KR101594979B1 - Compositions for treating or preventing obesity containing extract or fractions of Euphorbia supina Raf. - Google Patents

Abstract

The present invention relates to an anti-obesity food composition and a composition for weight reduction comprising an extract or fraction of Euphorbia supina Raf. As an active ingredient, an anti-obesity pharmaceutical composition, . In addition to inhibiting the differentiation and lipid accumulation of adipose tissue and the fractionation of the terbinafine according to the present invention, the obesity-suppressing effect such as weight and fat tissue weight reduction in the obesity-induced group is excellent, Can be usefully used to prevent, ameliorate or treat various obesity-related diseases.

Description

Technical Field [0001] The present invention relates to a composition for preventing or treating obesity, which comprises an extract of leaves, or a fraction thereof, as an active ingredient.

The present invention relates to an anti-obesity food composition and a composition for weight reduction comprising an extract or fraction of Euphorbia supina Raf. As an active ingredient, an anti-obesity pharmaceutical composition, .

Obesity is a state of excess body fat, refers to the state of body fat is 25% (male), 30% (female) or more. In modern society, the eating habits have been improved and the nutritional intake through the food has been improved. As a result, life expectancy and quality of life have been improved. However, over consumption of food, decrease in physical exercise due to changes in means of transportation and westernization of eating habits Obesity patients are increasing rapidly every year. The World Health Organization (WHO) explains that worldwide obesity is caused by a decrease in body movements due to an increase in the frequency of intake of foods with low nutrient content, high calorie content, and changes in means of transport and lifestyle. That is, the fundamental cause of obesity is the energy imbalance in which energy accumulation occurs because the calories consumed are larger than the calories consumed. Obesity is also seen as a risk factor for adult diseases and is involved in diseases such as diabetes, arteriosclerosis, hypertension, stroke, heart failure, and various inflammatory tumors. Obesity is accompanied by a serious weight gain, which can also affect joints and bones due to excessive weight gain, leading to skeletal abnormalities. For this reason, obesity is not only an external problem, but also a serious health problem.

Prevention of obesity should be controlled through balanced diet and regular eating habits, and physical exercise should be increased to increase basal metabolic rate to ensure normal metabolism. As obesity becomes known as a problem in modern society, the word diet is attracting much attention. There are many ways to lose weight by taking advantage of the fact that people who are obese are interested in weight loss. Typical examples are fasting and fasting, enzymes, liposuction, laxatives, diuretics, and treatments. These methods provide easy and comfortable weight loss to modern people, but it is common for side effects to occur. In general, modern people are interested only in quick and easy diet, but they are not interested in fundamental therapy. Therefore, they overlook clear and scientific reasoning and the mechanism of metabolism and nutritional physiological approach with fundamental therapeutic purpose If you only focus on weight loss, you will have another problem and a yo-yo phenomenon of weight. As such, obesity can be regarded as an independent disease, but as it causes other diseases, it is becoming more popular, easy to use, easy to lose weight, less side effects such as yo-yo phenomena It is time to develop a therapeutic agent.

To date, obesity treatments for weight loss are classified as appetite suppressants, fat absorption inhibitors, and energy consumption stimulants. Among them, pharmacotherapy using an appetite suppressant such as amphetamine inducing agent is a temporary problem in which the ability to inhibit appetite is not sustained, and problems such as headache, insomnia, hypertension, tension, anxiety, hallucination symptoms, dizziness, ≪ / RTI > A representative drug is sibutramine. It acts on the hypothalamus of the brain and is known to cause an appetite suppressing effect. Serotonin and norepinephrine (Norepinephrine) inhibit the reuptake of the nervous hormones that regulate the appetite, thereby reducing appetite, preventing overeating, and inhibiting high energy intake to reduce body weight. Most of these appetite suppressants prohibit the use of these drugs in cardiovascular patients because of the adverse effects of elevated blood pressure, which can put a strain on the heart. Also, the fat absorption inhibitor is Orlistat. Orlistat is a lipase inhibitor secreted by the pancreas and digestive system. It inhibits the lipase secreted from the pancreas and digestive system, inhibiting the absorption of fat and inhibiting the accumulation of fat in the body. However, orlistat may inhibit the absorption of fat-soluble vitamins, which can lead to vitamin imbalance, gastric intussusception, gas-induced gastric bloating, and oil-bladder stools, thereby causing inconvenience to the digestive system.

Therefore, in recent years, there has been a need to develop a composition which enables not only a therapeutic agent used in an artificial method, but also a herb medicine derived from a natural product to be ingested in the form of a universal food. Particularly attention is focused on the treatment of obesity derived from natural products without side effects.

