KR101669717B1 - Antiobesity composition comprising Capsicoside G - Google Patents

Abstract

The present invention relates to an anti-obesity composition comprising Capsicoside G, and more particularly to an anti-obesity composition comprising Capsicoside G having an anti-obesity effect as an active ingredient through inhibition of differentiation into adipocytes A pharmaceutical composition for therapeutic use, a food composition for preventing or ameliorating obesity, a quasi-drug composition and a cosmetic composition. The composition comprising the capsicoside G of the present invention as an active ingredient inhibits the differentiation of adipose precursor cells into adipocytes and inhibits the expression of PPAR-γ, C / EBP-α, aP2 and FAS involved in adipocyte differentiation Therefore, it can be effectively used as a food or medicament for preventing or treating various diseases that may be caused by obesity or obesity, and as a body line management cosmetic which inhibits fat production.

Description

Antiobesity composition comprising Capsicoside G < RTI ID = 0.0 >

The present invention relates to an anti-obesity composition comprising Capsicoside G, and more particularly to an anti-obesity composition comprising Capsicoside G having an anti-obesity effect as an active ingredient through inhibition of differentiation into adipocytes A pharmaceutical composition for therapeutic use, a food composition for preventing or ameliorating obesity, a quasi-drug composition and a cosmetic composition.

Obesity or obesity refers to the excess weight of a person's body weight, which is pathologically referred to as the World Health Organization's (WHO) standard body mass index (BMI) of 30 or more. If it is 25 or more, it is diagnosed as obesity. In most cases, weight means more than normal, but even if you do not have a lot of weight, the proportion of fat in your body is high. Obesity may not be a problem in itself, but it has serious social problems and secondary complications caused by excessive fat. For example, obesity can significantly increase the incidence of hyperlipidemia, hypertension, arteriosclerosis, diabetes, fatty liver, joint disorders, and the like.

As such, obesity causes various adult diseases. Increased weight can lead to diabetes and lower blood sugar, which can lead to hypertension. If the body weight is 20% higher than normal weight, the likelihood of developing diabetes increases to 10 times that of normal people. In addition, cardiovascular and pulmonary function may be impaired and life expectancy may be shortened. Complex factors such as hypertension, diabetes, hypercholesterolemia, and atherosclerosis may put pressure on the heart function and lead to heart failure and sudden death. In addition to women, the incidence of endometrial cancer is high, and cholelithiasis is more likely to occur. Men are more likely to develop colon cancer and prostate cancer. In addition, as the weight increases, people can not help but be conscious of the gaze that they are looking at, and if the obsession about it becomes worse, the symptoms of avoiding people and living alone are generated. The inferiority feeling that comes from the body also affects the self by losing confidence in his life.

In order to prevent obesity and maintain a healthy weight, balanced diet and regular exercise should be maintained throughout the lifetime. In particular, fat cells can be reduced in size once they are made, but it is impossible to remove them naturally and it is left permanently in the human body, so it is important to prevent obesity from beginning. Thus, primarily, you can reduce your weight by reducing the amount of calories consumed and increasing activity and exercise.

However, as described above, since the fat cells once formed can not be removed, there are cases where a drug such as a diet meal intake and a fat absorption reduction is taken.

Current treatments for treating obesity include drugs that act on the central nervous system, affect appetite, and drugs that act on the gastrointestinal tract to inhibit absorption. Drugs such as fenfluramine and dexfenfluramine which inhibit the serotonin (5HT) nervous system, drugs such as ephedrine and caffeine through the noradrenergic nervous system, and recently, drugs acting on the serotonin and noradrenergic nervous system And sibutramine, which inhibits obesity, are commercially available. In addition, drugs that inhibit obesity by acting on the gastrointestinal tract are typically Orrustat, which has recently been approved as a treatment for obesity, by inhibiting the lipase produced by the pancreas and reducing fat absorption. However, pfuramilamine has been recently banned due to side effects such as primary pulmonary hypertension or heart valve lesion. Other drugs have also been associated with problems such as decreased blood pressure and lactic acidosis, There is a problem that patients can not use.

