract and sesamin showed an effect of promoting angiogenesis in vitro and in vivo (see FIGS. 1 and 2), and thus usefully used for the prevention or treatment of angiogenic dependent diseases. Can be.

The angiogenic dependent disease may include, but is not limited to, ischemic disease and delayed wound healing. The ischemic diseases include ulcers, chronic ulcers, ischemia, myocardial infarction, arteriosclerosis, angina pectoris or cerebrovascular dementia.

The sesamine may be used in the form of a pharmaceutically acceptable salt, and includes all salts, hydrates and solvates prepared by conventional methods. The pharmaceutically acceptable salts are useful addition salts formed with free acid. Suitable free acids include organic and inorganic acids, and inorganic acids include hydrochloric acid, bromic acid, sulfuric acid and phosphoric acid. Organic acids include citric acid, acetic acid, lactic acid, tartariac acid and maleic acid. , Fumaric acid, formic acid, propionic acid, oxalic acid, trifluoroacetic acid, benzoic acid, gluconic acid, methanesulfonic acid, glycolic acid, succinic acid, 4-toluenesulfonic acid, galluxuronic acid, embonic acid, glutamic acid or aspart Acids and the like can be used. Furthermore, sesamines according to the present invention may include all salts, hydrates and solvates that can be prepared by conventional methods, as well as pharmaceutically acceptable salts.

Pharmaceutical dosage forms of the pharmaceutical compositions 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 compositions according to the present invention may be formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like, oral preparations, suppositories, and sterile injectable solutions, respectively, according to conventional methods. Can be used. Carriers, excipients and diluents that may be included in the composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl Cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.

When formulated, diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, and surfactants are usually used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, which may include at least one excipient such as starch, calcium carbonate, sucrose in the extract. ) Or lactose, gelatin and the like are mixed. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Oral liquid preparations include suspensions, solvents, emulsions, and syrups, and may include various excipients, such as wetting agents, sweeteners, fragrances, and preservatives, in addition to commonly used simple diluents such as water and liquid paraffin. . Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, suppositories. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, injectable esters such as ethyl oleate and the like can be used. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycero gelatin and the like can be used.

The pharmaceutical composition according to the present invention may be administered in the form of an injection to a site to be mixed with an injectable composition to induce angiogenesis in a mammal, and the injectable composition is preferably an isotonic aqueous solution or suspension. The silver may be sterile or contain adjuvant or other therapeutically useful substances such as preservatives, stabilizers, wetting agents, emulsifier solution accelerators, salts and / or buffers for osmotic pressure control.

The present invention also provides a health food for preventing or improving angiogenic dependent diseases containing sesamin or a pharmaceutically acceptable salt thereof as an active ingredient.

The sesamin or pharmaceutically acceptable salt thereof of the present invention may be added to a food as it is or used with other food or food ingredients, and may be appropriately used according to a conventional method. The amount of the active ingredient to be mixed can be suitably determined according to its use purpose (for prevention or improvement). Generally, in the preparation of food or beverages, the composition of the present invention is added at 0.2 to 20% by weight, preferably 0.24 to 10% by weight relative to the raw materials. However, in the case of long-term intake for health and hygiene or health control purposes, the amount may be below the above range, and the active ingredient may be used in an amount above the above range because there is no problem in terms of safety.

The health food of the present invention may contain various flavors or natural carbohydrates as additional ingredients. Such natural carbohydrates are monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, and polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol and erythritol. Examples of sweeteners include natural sweeteners such as tau martin and stevia extract, synthetic sweeteners such as saccharin and aspartame, and the like. The ratio of the natural carbohydrate is preferably selected in the range of 0.01 to 0.04 parts by weight, preferably about 0.02 to 0.03 parts by weight, per 100 parts by weight of the health food of the present invention.

There is no particular limitation on the kind of food. Examples of the food to which the substance may be added include drink, meat, sausage, bread, biscuit, rice cake, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gums, dairy products including ice cream, various soups , Beverages, alcoholic beverages and vitamin complexes, and includes all healthy foods in the conventional sense.

In addition to the above, the health food of the present invention includes various nutrients, vitamins, electrolytes, flavors, coloring agents, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols. And carbonation agents used in carbonated beverages. In addition, the health food of the present invention may contain a flesh for producing natural fruit juice, fruit juice beverage and vegetable beverage. These components can be used independently or in combination. Although the ratio of such an additive is not critical, it is generally selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the health food of the present invention.