On the other hand, Euphorbia supina Raf. Is an annual herbaceous plant belonging to the Euphorbiaceae. Its main stem extends along the ground and is 10 ~ 25cm long. There are some hairs with leaves and reddish brown spots at the center. It contains white fluid. Leaves are long oval with length of 5 ~ 10mm and width of 4 ~ 6mm, with sawtooth on the edge, spread horizontally and arranged in two lines. The upper surface of the leaf is dark green, the back is rusty white, and the petiole is very short. The bedbugs are widely distributed in temperate and tropical regions of Korea, China, Japan, Southeast Asia, and North and South America. It has excellent antioxidant, antitumor, and sedative effects and is widely used for various kinds of cancer, asthma, diabetes, heart disease, kidney disease, malignant headache and mental anxiety. It has excellent hemostatic effect, It is known to be effective in treating blood and hematuria. As mentioned above, various pharmacological effects of aphids are known, but it has not yet been elucidated whether the extracts and fractions of the phytophthora infestans have anti-obesity activity.

Accordingly, the inventors of the present invention have conducted studies on extracts and fractions of the genus Bacillus thuringiensis in order to develop a new drug. As a result, the present inventors have completed the present invention by confirming that the extract or fraction of the genus Bacillus thuringiensis has an excellent anti-obesity effect.

It is an object of the present invention to provide a pharmaceutical composition for anti-obesity which comprises as an active ingredient an extract of Curcuma longa thunbergii or a fraction thereof.

Another object of the present invention is to provide an anti-obesity food composition comprising an extract of Ajiji tsukuni or a fraction thereof as an active ingredient, and a composition for weight loss comprising the extract or fraction as an active ingredient.

In order to accomplish the above object, the present invention provides a pharmaceutical composition for anti-obesity comprising an extract of Curcuma longa thunbergii or a fraction thereof as an active ingredient.

In addition, the present invention provides an anti-obesity food composition comprising as an active ingredient an extract of Ajiwa tsubaki or a fraction thereof.

In addition, the present invention provides a composition for weight loss comprising an extract of Curcuma longa larvae or a fraction thereof as an active ingredient.

In addition to inhibiting the differentiation and lipid accumulation of adipose tissue and the fractionation of the terbinafine according to the present invention, the obesity-suppressing effect such as weight and fat tissue weight reduction in the obesity-induced group is excellent, Can be usefully used to prevent, ameliorate or treat various obesity-related diseases.

BRIEF DESCRIPTION OF THE DRAWINGS FIG.
FIG. 2 is a graph showing the cell survival rate of adipocytes in the treatment of a terbinafine extract (Vehicle: normal control, EtOH: rice terbinafine extract treatment group, EGCG: positive control group).
FIG. 3 is a graph showing the cell survival rate of adipocytes in the treatment of the terrestrial broad-leaved fraction (Vehicle: normal control group, CH ₂ Cl ₂ : treatment group of dichloromethane fraction of baby root, EtOAc: ₂ O: cleavage site water bed fraction treatment group, EGCG: positive control group).
FIG. 4 is a graph showing fat accumulation rate in adipocytes in the treatment of aripipterous moth larvae extract (Vehicle: normal control, MDI: negative control, EtOH: EGCG: positive control).
FIG. 5 is a graph showing fat accumulation rate in adipocytes during the treatment of the petioles of the landing zone (Vehicle: Normal control, MDI: Negative control, CH ₂ Cl ₂ : Dipeptidyl dichloromethane fraction treatment group, EtOAc: Ethyl acetate fraction treated group, H ₂ O: teratomas water fraction treated group, EGCG: positive control group).
FIG. 6 is a graph showing the results of ELISA assay of mouse blood leptin levels in the treatment of the terrestrial broad-leaved birds (Normal: normal control, HFD: obese-induced group, EtOH: Ethyl acetate fraction treated group, H ₂ O: disrupted ground water bed fraction treatment group, GC: positive control group).
FIG. 7 is a graph showing the results of measurement of the level of adiponectin in mouse blood by ELISA in the treatment of the infected ground wing fraction.
FIG. 8 is a graph showing the results of real-time PCR measurement of PPAR-γ levels in mouse liver tissues during the treatment with the extracts or fractions of A.beta.
FIG. 9 is a graph showing the results of real-time PCR measurement of C / EBP alpha levels in mouse liver tissues in the treatment of the terbinafine extract or fraction.
FIG. 10 is a graph showing the results of real-time PCR measurement of PPAR-γ levels in mouse adipose tissues in the treatment of the terbinafine extract or fraction.
11 is a graph showing the results of real-time PCR measurement of C / EBP alpha levels in mouse adipose tissues in the treatment of the terbinafine extract or fraction.

Hereinafter, the present invention will be described in detail.

The present invention relates to the use of Euphorbia supina Shelf . ) Extract or fraction thereof as an active ingredient.

The Euphorbia of the present invention supina Shelf . ) Is a perennial herb that belongs to the Euphorbiaceae and is widely distributed in temperate and tropical regions such as Korea, China, Japan, Southeast Asia, and North and South America. It is known that the bedbugs of the baby are excellent in antioxidant, anticancer, detoxifying, soothing, and hemostatic functions.

The term "extract " in the present invention means a preparation obtained by squeezing a herbal medicine with an appropriate leaching solution and concentrating by evaporating the leaching solution. The extract is obtained by extracting the diluted solution or concentrate of the extract, the dried product obtained by drying the extract, Or purified water.

The above-mentioned baby bedbug can be used without limitation such as cultivated or marketed, and can be used as it is or can be used by drying. As the drying method, it is possible to use both blanket, shade, hot air drying and natural drying.