As a result, an adipocyte differentiation inhibitor has emerged as a preventive or therapeutic method for obesity with few side effects. Fat stored in fat cells is used as an important energy source in the body. However, as the obesity progresses, not only the number of fat cells increases but also the synthesis of a large amount of triglyceride (TG, Triglyceride) And concomitant with changes in the expression of various genes, as well as morphological changes including increased size of adipocytes. On the other hand, according to the storage of fat, the increase in the size of adipocytes can increase its diameter to about 20 times, and as a result, the cell volume is increased to several thousand times. However, since the process of differentiation of new lipid precursor cells into adipocytes is not effective as a dietary regulator, in order to control the fundamental treatment or inhibition of obesity, the differentiation process of adipocytes It is important to adjust. The adipocyte differentiation is stimulated by stimuli such as insulin-like growth factor-1 and growth hormone. In this process, the CCAAT enhancer-binding protein (C / EBP) family, PPAR-γ proliferator-activated receptor-γ). These transcription factors promote the differentiation of adipocytes with adipocyte regulatory factors and increase the expression level of enzymes such as adipocyte fatty acid binding protein (aP2) and fatty acid synthase (FAS) . On the other hand, it has been reported that excessive accumulation of triglycerides is involved in the progress of fatty liver [J. Clin. Invest., 98,1575-1584 (1996)].

In recent years, studies have been conducted to search for substances that inhibit adipocyte differentiation based on the idea that the number of adipocytes produced by inhibiting differentiation of adipocytes can be regulated and the extra energy accumulated thereby can be released It is actively proceeding. In general, among the various methods for developing a new drug, experimental modification of the existing drug or synthesis and function of a new substance requires much time and investment.

On the other hand, Capsicoside G is a novel furostanol saponin extracted from a plant of the genus Capsicum, and has a basic structure in relation to the Capsicoside G and is used for treatment of skin diseases (U.S. Patent Publication No. 2011-0046080) and Antimicrobial activity ( J. Agric. Food Chem . 2002. Jun; 50 (15): 4310-16) and the like have been known, but the use of the above-mentioned anti-obesity has not been known at all. In addition, it has been known that the capsicoside G can be separated from red pepper seeds. It has been known that a compound isolated from red pepper or red pepper can have an anti-obesity effect. Heretofore, however, the anti-obesity activity has been limited to capsaicin and capsiate Capsicoside G is known to be only due to compounds such as capsiate ( Nat . Prod . Rep . 2011. Jul; 28 (9): 1493-1533).

Under such circumstances, attention has been paid to capecicoside G which can be separated from red pepper seeds. It has been found that capecicoside G has an effect of inhibiting differentiation into adipocytes in lipoatocytes and is useful for preventing or treating obesity The present invention has been completed.

It is an object of the present invention to provide a pharmaceutical composition for preventing or treating obesity which comprises capsicoside G as an active ingredient.

Another object of the present invention is to provide a food composition for preventing or improving obesity comprising caprosicoside G as an active ingredient.

It is still another object of the present invention to provide a quasi-drug composition for preventing or improving obesity, which comprises capsicoside G as an active ingredient.

It is still another object of the present invention to provide a cosmetic composition for preventing or improving obesity comprising caprosicoside G as an active ingredient.

In order to achieve the above object, the present invention provides a pharmaceutical composition for preventing or treating obesity, which comprises capsicoside G as an active ingredient.

The term "capricoside G" in the present invention is a compound represented by the following formula (1), which may be synthesized by a chemical synthesis method or separated from red pepper, red pepper seed or red pepper seed root. The capsicoside G has been known for its basic structure, skin diseases, antimicrobial use, etc. However, the present invention has revealed a new anti-obesity use.

[Chemical Formula 1]

The term "obesity " in the present invention means an excessive state of body fat, which means that the body mass index is clinically 25 in the case of Korea and 30 or more according to the World Health Organization (WHO). The body mass index (BMI) is calculated as body weight (kg / m ² ) relative to the height of the kidney. Generally, it means that the body weight is higher than the normal value. However, when the body fat percentage of the body composition is high, even if the body weight does not go out, it is diagnosed as obesity. Obesity can shorten life and put you at risk for diseases like hypertension, diabetes, heart disease and some cancers. Obesity is likely to cause health problems, and as the degree of obesity increases, the risk also increases. Obesity is caused by a wide variety of reasons and many causes are often combined. The most frequent causes are genetic influences, physiological effects, food intake and eating disorders, lifestyle, weight, pregnancy or drugs. The first signs of obesity are above average weight and symptoms are as follows: sleep problems, sleep apnea, shortness of breath, varicose veins, skin problems, osteoarthritis due to gallstones or weight loss, Obesity is also associated with diseases such as hypertension, hyperglycemia (diabetes), hypercholesterolemia, and hypertriglyceridemia. Once you are overweight, it is not easy to get out of it, so obesity is a lifelong experience. However, diseases or problems caused by obesity can improve if weight is reduced.