In addition, the present invention provides a method for preventing or treating angiogenic dependent disease in a subject, comprising administering to the subject a pharmaceutically effective amount of the pharmaceutical composition.

In a specific embodiment of the present invention, white hair extract and sesamin have been shown to promote angiogenesis in vitro and in vivo (see FIGS. 1 and 2), and thus may be usefully used for the prevention or treatment of angiogenic dependent diseases. Can be.

The subject is a vertebrate and preferably a mammal, more preferably an experimental animal such as a rat, rabbit, guinea pig, hamster, dog, cat, and most preferably an ape-like animal such as a chimpanzee or gorilla.

The method of administration may be administered orally or parenterally, intraperitoneal, rectal, subcutaneous, intravenous, intramuscular, intrauterine dural, intracerebroventricular or intrathoracic It can be administered by injection.

The pharmaceutically effective amount is 0.0001 to 100 mg / kg, preferably 0.001 to 10 mg / kg, but is not limited thereto. Dosage may vary depending on the weight, age, sex, health status, diet, duration of administration, method of administration, elimination rate, severity of disease, and the like of a particular patient. The pharmaceutical composition of the present invention may be administered several times daily to once a week depending on the half-life. The dosage does not limit the scope of the invention in any aspect.

Hereinafter, the present invention will be described in detail by examples and production examples.

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

< Example  1> Sesamin  refine

<1-1> In the white haired organ Sesamin  refine

Acanthopnax divaricatus var.albeofructus HS-70) 5 kg of stem (Cheonan Materials Co., Ltd. Receiving Ogalpi Farm) was immersed in 4 liters of methanol and stirred well, and then extracted by heating for 5 hours at an extraction temperature of 70 ° C. And filtered through a filter paper. The extract was concentrated under reduced pressure using a concentrator at 50 to 55 ° C., followed by freeze drying to obtain an extract, and 500 g of methanol extract was partitioned between CHCl ₃ and H ₂ O to form CHCl _3. 100 g of fractions were obtained. CHCl ₃ using a gradient system (20: 1-100% EtOAc) with Hexane-EtOAc as the developing solvent. The fractions were subjected to silica gel column chromatography (Kiegelgel 60, 70-230 mesh, manufactured by Merck) to obtain four fractions. From these fractions 3 (23.5 g), 512 mg of sesamine of colorless prism was obtained. Melting point (mp) was 122-124 and identified as sesamine from the following ¹ H-NMR and ¹³ C-NMR spectrum results.

Colorless prism, mp 122-124, ¹ H-NMR (300 MHz, CDCI ₃ ) d: 3.07 (2H, m, H-1 and H-5), 3.90 (2H, dd, J = 9.2, 3.5 Hz, He -4 and He-8), 4.25 (2H, dd, J = 8.7, 7.0 Hz, Ha-4 and Ha-8), 4.74 (2H, d, J = 4.3 Hz, H-2 and H-6), 5.96 (4H, s, 2 x OCH ₂ O), 6.80 (4H, m, H-5 ', H-5, H-6' and H-6), 6.86 (2H, m, H-2 'and H -2). ¹³ C-NMR (75 MHz, CDCI ₃ ) d: 148.3 (s, C-3 'and C-3), 147.5 (s, C-4' and C-4), 135.5 (s, C-1 'and C-1), 119.7 (s, C-6 'and C-6), 108.5 (s, C-5' and C-5), 106.8 (s, C-2 'and C-2), 101.4 (t , 0-CH ₂ -O), 86.1 (d, C-2 and C-6), 72.1 (t, C-4 and C-8), 54.7 (d, C-1 and C-5).

<1-2> In sesame gourd Sesamin  refine

100 g of sesame gourd (the remaining material oiled from sesame oil) was immersed in 1 liter of water, stirred well, extracted with heating for 3 hours at an extraction temperature maintained at 80 ° C, and filtered through a filter paper. The water extract was discarded and the residue was extracted twice with 70% for 3 hours at 80% ethanol (0.6 liter). The extract was concentrated under reduced pressure using a concentrator at 50 to 55 ° C., and then the concentrate was extracted twice with 100 ml of ethyl acetate. The extract was concentrated under reduced pressure using a concentrator at 50 to 55 ° C., and then freeze-dried to obtain an extract, which was added to the ethyl acetate extract with a gradient system (20: 1-100% EtOAc) using hexane-EtOAc as a developing solvent. Silica gel column chromatography was performed to obtain four fractions, and from these fractions 3 (23.5 g), 130 mg of sesamin of colorless prism was obtained.