The above-mentioned baby bird bed bug extract or fraction can be extracted or fractionated from various parts of the baby bed bed, and is preferably extracted or fractionated from flowers, leaves, stems, roots or outposts of baby bed bugs, It is extracted or fractionated from leaves, stems or outposts of the ground worms, and most preferably extracted or fractionated from the outposts of the worm worms.

The baby cotyledon extracts or fractions can be obtained from nature using methods of extraction, separation and fractionation known in the art. The baby cotyledon extract or fraction may be obtained according to various extraction solvents and extraction methods. As an appropriate solvent for obtaining the extract or fraction from the infant bed cavity, water or an organic solvent may be used, and any pharmaceutically acceptable organic solvent may be used. Examples of the solvent include water, alcohols having 1 to 4 carbon atoms such as methanol, ethanol, propanol, isopropanol, and butanol, etc. These solvents may be used alone or as a mixture of two or more thereof . Preferably, methanol or ethanol (alcohol) can be used, and more preferably, ethanol can be used.

Preferably, the extraction temperature for obtaining the above-mentioned terbinafine extract is 50 to 100 < 0 > C. In one embodiment of the present invention, the extract of A. terbinafine was obtained at a temperature of 70 DEG C, but is not limited thereto. The extraction method may include various methods such as a hot water extraction method, a cold extraction method, a reflux cooling extraction method, a solvent extraction method, a steam distillation method, an ultrasonic extraction method, a leaching method and a pressing method, but is not limited thereto, Extraction method.

The above-mentioned pit stopper fractions can be obtained as individual solvent fractions through a process of separating an organic solvent from a non-polar to polar fraction. Suitable solvents for the fractionation of the soil bedbug fractions may be water, ethanol, methanol, hexane, chloroform, dichloromethane, ethyl acetate, butanol or mixtures thereof, preferably water, dichloromethane, ethyl acetate Most preferably water, ethyl acetate or a mixture thereof.

The baby cotyledon extracts or fractions can be obtained by freezing, heating and filtering at room temperature by conventional methods known in the art, or the solvent may be further evaporated, spray dried or lyophilized. In addition, the infant extract or fraction may be produced in powder form by an additional process such as vacuum distillation and freeze-drying or spray-drying. The extract or fraction may be further purified by further chromatography using various chromatographies such as silica gel column chromatography, thin layer chromatography, high preformance liquid chromatography and the like Can be obtained.

Therefore, the term "birdwatching" or "fragrance" used in the present invention includes all the extracts, fractions and purified products obtained in each step of extraction, fractionation or purification, their diluted solutions, concentrates or dried products.

In the present invention, the term "obesity " refers to a state having excess body fat, which means that the body fat is 25% (male) or 30% (female) or more of body weight.

The term "anti-obesity" in the present invention means treating, preventing or ameliorating an obese condition.

According to one embodiment of the present invention, the Ajiwi groundbreaking extract or fraction inhibits the differentiation of adipose precursor cells into adipocytes, reduces the amount of leptin expression in the blood, and increases the expression level of adiponectin. In addition, the expression level of PPAR-γ protein, C / EBP α protein, C / EBP β protein and C / EBP δ protein, which are transcription factors involved in adipocyte differentiation, . Therefore, the extracts or fractions of the infant of the present invention may be useful as medicines and food compositions for preventing or treating obesity-related diseases or complications due to obesity.

The inventive terbinafine extract or fractions include, but are not limited to, metabolic syndrome, hypertriglyceridemia, hyperlipidemia, hypolipidemia, angina pectoris, myocardial infarction, sexual dysfunction, sleep apnea syndrome, premenstrual syndrome, Hyperlipidemia, diabetes mellitus, hypertension, impaired glucose tolerance, coronary artery thrombosis, diaphoresis, depression, anxiety, hyperlipidemia, hypertension, diabetes mellitus, , Anorexia nervosa, polycystic ovarian disease, atherosclerosis, atherosclerosis, atherosclerosis, gallbladder disease such as atherosclerosis, gallstone disease, anorexia nervosa, anorexia nervosa, Dermatological conditions such as reproductive disorders, infections, varicose veins, epidermal hyperplasia and eczema, insulin resistance, chronic arteriolar obstruction, orthopedic surgery May be used to prevent or treat injury, thromboembolic events, cardiovascular disease, urinary disorders, lipid syndromes, hyperglycemia, and stress at least one obesity-related disorder selected from the group consisting of.

The present invention also provides a method for preventing, ameliorating or treating obesity, abdominal obesity or visceral fat accumulation in each part of the body, and preferably for preventing, improving or treating visceral fat accumulation Can be used. The abdominal obesity or visceral fat accumulation includes, but is not limited to, living in a chair for a long time, or caused by lack of exercise, alcohol consumption, stress or the like.

Preferably, the pomegranate extract or fraction is contained in an amount of 0.01 to 95% by weight, more preferably 1 to 80% by weight, based on the total weight of the pharmaceutical composition of the present invention.