The term "prevention" in the present invention means all actions that inhibit or delay the onset of obesity by the administration of the pharmaceutical composition according to the present invention, and "treatment" Means any action that alters or alleviates symptoms of an individual.

In the present invention, the prevention or treatment of obesity can be achieved by the activity of inhibiting the differentiation of adipose precursor cells into adipocytes. More specifically, PPAR-gamma, C / EBP-alpha, aP2, May be accomplished by reducing the expression of FAS and ultimately by inhibiting the production of intracellular triglycerides (TG, Triglycerides).

In the present invention, the term "lipid precursor cell" refers to a cell having the ability to differentiate into a lipoprotein, which is first produced from stem cells, during the differentiation stage of adipocytes. When lipid precursors grow to 100% confluency, And the differentiation into the local taxa proceeds slowly. The process by which lipid precursor cells are differentiated into adipocytes is a phenomenon caused by differentiation inducers in the serum added to the culture medium. Therefore, the addition of differentiation-promoting factors (insulin, dexamethasone, 3-isobutyl-1-methylxanthine) to the culture medium can further promote differentiation of cells. On the other hand, differentiation inhibitory factors (actinomycin D, Differentiation may be inhibited by the addition of tumor necrosis factor a, bromo-deoxyuridine, or the like. Finally, the differentiated lipid precursor cells become mature adipocytes such as accumulation of intracellular fat, and the activation of enzymes inducing adipocyte-specific gene expression and fat accumulation occurs. The most common adipocyte differentiation promoting factors known so far include insulin and insulin-like growth factor-I (IGF-I).

The term "peroxisome proliferator activated receptor-γ (PPAR-γ)" as used herein refers to a nuclear hormone receptor superfamily which is PPARs (peroxisome proliferator activated receptors) And PPAR-δ are included in the superfamily and are known to function as regulators of adipocyte differentiation. In the present invention, the term "C / EBP (CCAAT enhancer binding protein)" is one of transcription factors having a basic-leucine zipper motif. These are not expressed in adipocytes, (granulocyte) or hepatocyte, and is known to play an important role in the final differentiation of the cells. As used herein, the term "aP2 (adipocyte protein 2) " refers to an adipocyte-specific fatty acid binding protein which is induced by PPAR-γ in adipocytes, It is known as a gene that functions in induction of differentiation. In the present invention, the term "FAS (fatty acid synthase)" refers to a major enzyme involved in the fatty acid biosynthesis pathway. It is composed of a multifunctional polypeptide and includes acetyl-CoA and malonyl- It is known to be an important regulator of obesity, involved in energy metabolism, while producing long chain fatty acids from coenzyme A (malonyl-CoA).

Adipocyte differentiation refers to a process in which a preadipocyte is transformed into a mature adipocyte through proliferation and differentiation. In this process, lipid accumulation, aP2 involved in lipogenesis involved in hormone production, And FAS expression. Such gene expression is known to be regulated by the transcription factors, PPAR and C / EBP. These transcription factors induce expression at different time points in the differentiation process of adipocytes, and gradually induce adipocyte differentiation by controlling the expression of various adipocyte-specific genes through mutual interaction. Thus, the transcription factors regulate the expression of a series of genes involved in adipogenesis, which is the process of becoming a mature adipocyte through the process of proliferation and differentiation of preadipocytes, In particular, the C / EBP family and PPAR-γ are transcription factors that play an important role in the process of inducing adipocyte differentiation by the hormone, and play a role in regulating adipocyte differentiation as a whole, . In addition, C / EBP-β plays an important role in the early differentiation process of lipid precursor cells by promoting PPAR-γ expression, and promotes the late differentiation process of lipid precursor cells through strong synergism with PPAR-γ do.

Capsicoside G of the present invention has effects of weight loss, reduction of body fat, reduction of blood triglyceride content and reduction of blood cholesterol content. According to a specific embodiment of the present invention, when differentiation of 3T3-L1 adipose precursor cells is induced using an adipocyte differentiation medium containing IBMX (3-isobutyl-1-methylxanthine), dexamethasone and insulin (MDI solution) As a result, it was confirmed that the content of triglyceride was decreased according to the treatment concentration of the composition by inhibiting adipocyte production by adding the capsicose G of the present invention (FIG. 5). In addition, it was confirmed that the addition of capecicoside G of the present invention to the 3T3-L1 adipose precursor cells induced differentiation, and the effect of inhibiting the generation of triglycerides was greatest at the early stage of differentiation into adipocytes 6).