< Example  2> In vitro  Investigate effect on cell proliferation

White hair organolpi on cell proliferation, the most basic step in angiogenesis The effect of the extract and sesamin was confirmed.

<2-1> Purchasing Reagent

Heparin, streptomycin, penicillin, and M199 (medium 199) were purchased from Invitrogen life technologies (USA), VEGF and basic Fibroblast Growth Factor (bFGF) were purchased from Upstate Biotechnology (USA), N ^G -methyl- _L -arginine) was purchased from Calbiochem (USA) and Matrigel with reduced phosphorylation-specific antibodies and growth factors specific for eNOS and eNOS was purchased from BD Bioscience (USA). Other reagents were purchased from Sigma (USA).

<2-2> Cell Culture

Human umbilical vein endothelial cells (HUVECs) are isolated from human umbilical veins by known methods of collagenase digestion (Lee SJ et. al ., Am . J. Physiol. Heart . Circ . Physiol . 291: 2836-2846, 2006) and then used for 3-7 generations. Cells were cultured in conditions of controlled humidity of 37 ° C., 5% CO ₂ in M199 medium supplemented with 20% 100 U / ml penicillin, 100 μg / ml streptomycin, 3 ng / ml bFGF and 5 U / ml heparin It was.

<2-3> Endothelial Cell Proliferation Test

MTT [3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide] (Carmichael J et al. Cancer Endothelial cell proliferation was measured according to Res 47 (1987) 943-946, 1987). HUVECs were inoculated in gelatin-coated 24-well plates at a density of 2 × 10 ⁴ cells / well. After 24 hours, cells were washed twice with M199 and incubated for an additional 6 hours in M199 with 1% FBS. After 24 hours of VEGF (10 ng / ml), various concentrations of sesamine (SE: 5, 10, 20 and 30 μM), 100 μl of MTT solution (5 mg / ml) was added and the absorbance was 570 nm. Measured at All data are reported as mean ± SD of at least three independent experiments. Statistical comparisons between experimental groups were performed by Student's t-test using one-way ANOVA. P <0.05 was considered statistically significant.

As a result, as shown in Figure 1a it was confirmed that the cells proliferate depending on the concentration of SE.

< Example  3> In vitro  Investigate effects on cell migration

The effect of sesamin on the movement of cells, the most basic step in angiogenesis, is transwell plate (Corning Costar, USA) equipped with 6.5-mm diameter polycarbonate filter (8-mm pore size) according to the known method. (Lee SJ et al ., 2006). Briefly, the bottom surface of the filter was coated with 10 μg gelatin. Freshly made VEGF (10 ng / ml), M199 (1% FBS) containing various concentrations of sesamine (SE: 5, 10, 20 and 30 μM) were placed at the bottom of the wells. The cells were trypsinized and then suspended in M199 containing 1% FBS at a final concentration of 1 × 10 ⁶ cells / ml. Various concentrations of sesamine were treated with the cells for 30 minutes at room temperature before cell inoculation. 100 μl of cell suspension was loaded on top of the wells. The chamber was incubated at 37 ° C. for 4 hours. The cells were fixed and stained with hematoxylin and eosin. Unmoved cells located on the top side of the filter were removed by cotton swap, and cell migration was quantified by measuring the number of cells that migrated to the bottom side of the filter under an optical microscope (200X). Ten fields were measured for each analysis.

As a result, as shown in FIG. 1b, the SE was more effective than the positive control group VEGF.

< Example  4> In vitro  Investigate the effect on tube formation

White hair organs for the formation of in vitro tubular structures The effect of extract and sesamin was determined. Briefly, Matrigel with reduced 250 μl growth factor was injected into a 16 mm diameter culture plate and then polymerized at 37 ° C. for 30 minutes. HUVECs were incubated for 6 hours in M199 medium containing 1% FBS and then harvested by trypsin treatment and suspended in M199 containing 1% FBS. The cells were incubated with VEGF (10 ng / ml), various concentrations of sesamine (SE: 10, 20 and 30 μM) at room temperature for 30 minutes and injected into the Matrigel layer at a density of 2 × 10 ⁵ cells / well. . After 36 hours, the formation of a pipe-like structure was photographed (40 ×). The area covered by the tube network was determined using an optical imaging technique that scans the photo and quantifies it using Image-Pro Plus (Media Cybermetics, USA).

As a result, as shown in Figure 1c by SE was confirmed to have a higher tube forming effect compared to the control, as shown in Figure 1d was confirmed that the effect increases in a concentration-dependent.