The pharmaceutical compositions of the present invention may further comprise suitable carriers, excipients and diluents conventionally used in the art. In addition, it can be formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, oral preparations such as syrups and aerosols, external preparations, suppositories and sterilized injection solutions according to a conventional method. Suitable formulations known in the art are preferably, but not limited to, those disclosed in Remington's Pharmaceutical Science (Mack Publishing Company, Easton PA). Examples of the carrier, excipient and diluent include lactose, dextrose, sucrose, sorbitol, mannitol, xylitone, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, Cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, mineral oil and the like. When the pharmaceutical composition is formulated or formulated, 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, lactose, Gelatin and the like. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used.

Examples of the liquid preparation for oral use include suspensions, solutions, emulsions, and syrups. In addition to water and liquid paraffin, simple diluents commonly used, various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included . Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, suppositories, and the like. 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. The components may be added to the active ingredient, i. E., The pomegranate extract or fraction, either independently or in combination.

The term "administering" as used herein means providing a pharmaceutical composition of the present invention to a subject in any suitable manner.

The present invention also provides pharmaceutical compositions comprising an amount of an active ingredient or pharmaceutical composition that induces a biological or medical response in a tissue system, animal or human, as contemplated by a researcher, veterinarian, physician or other clinician, Or a therapeutically effective amount which is sufficient to induce relief. It will be apparent to those skilled in the art that the therapeutically effective dose and the frequency of administration for the pharmaceutical compositions of the present invention will vary with the desired effect. Thus, the optimal dosage to be administered can be readily determined by those skilled in the art and will vary with the nature of the disease, the severity of the disease, the amount of active and other ingredients contained in the composition, the type of formulation, and the age, The age, body weight, sex, diet, time of administration, route of administration and fraction of the composition, duration of treatment, concurrent medication, and the like. The pharmaceutical composition of the present invention may be administered to the individual by various routes. But are not limited to, intravenous, intraperitoneal, intramuscular, intraarterial, oral, intracardiac, intramedullary, intradermal, transdermal, intestinal, subcutaneous, sublingual or topical administration.

The pharmaceutical composition of the present invention may be administered at a dose of 1 to 10,000 mg / kg / day, preferably 1 to 2000 mg / kg / day, and may be administered once a day or divided into several doses.

The pharmaceutical composition of the present invention may be administered to a subject in various routes. All modes of administration may be expected, for example, oral, rectal or intravenous, intramuscular, subcutaneous.

In addition, the pharmaceutical composition of the present invention can be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy or biological response modifiers for the prevention or treatment of obesity, obesity-related diseases or complications .

In addition, the present invention provides an anti-obesity food composition comprising as an active ingredient an extract of Ajiwa tsubaki or a fraction thereof.

The inventive terbinafine extract or fraction can be used as a health functional food, a food additive or a dietary supplement.

In the case of using the baby food extract or fraction of the present invention as a food additive, it may be suitably used according to a conventional method such as adding the baby food extract or fraction as it is, or mixing it with other food or food ingredients .

In addition, the amount of mixture of the above-mentioned active ingredients, the soil bed bug extract or the fractions, may be suitably changed according to the purpose of use (prevention, health or therapeutic treatment), and the above- By weight, preferably 0.01 to 95% by weight, more preferably 1 to 80% by weight. As a specific example, in the production of a food or beverage, the rice terra cotta extract or fraction of the present invention is added in an amount of 15% by weight or less, preferably 10% by weight or less based on the raw material.

There is no particular limitation on the type of the food, but examples of the food to which the present invention is added include a meat, a sausage, a bread, a chocolate, a candy, a snack, a confectionery, a pizza, a ramen, Dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages, vitamin complexes, and the like.

When the food composition of the present invention is prepared as a beverage, it may contain additional ingredients such as various flavors or natural carbohydrates such as ordinary beverages. Examples of the natural by-products include monosaccharides such as glucose and fructose; Disaccharides such as maltose and sucrose; Natural sweeteners such as dextrin and cyclodextrin, synthetic sweeteners such as saccharin and aspartame, and the like. The natural carbohydrate is contained in an amount of 0.01 to 10% by weight, preferably 0.01 to 0.1% by weight based on the total weight of the food composition of the present invention.

The food composition of the present invention can be used for various foods, vitamins, electrolytes, flavors, colorants, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloid thickening agents, pH adjusters, stabilizers, preservatives, glycerin, Carbonation agents used in beverages, etc., and may include, but is not limited to, natural fruit juices, fruit juice beverages, and flesh for the manufacture of vegetable beverages. These components may be used independently or in combination. The proportion of the above additives is not particularly limited, but is preferably within the range of 0.01 to 0.1% by weight based on the total weight of the food composition of the present invention.

For long-term consumption intended for health and hygiene purposes or health control purposes, the food composition of the present invention has no problem in terms of safety and can be taken for a long period of time.

In addition, the present invention provides a composition for weight loss comprising an extract of Curcuma longa larvae or a fraction thereof as an active ingredient.