In order to confirm the effect of the capsicoside G of the present invention on the effect of inhibiting the differentiation of lipid precursor cells, the expression of PPAR-γ, C / EBP-α, aP2 and FAS, which are the differentiation factors of lipid precursor cells, As a result, it was confirmed that the expression of all of the genes was decreased, and it was confirmed that the composition of the present invention had an activity of inhibiting the differentiation of lipid precursor cells (Fig. 7). Accordingly, the composition of the present invention inhibits the differentiation of lipid precursor cells by inhibiting the expression of adipogenic proteins by reducing the expression of major transcription factors, thereby ultimately reducing intracellular triglyceride production , Thereby indicating that it is effective in improving or treating obesity.

The pharmaceutical composition comprising the compound of Formula 1 of the present invention may further comprise a pharmaceutically acceptable carrier. The term "pharmaceutically acceptable" of the present invention means that it exhibits properties that are not toxic to the cells or humans exposed to the composition. Such carriers may be used without limitation as long as they are known in the art such as buffers, preservatives, wetting agents, solubilizers, isotonic agents, stabilizers, bases, excipients and lubricants. In addition, the pharmaceutical composition of the present invention can be formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups and aerosols, oral formulations, external preparations, suppositories, . Furthermore, it can be used in the form of an external preparation for skin in the form of ointments, lotions, spray agents, patches, creams, powders, suspensions, gels or gels. Examples of carriers, excipients and diluents that can be included in the composition of the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate cellulose, polyvinylpyrrolidone , Water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid formulations for oral administration include tablets, pills, powders, granules, capsules and the like, which may contain at least one excipient such as starch, calcium carbonate, sucrose, sucrose), lactose, gelatin, and the like. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid preparations for oral use include water, liquid paraffin, which is a simple diluent commonly used in suspension, liquid, emulsion, syrup, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, Excipients such as wetting agents, sweeteners, perfumes, preservatives, and the like. Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Examples of the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. Examples of the suppository base include witepsol, macrogol, tween 61, cacao butter, laurin, glycerogelatin and the like.

Meanwhile, the pharmaceutical composition of the present invention is administered in a pharmaceutically effective amount. The term "administering" of the present invention means introducing a predetermined substance into an individual by an appropriate method, and the administration route of the composition can be administered through any conventional route so long as it can reach the target tissue. But are not limited to, intraperitoneal, intravenous, intramuscular, subcutaneous, intradermal, oral, topical, intranasal, intrathecal, rectal. The composition may also be administered by any device capable of transferring the active agent to the target cell.

The term "individual " refers to all animals, including mice, mice, livestock, etc., including humans who have developed or are capable of developing obesity. Preferably, it may be a mammal, including a human.

The term "pharmaceutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment and not causing side effects, and the effective dose level is determined by the patient's sex, Including, but not limited to, medicaments and other medical fields that are used in combination, or in combination with, or in combination with, a pharmaceutically acceptable carrier, excipient, condition, type of condition, severity, activity of the drug, sensitivity to the drug, Can be readily determined by those skilled in the art according to known factors. However, for the desired effect, the composition of the present invention is preferably administered at 0.2 to 200 mg / kg of body weight per day. The administration may be carried out once a day or divided into several times. The dose is not intended to limit the scope of the invention in any way.

Accordingly, in another aspect, the present invention provides a method of treating obesity comprising the step of administering to a subject a pharmaceutical composition comprising the compound represented by Formula 1 above.

In the present invention, the term "individual" means all animals, including humans, who have developed or are suffering from obesity as described above, and the obesity can be effectively prevented or treated by administering the pharmaceutical composition of the present invention to a subject. The pharmaceutical composition of the present invention can be administered in parallel with the existing therapeutic agent for obesity.

In another aspect, the present invention provides a food composition for preventing or improving obesity, which comprises capsicoside G as an active ingredient. The Capsicoside G is a compound represented by Formula 1, wherein the Capsicoside G and its anti-obesity use are as described above, and thus can be used as an active ingredient of an anti-obesity food composition.