< Example  5> In vivo  Microscopic analysis Intravital microscopy assay Angiogenesis through side tablet

White hair organ skin for angiogenesis in vivo The effect of extract and sesamin was determined. Male BALB / c mice (6-8 weeks old) were imported from Orient (Korea) and cultured in a specific pathogen-free (SPF) culture facility. All procedures performed on the experimental animals were performed in accordance with the guidelines of the University Animal Care and Use Committee. For measurement of in vivo angiogenesis using in vivo microscopy, abdominal wall windows were implanted in BALB / c mice according to known methods (Lee SJ et. al ., 2006). Matrigel (100 mL) containing VEGF (100 ng) or sesamine (SE: 20 nmoles; 7.08 μg) was applied to the interior space of the pore. After 4 days, the animals were anesthetized and intravenously injected with 50 ml of 25 mg / ml FITC-labeled dextran (MW 250,000) into the tail vein. Angiogenesis was measured under a Zeiss Axiovert 200M microscope using a 100-W mercury lamp and a blue filter set (here 440-475 nm; emission is 530-550 nm). Fluorescence images were digitized for continuous offline analysis using MetaMorph program (Universal Imaging Corp., USA).

As a result, as shown in Figures 1e and 1f SE was confirmed to have an angiogenic effect compared to the control, especially showed a higher effect than when treated with a positive control VEGF.

< Example  6> eNOS Of activity against

 eNOS is well known to be involved in angiogenesis signaling pathways. Therefore, the effect of sesamin on eNOS activity was determined. Cytolytic buffer (50 mM Tris) in HUVECs treated with 30 μM SE for defined time (10, 20, 30, 45 and 60 minutes) and HUVECs treated with SE (10, 20 and 30 μM) at specified concentrations for 30 minutes. -HCl, pH 8.0, 150 mM NaCl, 1% Nonidet P-40, 0.5% deoxycholic acid, 0.1% SDS) to prepare a cell lysate. Cell lysate (40 μg of protein) was electrophoretically separated on SDS-PAGE and then transferred to PVDF membrane. The membrane was incubated with antibodies to the target protein for 2 hours after reaction with BSA. After culturing the membrane, the expression level of the target protein was measured using an enhanced chemiluminescence system.

As a result, as shown in FIG. 2A, sesamine treatment increased the activity of eNOS in a time and concentration-dependent manner.

< Example  7> NO  Measurement of production

Cellular nitric oxide (NO) production was measured according to the known method in situ using DAF-FM (4-amino-5-methylamino-27-difluorofluorescein) diacetate (Molecular Probes, USA) (Lee SJ et al ., 2006). Briefly, after 1 hour simultaneous treatment with sesamin (10, 20 and 30 μM) and nitric oxide synthase (NOS) inhibitor 1 mM N-methyl-L-arginine (NMA), the cells were serum-free M199 After washing twice, the cells were incubated at 37 ° C. for 1 hour with 5 μM DAF-FM diacetate. After removal of excess probe, the relative amount of intracellular NO was determined from the fluorescence luminescence of DAF-FM using confocal microscopy.

As a result, as shown in Figures 2b and 2c by sesamin treatment, NO production was increased in a concentration-dependent manner, which was suppressed to the ground level by NMA, a nitric oxide synthase (NOS) inhibitor.

< Manufacturing example  1> Preparation of Pharmaceutical Formulations

1. Preparation of powder

2 g of sesamin of the present invention or a pharmaceutically acceptable salt thereof

1 g lactose

The above components were mixed and packed in airtight bags to prepare powders.

2. Preparation of tablets

100 mg of sesamin of the present invention or a pharmaceutically acceptable salt thereof

Corn starch 100 mg

100 mg of milk

2 mg magnesium stearate

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

3. Preparation of Capsule

100 mg of sesamin of the present invention or a pharmaceutically acceptable salt thereof

Corn starch 100 mg

100 mg of milk

2 mg magnesium stearate

After mixing the above components, the capsule was prepared by filling in gelatin capsules according to the conventional method for producing a capsule.

4. Manufacture of rings

1 g of sesamin of the present invention or a pharmaceutically acceptable salt thereof

Lactose 1.5 g

Glycerin 1 g

0.5 g of xylitol

After mixing the above components, it was prepared to be 4 g per ring in a conventional manner.