The composition for weight reduction according to the present invention may be a pharmaceutical composition or a food composition. When formulated into a pharmaceutical composition, various auxiliaries used in medicine such as carriers and other additives such as stabilizers, emollients, emulsifiers and the like may be added as needed unless adversely affecting the active ingredient. The formulations may be any formulations suitable for pharmaceutical preparations of herbal medicine, including tablets, pills, capsules, granules, powders, excipients, precursors, infiltrates and the like.

As used herein, the term "prophylactic " means any act that inhibits obesity or delays progression by administration of the composition of the present invention.

As used herein, the term "treatment" refers to any action in which the administration of the composition of the present invention improves or alleviates obesity.

Terms not otherwise defined herein have meanings as commonly used in the art to which the present invention belongs.

Hereinafter, the present invention will be described in detail with reference to Examples and Preparation Examples. However, the following examples and preparative examples are merely illustrative of the present invention, and the content of the present invention is not limited to the following examples and preparative examples.

Example  One. Baby bedbug ( Euphorbia supina Shelf .) Preparation of extract

2.5 kg of Euphorbia supina Raf. Plant which was dried in the air was repeatedly extracted with 70% ethanol at 70 ° C three times using a reflux extracting apparatus to obtain 250 g (yield: 10.06%) of a baby grasshopper extract. For example.

Example  2. Baby bedbug Fraction  Produce

The procedure of suspending 250 g of the A. niger extract obtained in Example 1 in distilled water and separating the dichloromethane (CH ₂ Cl ₂ ) layer was repeated three times. The dichloromethane layer thus obtained was combined and concentrated under reduced pressure to obtain 128 g (yield: 51.2%) of a dichloromethane fraction in the form of a flower bed. Ethyl acetate (CH ₃ COOC ₂ H ₅ ) was added to the remaining water layer and fractionation was carried out in the same manner as above. The filtrate was concentrated under reduced pressure and lyophilized to obtain 48 g (yield: 19.2%) of the ethyl acetate fraction 74 g (yield: 29.6%) of the fractions were obtained and used in the following examples.

The production process of the baby cotto leaves extract and fractions is briefly shown in Fig.

Example  3. Cytotoxicity and viability analysis

3-1. Lipo-precursor cells (3 T3 - L1 ) And preparation

3T3-L1 cells are lipogenic precursor cells widely used to study the metabolism of adipocytes. As 3T3-L1 cells are actively differentiated, fat accumulation in adipocytes is also actively induced. 3T3-L1 cells were purchased from ATCC (American Type Culture Collection, USA). Cell lines obtained from ATCC were cultured in DMEM medium containing 10% bovine calf serum (BCS) and 1% penicillin in an incubator maintained at 37 ° C with 5% CO ₂ . When the cells were stabilized through two or more subcultures, they were used for the experiment.

3-2. Cytotoxicity and viability analysis

(MTT (3- (4,5-dimethylthiazol-2-yl) -1,2,3,4-tetrahydronaphthalene-2-yl) -benzonitrile was used in order to determine the cytotoxicity, -2,5-diphenyl tetrazolium bromide) analysis was performed. 3T3-L1 cells were inoculated into a 48-well plate at a concentration of 2 × 10 ⁵ cells / well, and the culture medium was changed once every two days and cultured to 100% confluent. The cultured cells were treated with 4, 20, and 100 μg / ml of each of the pituitary extracts, respectively. The dichloromethane fraction, the ethyl acetate fraction, and the pituitary extract fraction were 0.8, 4, 20 ㎍ / ㎖, and cultured at 37 ° C and 5% CO ₂ for 8 days. After incubation, absorbance was measured using MTT buffer. (EGCG) treated with Epigallocatechin gallate (EGCG) was used as a positive control group, and the above procedure was repeated three times as a normal control group Lt; / RTI > Cell viability was calculated by the following formula 1, and the results are shown in FIG. 2 and FIG.

[ Equation  One]

Cell viability (%) = (absorbance of sample-treated group / absorbance of control group) x 100

As shown in FIG. 2 and FIG. 3, it was confirmed that the active ingredient of the present invention was not toxic at high concentration.

Example  4. Baby bedbug  Extract or Fraction  Identification of inhibitory effect on adipocyte differentiation

The fat accumulation rate was checked to determine if the extracts or fractions of the terrestrial bandlets inhibit the differentiation of 3T3-L1 cells into adipocytes. To confirm this, stabilized 3T3-L1 cells were plated in a 48-well plate at a density of 2 × 10 ⁵ cells / well and cultured. The cells were maintained for 2 days at 100% confluent time. 3T3-L1 cells were treated with 10% Fetal Bovine Serum (FBS) containing 1% MDI (0.5 mM 3-isobutyl-1-methylxanthine (IBMX), 1 μM dexamethasone, Penicillin-containing DMEM medium for induction of differentiation into adipocytes for 2 days. After 48 hours of culture, the cells were cultured in DMEM medium containing 10% FBS and 1% penicillin containing 1 μg / mL of insulin for 2 days. During the adipocyte differentiation process, the extracts of the rice seedlings and the leaves of the rice seedlings were treated at the concentrations of 4, 20 and 100 μg / mL, and the dichloromethane fraction, the ethyl acetate fraction, Water fractions were treated at 0.8, 4, and 20 μg / mL in culture. Then, the degree of adipocyte differentiation was observed on the 8th day when the differentiation was completed. As a normal control group, vehicle treated group (Vehicle) was used. Negative control group was treated with IBMX, Insulin, Dexamethasone to induce lipid accumulation (MDI).