When Capsicoside G of the present invention is used as a food additive, Capsicoside G can be added as it is, or it can be used together with other food or food ingredients, and can be suitably used according to ordinary methods. The amount of the active ingredient to be mixed can be suitably determined according to the intended use (prevention, health or therapeutic treatment). Generally, in the production of food or beverage, the composition of the present invention is added in an amount of 0.01 to 10% by weight, preferably 0.05 to 1% by weight based on the raw material. However, in the case of long-term ingestion intended for health and hygiene purposes or for the purpose of controlling health, the amount can also be used in the above-mentioned range.

There is no particular limitation on the kind of the food. Examples of the food to which the above substances can be added include dairy products including meat, sausage, bread, chocolate, candy, snack, confectionery, pizza, ramen, other noodles, gums, ice cream, various soups, drinks, tea, Alcoholic beverages, and vitamin complexes, all of which include healthy foods in a conventional sense.

The health beverage composition of the present invention may contain various flavors or natural carbohydrates as an additional ingredient such as ordinary beverages. 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 generally about 0.01 to 0.04 g, preferably about 0.02 to 0.03 g per 100 ml of the composition of the present invention.

In addition to the above, the composition of the present invention may further contain various nutrients, vitamins, electrolytes, flavors, colorants, pectic acids and salts thereof, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, A carbonating agent used in a carbonated beverage, and the like. In addition, the composition of the present invention may contain flesh for the production of natural fruit juices, fruit juice drinks and vegetable drinks. These components may be used independently or in combination. The proportion of such additives is not critical, but is generally selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the composition of the present invention. These components can be used independently or in combination.

In another aspect, the present invention provides a quasi-drug composition for preventing or ameliorating obesity, which comprises capsicoside G as an active ingredient.

That is, the capsicoside G of the present invention can be added to a quasi-drug composition having the anti-obesity activity and promoting the degradation of triglycerides as well as the cellulite reducing effect like capsaicin, which is well known for the purpose of preventing or improving obesity. When Capsicoside G of the present invention is used as a quasi-drug additive, Capsicoside G may be added as it is or may be used in combination with other quasi-external components, 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 the intended use (prevention, health or therapeutic treatment).

Preferably, the quasi-drug composition may be disinfectant cleaner, shower foam, gagrin, wet tissue, detergent soap, hand wash, humidifier filler, mask, ointment or filter filler.

In another aspect, the present invention provides a cosmetic composition for preventing or ameliorating obesity comprising caprosicoside G as an active ingredient.

That is, the capsicoside G of the present invention can be added to a cosmetic composition having a cellulite reducing effect which promotes the decomposition of triglycerides for the purpose of preventing or improving obesity. When Capsicoside G of the present invention is used as a cosmetic additive, Capsicoside G can be added as it is or can be used in combination with other cosmetic ingredients, and can be suitably used according to a conventional method. The amount of the active ingredient to be mixed can be suitably determined according to the intended use (prevention, health or therapeutic treatment).

Preferably, the formulation of the cosmetic composition is selected from the group consisting of softening agents, convergent lotion, nutritional lotion, nutritional cream, massage cream, essence, eye cream, eye essence, cleansing cream, cleansing foam, cleansing water, pack, powder, Cream, body oil, body essence, makeup base, foundation, hair dye, shampoo, rinse or body cleanser.

The composition comprising the capsicoside G of the present invention as an active ingredient inhibits the differentiation of adipose precursor cells into adipocytes and inhibits the expression of PPAR-γ, C / EBP-α, aP2 and FAS involved in adipocyte differentiation Therefore, it can be effectively used as a food or medicament for preventing or treating various diseases that may be caused by obesity or obesity, and as a body line management cosmetic which inhibits fat production.

1 is a graph showing the results of ¹ H-NMR (nuclear magnetic resonance) spectroscopy and ¹³ C-NMR spectroscopy of Capsicoside G as an active ingredient of the present invention.
FIG. 2 is a graph showing the results of measurement of ghost MBR (gHMBC) spectrum of Capsicoside G, an active ingredient of the present invention.
3 is a graph showing electrospray ionization mass spectrometry (ESI-MS) spectral results of Capsicoside G, an active ingredient of the present invention.
Fig. 4 is a chart showing the cytotoxicity of the composition comprising the capsicoside G of the present invention as an active ingredient. Fig.
Figure 5 is a graph depicting concentration-dependent (0, 1, 5, 10, 20 [mu] g / ml) fat production inhibition. The effect of inhibiting the differentiation of adipocyte precursor cells into adipocytes according to the concentration of the capsicoside G of the present invention was quantified by oil red dyeing and analysis and microscopic observation photographs were shown.
6 is a diagram showing time-dependent lipogenesis inhibition. The conditions of treatment with the capsicoside G of the present invention after induction of differentiation of adipose precursor cells to the respective times (0 to 2, 0 to 4, 0 to 6, 2 to 4, 2 to 6 and 4 to 6 days) The results of the quantitative determination and the microscopic observation of the effect of inhibiting the differentiation of adipocytes from preadipocytes by oil red staining and analysis are shown.
Figure 7 shows the effect on lipogenesis gene and protein expression. The degree of expression of lipogenesis-related genes and proteins, (A) PPAR-γ, (B) C / EBP-α, (C) aP2 and (D) FAS according to the concentration of the capsicoside G of the present invention was Western blot And quantification was carried out.