5. Manufacture of granules

150 mg of sesamin or pharmaceutically acceptable salt thereof of the present invention

Soybean Extract 50mg

Glucose 200 mg

600 mg of starch

After mixing the above components, 100 mg of 30% ethanol was added and dried at 60 ° C. to form granules, and then filled in fabric.

Preparation Example 2 Preparation of Food

Foods containing sesamine or a pharmaceutically acceptable salt thereof of the present invention were prepared as follows.

1. Preparation of Cooking Seasonings

20 to 95 parts by weight of sesamin or a pharmaceutically acceptable salt thereof of the present invention was prepared for health promotion cooking seasoning.

2. Manufacturing of Flour Foods

0.5-5.0 parts by weight of sesamine or a pharmaceutically acceptable salt thereof of the present invention was added to flour, and bread, cake, cookies, crackers and noodles were prepared using the mixture to prepare health promoting food.

3. Manufacture of dairy products

5-10 parts by weight of sesamin or a pharmaceutically acceptable salt thereof of the present invention was added to milk, and various milk products such as butter and ice cream were prepared using the milk.

4. Manufacture of wire

Brown rice, barley, glutinous rice, and yulmu were alphanated by a known method to distribute the dried ones, and then prepared into a powder having a particle size of 60 mesh.

Black beans, black sesame seeds, and perilla were also steamed and dried by a known method, and then ground to a powder having a particle size of 60 mesh.

The dry powders of the grains, seeds, and sesamin of the present invention or pharmaceutically acceptable salts thereof prepared in the following ratio were prepared.

Cereals (30 parts by weight brown rice, 15 parts by weight brittle, 20 parts by weight of barley),

Seeds (7 parts by weight of perilla, 8 parts by weight of black beans, 7 parts by weight of black sesame seeds)

Sesamin of the present invention or a pharmaceutically acceptable salt thereof (3 parts by weight),

(0.5 part by weight),

Foxglove (0.5 part by weight)

< Manufacturing example  3> Manufacturing of beverage

1. Manufacture of health drinks

1000 mg of sesamin or pharmaceutically acceptable salt thereof of the present invention

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 the conventional healthy beverage manufacturing method, and stirred and heated at 85 ℃ for about 1 hour, the resulting solution is filtered and obtained in a sterilized 2 L container, sealed sterilization and then refrigerated and stored Used to prepare the healthy beverage composition of the invention.

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

1 is a diagram showing that angiogenesis was promoted by sesamin:

a: SE efficacy on vascular endothelial cell proliferation;

b: SE efficacy on vascular endothelial cell migration;

c & d: SE potency (c) and its quantitative value (d) for tube formation in vascular endothelial matrigel; And,

e & f: Efficacy of SE on angiogenesis in vivo (e) and its quantitative value (f).

2 is a diagram confirming the increase in the activity of eNOS and intracellular NO production by sesamin:

a: SE efficacy on activation of eNOS protein important for vascular endothelial cell proliferation;

 Left: treatment for a designated time with SE of 30 μM;

 Right: treatment for 30 min with SE at the indicated concentration;

b: Induction effect of nitric oxide production in vascular endothelial cells of SE (30 μM); And,

c: NOS inhibitor inhibits the nitric oxide production induction effect of SE in vascular endothelial cells.

Claims (11)

Angiogenesis promoter containing sesamine or a pharmaceutically acceptable salt thereof as an active ingredient. The method of claim 1, wherein the sesamin is separable from any one plant selected from the group consisting of Acanthopnax divaricatus var. Albeofructus HS-70, Acanthopanax senticosus, sesame, sesame oil and sesame gourd Accelerator, characterized in that. A pharmaceutical composition for preventing or treating angiogenic dependent diseases containing sesamin or a pharmaceutically acceptable salt thereof as an active ingredient. 4. The composition of claim 3, wherein the angiogenic dependent disease is ischemic disease or delayed wound healing. The composition of claim 4, wherein the ischemic disease is selected from the group consisting of ulcers, chronic ulcers, ischemia, myocardial infarction, arteriosclerosis, angina pectoris and cerebrovascular dementia. Health food for the prevention or improvement of angiogenic dependent disease containing sesamin or a pharmaceutically acceptable salt thereof as an active ingredient. 7. The health food according to claim 6, wherein the angiogenic dependent disease is ischemic disease or delayed wound healing. 8. The health food of claim 7, wherein the ischemic disease is selected from the group consisting of ulcers, chronic ulcers, ischemia, myocardial infarction, arteriosclerosis, angina pectoris and cerebrovascular dementia. delete delete delete

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