In order to observe the degree of differentiation of adipocytes, Oil Red O staining, which is specific to lipid droplet, was performed. Specifically, the medium was removed from adipocyte differentiation-induced cells and fixed with 10% formalin for 1 hour at room temperature. Then washed with 60% isopropanol and each well was thoroughly dried. Oil Red O dye (60% Oil Red O stock, 40% sterilized distilled water) was treated for 1 hour and washed twice with 60% isopropanol and once with sterile distilled water. For the elution of the combined Oil Red O, it was treated with 100% isopropanol for 10 minutes, and the absorbance of the eluate was measured at 520 nm.

Fat accumulation rate in adipocyte was calculated by the following formula 2, and the results are shown in FIG. 4 and FIG.

[ Equation  2]

Fat accumulation rate (%) = (absorbance of sample-treated group / absorbance of control group) x 100

As shown in FIG. 4 and FIG. 5, it was confirmed that fat accumulation in adipocytes was inhibited in a concentration-dependent manner by the treatment with the active ingredient of the present invention, the soil bed band extract, the ethyl acetate fraction, the dichloromethane fraction and the water fraction. Therefore, it can be seen that the pituitary gland extract or the pit bark of the present invention inhibits the differentiation of adipose precursor cells into adipocytes, and thus the prevention or treatment effect of obesity is excellent.

Example  5. Baby bedbug  Extract or Fraction  Check your weight loss effect

5-1. Construction of Obesity-Induced Mouse Model

5 weeks old C57BL / 6J male mice (20 ~ 25g) were purchased from Samtako (Osan, Korea) and were allowed to freely eat solid feed and water for 1 week. ± 10%), constant temperature (22 ± 2 ℃), and 12-hour cycle. After the adaptation period for one week, the animals were divided into 12 groups. The normal control group (normal), obesity induction group treated with 45% high fat diet (HFD) (0.8, 4, 20 mg / kg, ESEA + HFD) and the control group (0.8, 4, 20 mg / kg, ESEE + HFD) prepared in Example 2, 4, 20 mg / kg, ESH ₂ O + HFD) and the positive control group, Garminia cambogia extract treatment (200 mg / kg, GC + HFD).

Obesity group, obesity group, obesity group, obesity group, obesity group, obesity group, obesity group, obesity group, obesity group, After dissolving in cellulose (sterilized distilled water + carboxymethylcellulose), 45% high fat diet was freely taken and oral administration of extract or fraction was performed once a day for 6 weeks. Normal control diet was given freely, and 0.9% of saline was orally administered to the obese group. A mouse model was constructed as described above and used in the following examples.

5-2. Check your weight change

In order to confirm the anti-obesity effect of the extracts or fractions of the birds, the body weight was measured at intervals of one week after the start of the experiment and the start of the experiment in the mouse experimental group constructed by the method of Example 5-1, Table 3 shows the results.

As shown in Tables 1 to 3, it was confirmed that the body weight of the obese-induced group was significantly increased compared with the normal control group. In contrast, the weight gain of the group treated with Garcinia cambogia was 34.13% lower than that of the obese group.

Specifically, as shown in Table 1, the body weight gain was decreased by 13.52% and 34.54%, respectively, compared with the obesity-induced group, and as shown in Table 2, the ethyl acetate fraction fraction Were decreased by 23.02% and 52.27%, respectively, in comparison with the obese group. In addition, as shown in Table 3, it was confirmed that the body weight gain was reduced by 20.21% and 52.28% in the group treated with the parasitic infestation water, compared with the obesity-induced group.

Therefore, the extract of the present invention, which is the active ingredient of the present invention, decreased the body weight to a significant level, and the weight loss was significantly larger in the 20 mg / kg-treated group than the control group . In addition, 20mg / kg - treated group showed higher weight loss than the positive control group. From these results, it was confirmed that the extracts or fractions of the present invention were excellent in anti-obesity effect by inducing weight loss.

Example  6 - Baby bedbug  Extract or Fraction  Long-term weight and fat tissue weight reduction effects

In order to confirm the anti-obesity effect of the A. niger extract or fraction of the present invention, the weight of liver tissue, kidney tissue, white epididymal fat tissue and spleen tissue of the experimental group constructed in 5-1 above was measured. Specifically, after the experiment period of 5-1, the mouse model was sacrificed by ether anesthesia, the abdomen was excised, each tissue was extracted, and blood and foreign substances were removed using physiological saline and the weight was measured. Are shown in Tables 4 to 6 below.

As shown in Tables 4 to 6, the weight of white epididymal adipose tissue was significantly increased in the obese-induced group compared to the normal control group, but the weight of the treated group was significantly decreased in the group treated with Garcinia cambogia . In addition, it was confirmed that the weight of white epididymal adipose tissue was significantly decreased in the concentration -

Accordingly, the present invention relates to a method for producing an anti-obesity food composition, an anti-obesity food composition, or a composition for weight loss, which is effective for decreasing the weight of adipose tissue.