Hereinafter, the constitution and effects of the present invention will be described in more detail through examples. These examples are only for illustrating the present invention, and the scope of the present invention is not limited by these examples.

Example  1: red pepper From the degreasing pail Anti-obesity  Extraction, separation and purification of physiologically active substances

A 500-mg portion of n-BuOH (n-BuOH) obtained by fractionation of the ethyl alcohol fraction of skimmed red pepper was subjected to solvent system fractionation using Medium Pressure Liquid Chromatography (MPLC) Separation was carried out by fractionation. The MPLC was a Combiflash Rf instrument from TELEDYNE ISCO, and the cartridge column was YMC-DispoPack AT (ODS-25, 120 g) from YMC. In addition, formic acid diluted with 0.1% of formaldehyde in water to a concentration of 0.1% in acetonitrile was used as solvent B. Solvent slope was applied at the ratio of 5%, 10%, 20%, 25%, 40% and 100% of the solvent B, respectively, and the detector was detected at 254 nm. The flow rate was maintained at 10 mL / min and the aliquot fraction was identified by thin layer chromatography (TLC) and concentrated. A total of 8 fractions were obtained.

Among them, open-column chromatography was carried out by filling sephadex LH-20 for the further purification of the fraction having the most excellent antioxidant activity, and the finally obtained compound, Capsicoside G (380 mg) Separated into a single material.

Example  2: Red pepper From the degreasing pail  Isolated Fraction  Chemical structure analysis

¹ (A), ¹³ C-NMR (FIG. 1), 2D-NMR (FIG. 2) and ESI-MS 3) analysis of the fractions. As a result, caprosicoside G, which is furostanol saponin isolated from red pepper was confirmed.

Capsicoside G (CBU-6). Yield, 380 mg, ¹ H NMR (400 MHz, pyridine- d ₅ ) (aglycon moiety) δ 3.58 (1H, m, H-26b), 1.18 (3H, d, J = 6.8 Hz, (3H, d, J = 6.4 Hz, H-27), 0.78 (3H, s, H-18), 0.61 (3H, s, H-19), 0.45 (1H, m, H-9), (anomeric protons) δ5.55 (1H, J = 7.6 Hz, anomeric proton, Glc ⅳ), 5.32 (1H, J = 6.4 Hz, anomeric proton, Glc ⅱ), 5.23 (1H, J = 7.2 Hz, anomeric proton, Glc ⅲ ), 5.18 (1H, J = 8.0 Hz, anomeric proton, Glc ⅰ), 4.92 (1H, J = 7.6 Hz, anomeric proton, Gal), 4.87 (1H, J = 7.6 Hz, anomeric proton, C-26-Glc ). ESI-MS (positive ion), m / z 1429 [M + Na] +, m / z 1267 [1429 - 162] +, m / z 1105 [1297 - 162] +, and m / z 943 [1135 - 162 ] ⁺ .

Example  3: Preparation of fat precursor cells

3T3-L1 adipose precursor cells were purchased from ATCC (American Type Culture Collection, Manassas, VA, USA; ATCC No. CL-173). 3T3-L1 is widely used in experiments for studying the metabolic process of adipocytes. As the cell differentiation becomes active, the accumulation of adipocytes in the adipocytes leads to obesity. When a substance considered to have an anti-obesity effect is treated on the cell, the smaller the differentiation of the cell, the greater the anti-obesity effect.

Example  4: Capricosid  Cytotoxicity of Preganglionic Cells of G cell   cytotoxicity) and cell viability cell viability ) Measure

In order to confirm the effect of Capsicoside G obtained in Example 1 on the cell viability, the LDH (Lactate dehydrogenase) assay and MTT (3- [4,5-dimethylthiazol-2-yl] -2,5- tetrazolium bromide) assay.