Example  7 - Measurement of triglyceride concentration in serum

In order to confirm the anti-obesity effect of the inventive terbinafine extract, the concentration of triglyceride in the blood of the mouse model was measured after completion of the experiment of the above example.

Blood was collected after the experiment was terminated and cardiac blood was drawn after anesthesia. The collected blood was allowed to stand at room temperature for 30 minutes and then centrifuged at 4 ° C for 3 minutes at 3000 rpm for 10 minutes to obtain serum (triglyceride ), Total cholesterol, high density lipoprotein-cholesterol and low density lipoprotein-cholesterol concentrations were measured using Asanpharm (Korea) kit, Table 9 shows the results.

As shown in Tables 7 to 9, the concentration of triglyceride, total cholesterol, and LDL-cholesterol in the serum of the obese-induced group was higher than that of the normal control group, and the concentration of HDL-cholesterol was found to be low. On the other hand, the concentration of triglyceride, total cholesterol and LDL-cholesterol in serum was lower and the concentration of HDL-cholesterol was higher in obesity-induced group than that in obesity-induced group Respectively.

Therefore, the inventive terbinafine extract or fraction thereof significantly reduces the concentration of triglyceride, total cholesterol and LDL-cholesterol in the test animal in a concentration-dependent manner and significantly increases the concentration of HDL-cholesterol in a concentration-dependent manner, And the obesity effect is excellent.

Example  8 - Baby bedbug  Extract or Fraction  Serum Leptin  And Depotectin  Check the effect on concentration

The leptin and adiponectin concentrations in the blood of animal models were measured after completion of the experiment of the above example in order to confirm the anti-obesity effect of the infant extracts or fractions. Leptin and adiponectin are both secreted proteins in adipocytes.

As in Example 7, blood collected from a mouse was centrifuged to obtain serum, and concentrations of leptin and adiponectin were measured using an ELISA kit. The results are shown in FIGS. 6 and 7.

As shown in FIG. 6, serum leptin levels were significantly increased in the obese-induced group compared to the normal control group, and the positive control group was significantly decreased as compared with the obese-induced group. In addition, it was confirmed that the concentration of leptin in the serum was decreased in a concentration-dependent manner in the group of administration of the active ingredient of the present invention, that is, the terbinafine extract or fraction, in comparison with the obese-induced group.

As shown in FIG. 7, serum concentrations of adiponectin were significantly lower in the obese-induced group than in the normal control group, and the group treated with Garcinia cambogia extract, which was a positive control, was significantly increased as compared with the obese-induced group. In addition, it was confirmed that the concentration of adiponectin in the serum was significantly increased in the group of administration of the active ingredient of the present invention as a control group, or in the fraction-administered group, as compared with the obesity-induced group.

Thus, the baby bedbug extract or fraction may have an anti-obesity effect due to the effect of decreasing the leptin level of blood and the effect of increasing the level of adiponectin.

Example  9 - Baby bedbug  Extract or Fraction  Confirming the inhibitory effect of obesity gene expression

In order to confirm whether or not the extract of the present invention is inhibiting obesity-associated genes, RNA was extracted from liver tissue and white epididymal adipose tissue, and expression of PPAR-γ, C / EBP α gene was analyzed by Real-time PCR Respectively.

Specifically, first, a lysis buffer was added to the liver tissue and white epididymal adipose tissue of the mice to homogenize them, and then a supernatant was obtained using a centrifuge. In the supernatant CHCl ₃ After adding the solution, the supernatant was obtained again by using a centrifuge. Isopropanol was added to the supernatant thus obtained, and the supernatant was removed using a centrifugal separator. After washing with 75% ethanol, a precipitate RNA was obtained. This precipitate was dissolved in water without Rnase and used. RNA was synthesized by cDNA synthesis kit and real-time PCR was performed.

Real-time PCR was performed using the StepOnePlus ^TM Instrument was used. First, add 10 μl of SYBR® Green Reagents, 0.4 μl of forward primer (10 pmol / l), 0.4 μl of reverse primer (10 pmol / μl), 7.2 μl of water without Rnase and 2 μl of template (30 ng / μl) -time PCR was performed on a plate and then proceeded using a StepOne Softewear 2.3v. The PCR conditions were 95 cycles of 10 minutes, 95 cycles of 10 seconds, and 60 cycles of 1 minute at 40 cycles. The results are shown in FIGS.

As shown in FIG. 8 to FIG. 11, it was confirmed that the extracts or fractions of A. terbinafine according to the present invention inhibited the expression of PPAR-γ and C / EBP α in liver and white epididymal adipose tissue in a concentration-dependent manner.

Therefore, it can be seen that the extract of Ajiwa tendonhopper of the present invention inhibits the expression of obesity factors, and thus the effect of preventing or treating obesity, obesity-related diseases or complications will be excellent.

Formulation example  1 - Manufacture of pharmaceutical preparations

Acid production

A mixture of 20 mg of the infant groundbug extract or fraction, 100 mg of lactose, and 10 mg of talt was mixed and filled into airtight bags to prepare powders.