Example  4-1: LDH Glass performance  Through measurement Capricosid  Cytotoxicity of Preganglionic Cells of G cell cytotoxicity ) Measure

To measure the cytotoxicity of Capsicoside G, the LDH glass activity was measured using a commercially available kit (LDH assay kit, Roche Applied Science, IN, USA). Capsicoside G samples were treated at 0, 1, 5, 10, and 20 ㎍ / mL for 24 and 48 hours, respectively. After the incubation, the supernatant was obtained to measure the amount of LDH released into the medium due to cell membrane damage, and the absorbance was measured using a microplate reader.

As a result, as shown in Fig. 4 (A), it was observed that the concentration-dependent cytotoxicity was not induced at all the concentrations tested.

Example  4-2: MTT assay Through Capricosid  Cell survival rate of adipose precursor cells of G cell viability ) Measure

To determine the effect of capecicoside G on cell viability, MTT assay was performed to measure the survival rate of early cells in the cell death process. The MTT assay is based on the principle that the dehydrogenase in the mitochondria of the cell organelles reduces MTT tetrazolium, which is a yellow water-soluble substance, into blue-violet, non-aqueous MTT formazan, By measuring the content of formazan, the cytotoxicity can be measured indirectly. Capsicoside G was treated with lipopolysaccharide at concentrations of 0, 1, 5, 10 and 20 ㎍ / mL, respectively, for 24 and 48 hours, respectively. After incubation, supernatants were collected to determine the content of formazan formed by surviving cells, transferred to 96-well plates, and reacted with MTT solution prepared at a concentration of 5 mg / mL in a cell incubator for 4 hours. After the reaction, the formed formazan was dissolved and the absorbance was measured using a microplate reader.

As a result, as shown in Fig. 4 (B), it was observed that the cell viability was not affected at all the concentrations tested.

From these results, it was confirmed that the capsicoside G of the present invention is a safe substance without cytotoxicity.

Example  5: Capricosid  G of Anti-obesity  Verification of effect

Example  5-1: Differentiation induction and sample treatment of lipid precursor cells

As the cell differentiation of lipid precursor cells becomes more active, the fat accumulation in the adipocyte becomes active and induces obesity. Therefore, in order to derive a substance having a large anti-obesity effect, in vitro In the experiment, 3T3-L1, which is a mouse adipose precursor cell, is induced to differentiate into adipocytes.

In order to induce the differentiation of 3T3-L1 mouse adipocytes into adipocytes, the adipocytes were placed in DMEM (dulbecco's modified Eagle's medium) medium containing 10% bovine calf serum (BCS) % CO ₂ . When the adipose precursor cells reach the confluent stage, they are stimulated with MDI (0.5 mM 3-isobuty1-IBMX; IBMX), 1 uM dexamethasone (DEX), 5 μg / mL Adipocyte differentiation was induced for 2 days by adding DMEM medium supplemented with 10% fetal bovine serum (FBS) supplemented with insulin. At this time, in order to confirm the inhibition of differentiation of capsicoside G obtained in Example 1 into adipocytes, Capsicoside G sample was added to the culture medium by concentration. However, from 2 days after induction of differentiation, the cells were matured into adipocytes for 4 days using a medium supplemented with insulin alone.

Example  5-2: Capricosid  G neutral fat cells ( TG , Triglyceride ) Production inhibition

In order to confirm whether the composition containing Capsicoside G as an active ingredient exhibits an anti-obesity effect, Capsicoside G was added to 3T3-L1 cells inducing lipid differentiation as in Example 5-1 (0, 1, 5, 10, 20 ㎍ / mL) for 6 days, and the degree of inhibition of adipocyte differentiation was measured by oil red o staining.

The oil red oocyte staining method is a method in which the medium is removed after the differentiation and maturation of the lipid precursor cells, and the cells are fixed using 10% formalin solution, and the cells are specifically reacted with the lipid droplets generated in the cells Lt; / RTI > solution.

As shown in Fig. 5, an oil-red stain image of 3T3-L1 cultured by treatment with each concentration of capricoside G was obtained by microscopic observation, and the stained cells after microscopic observation were dissolved in isopropanol The back absorbance was measured. As shown in FIG. 5, when the capsicoside G of the present invention was treated, the lipid accumulation capacity calculated by measuring the absorbance was compared with that of the control group not treated with the sample, It was found that the effect of inhibiting adipocyte differentiation was dose dependent.

Next, the same amount of Capsicoside G of 20 / / mL was treated at different dosing times in order to confirm the effect of inhibiting lipogenesis according to the administration time of Capsicoside G. Specifically, capsicoside G was treated at 0 to 2 days, 0 to 4 days, 0 to 6 days, 2 to 4 days, 2 to 6 days, and 4 to 6 days based on the time point of induction of the first lipid differentiation. As a result, as shown in FIG. 6, it was confirmed that the effect of suppressing fat production was greatest in the group treated with capecicose G (0 to 2 days, 0 to 4 days, 0 to 6 days) there was. That is, although the administration period was important, it was confirmed that the time point of administration was important, and when the capsicoside G of the present invention was administered at the early stage of differentiation into adipocytes, the effect of inhibiting triglyceride production was the greatest.

Taken together, it can be seen that the capsicoside G of the present invention effectively inhibits lipogenesis and is effective in preventing or treating obesity.

Example  5-3: Capricosid  Inhibitory effect of G on the expression of adipocyte differentiation protein

In order to confirm the specific mechanism of anti-obesity activity of the capsicoside G of the present invention, protein expression of PPAR-γ, C / EBP-α, aP2 and FAS, which are key factors involved in adipocyte differentiation, Respectively. That is, in addition to confirming the lipid formation inhibition of the composition observed visually in Example 5-2, the effect of the composition on the expression of the adipocyte differentiation-related protein was confirmed.

For protein expression analysis by Western blotting, cell lysis buffer was added to the cells that had been differentiated into adipocytes, left at 4 ° C for 30 minutes, and the cell solution was centrifuged to obtain a cell protein solution. Protein concentration was determined by BCA protein assay (BCA protein assay) and the same amount of protein was dissolved in 10% sodium dodecylsulfate polyacrylamide gel (SDS (sodium dodecyl sulfate) polyacrylamide gel) and electrophoresis was performed. The protein developed on the gel was transferred to a nitrocellulose membrane and then blocked with 5% skim milk for 1 hour. After blocking, the membrane was hybridized with the primary antibody of PPAR-γ, C / EBP-α, aP2 and FAS at a ratio of 1: 1000 at room temperature for 2 hours. After washing the hybridized membrane, Lt; / RTI > for a further hour. Subsequently, the membrane after the antibody reaction was treated with an enhanced chemiluminescent (ECL) detection reagent and exposed to X-ray fim for development. The degree of expression of the four proteins to be analyzed was determined by quantifying the protein band developed on the X-ray film using software image J. [

Specifically, 3T3-L1 cells that induced lipid differentiation were treated with capecicoside G at concentrations of 0, 1, 5, 10, and 20 ㎍ / mL for 6 days, PPAR-γ, C / EBP-α, aP2 and FAS. That is, as shown in FIG. 7, in the control group in which lipidation was induced by MDI treatment, four fat-forming genes (PPAR- γ, C / EBP-α, aP2 and FAS), whereas the expression of the increased genes was decreased in a dose-dependent manner by the treatment with the capsicoside G of the present invention.

Therefore, it has been confirmed that the composition containing the capsicose G of the present invention as an active ingredient inhibits the expression of a key factor involved in fat formation, thereby inhibiting the differentiation of lipid precursor cells into adipocytes, and ultimately, Respectively.

Claims (6)

A pharmaceutical composition for preventing or treating obesity comprising, as an active ingredient, Capsicoside G represented by the following formula (1).
[Chemical Formula 1]

The pharmaceutical composition according to claim 1, wherein the prevention or treatment of obesity is achieved by inhibiting the differentiation of adipose precursor cells into adipocytes.
The method according to claim 1, wherein the expression of PPAR-gamma (proliferator-activated receptor gamma), CEBP-alpha (CCAAT enhancer binding protein-beta), aP2 (adipocyte fatty acid binding protein), fatty acid synthetase Or a pharmaceutically acceptable salt thereof.
A food composition for preventing or improving obesity comprising, as an active ingredient, Capsicoside G represented by the following formula (1).
[Chemical Formula 1]

A quasi-drug composition for preventing or improving obesity, which comprises Capsicoside G represented by the following formula (1) as an active ingredient.
[Chemical Formula 1]

A cosmetic composition for preventing or improving obesity comprising, as an active ingredient, Capsicoside G represented by the following formula (1).
[Chemical Formula 1]

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