Tablet manufacture

The tablets were prepared by mixing 10 mg of cotyledonberry extract or fraction, 100 mg of corn starch, 100 mg of lactose and 2 mg of magnesium stearate and then tableting according to a conventional preparation method.

Capsule preparation

According to a conventional capsule preparation method, 10 mg of the extract or fraction of Aji, 9 mg of crystalline cellulose, 14.8 mg of lactose and 0.2 mg of magnesium stearate were mixed and filled in gelatin capsules to prepare capsules.

Injection manufacturing

1 ampoule per (2mL) according to the conventional method for preparing injections were prepared in a baby bed bugs land extract or fraction 10mg, mannitol 180mg, injectable sterile distilled water 2,974mg and _{Na 2 HPO 4 · 2H 2 O} 26mg.

Liquid preparation

In accordance with the usual preparation method of the liquid preparation, 20 mg of the extracts or fractions of Piggerychus, 10 g of isomerized sugar and 5 g of mannitol were dissolved in purified water, and the lemon flavor was added in an appropriate amount. Then, purified water was further added thereto, adjusted to a total volume of 100 mL, filled in a brown bottle, and sterilized to prepare a liquid preparation.

Formulation example  2 - Manufacture of food preparation

Health food manufacturing

100 mg of vitamin A mixture, vitamin A acetate 70 g, vitamin E 1.0 mg, vitamin B1 0.13 mg, vitamin B2 0.15 mg, vitamin B6 0.5 mg, vitamin B12 0.2 g, vitamin C 10 mg, biotin 10 g, nicotinic acid Amide 1.7 mg, folic acid 50 g, calcium pantothenate 0.5 mg, an appropriate amount of inorganic mixture, ferrous sulfate 1.75 mg, zinc oxide 0.82 mg, magnesium carbonate 25.3 mg, potassium phosphate monobasic 15 mg, calcium phosphate dibasic 55 mg, potassium citrate 90 mg, 100 mg of calcium and 24.8 mg of magnesium chloride were mixed and granules were prepared and a health food was prepared according to a conventional method. At this time, although the composition ratio of the vitamin and mineral mixture is relatively mixed with the ingredient suitable for health food, it may be arbitrarily modified.

Health drink manufacturing

100 g of vitamin C, 100 g of vitamin E (powder), 19.75 g of iron lactate, 3.5 g of zinc oxide, 3.5 g of nicotinic acid amide, 0.2 g of vitamin A, 0.2 g of vitamin B1 0.25 g, 0.3 g of vitamin B2 and a predetermined amount of water were mixed and heated at 85 DEG C for about 1 hour with stirring, and the resulting solution was filtered to obtain a sterilized 2L container, sealed sterilized, and refrigerated to prepare a health drink . At this time, although the composition ratio of the ingredients suitable for the beverage is comparatively mixed, the mixture ratio may be arbitrarily varied according to the demand, the demanded country, the intended use, and the regional or national preference.

Although the present invention has been described in terms of the preferred embodiments mentioned above, it is possible to make various modifications and variations without departing from the spirit and scope of the invention. It is also to be understood that the appended claims are intended to cover such modifications and changes as fall within the scope of the invention.

Claims (12)

A pharmaceutical composition for anti-obesity comprising an ethanol extract of Euphorbia supina Raf . Or a fraction thereof as an active ingredient.
The pharmaceutical composition for anti-obesity according to claim 1, wherein the ethanol is 70% ethanol.
delete The pharmaceutical composition according to claim 1, wherein the fraction is a fraction of at least one solvent selected from the group consisting of water, dichloromethane, ethyl acetate, and mixtures thereof.
2. The pharmaceutical composition according to claim 1, wherein the ethanol extract or fraction thereof of the petioles of the present invention inhibits differentiation of adipocytes.
The pharmaceutical composition for an anti-obesity drug according to claim 1, wherein the ethanol extract or the fraction thereof of the terrestrial herbaceous plant decreases the expression level of leptin in the blood or increases the expression level of adiponectin .
[Claim 2] The method according to claim 1, wherein the ethanol extract or fraction thereof of the baby food comprises at least one expression amount selected from the group consisting of PPAR-gamma protein, C / EBP alpha protein, C / EBP beta protein and C / EBP delta protein Lt; RTI ID = 0.0 > anti-obesity < / RTI >
The method according to claim 1, wherein the ethanol extract or fraction thereof of the baby's footbath comprises at least one kind of obesity-related disease selected from the group consisting of metabolic syndrome, hypertriglyceridemia, hyperlipidemia, hypolipidemia and insulin resistance Or a pharmaceutically acceptable salt thereof.
2. The pharmaceutical composition according to claim 1, wherein the ethanol extract or the fraction thereof of the baby's footbath is to prevent or treat visceral fat accumulation.
A food composition for anti-obesity comprising an ethanol extract or a fraction thereof as an active ingredient.
11. The food composition according to claim 10, wherein the food composition is a health functional food, a food additive or a dietary supplement.
A composition for weight loss comprising an ethanol extract or a fraction thereof as an active ingredient.

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