KR101710780B1 - Pharmaceutical composition for preventing or treating obesity or lipid related metabolic disease comprising extract of Atractylodis macrocephalae rhizoma - Google Patents

Abstract

The present invention relates to a pharmaceutical composition and a food composition for the treatment of obesity or lipid-related metabolic diseases comprising Atractylodis macrocephala rhizoma extract or Atractylenolide III as an active ingredient. In the present invention, it was confirmed that obesity, glucose consumption and energy metabolism were regulated by controlling the fat metabolism, glucose metabolism and insulin resistance in the fat and muscle cells using the above extract or compound, In the prevention or treatment of type 2 diabetes according to resistance, it is expected to be more fundamentally approachable and effectively treated.

Description

TECHNICAL FIELD The present invention relates to a pharmaceutical composition for preventing or treating obesity or a lipid-related metabolic disease, which comprises at least one extract obtained from Atractylodis macrocephalae rhizoma,

The present invention relates to a pharmaceutical composition and a food composition for the treatment of obesity or lipid-related metabolic diseases comprising Atractylodis macrocephala rhizoma extract or Atractylenolide III as an active ingredient.

Diabetes (diabetes) is a disease in which glucose present in the blood is excreted through the urine. It is one of the most non-communicable chronic diseases among modern people and a chronic intractable disease that does not fundamentally heal. This is a metabolic disorder characterized by hyperglycemia caused by insulin action and insulin secretion. It is a type 1 diabetes that insulin is absolutely lacking due to destruction of pancreatic beta cells by autoimmune mechanism and resistance to insulin action (insulin resistance ) And type 2 diabetes mellitus (diabetes mellitus). In particular, type 2 diabetes mellitus accounts for 90-95% of all diabetes mellitus. These diabetes mellitus causes hyperglycemia to be chronic and triggers lipid and protein metabolism disorders as well as carbohydrate metabolism, leading to vascular complications such as retinas, kidneys, nerves, and cardiovascular diseases and abdominal lesions. Various complications .

Currently, the prevalence of diabetes is 8.2%, with about 3 million people suffering from diabetes. Moreover, the death rate due to diabetes is also rapidly increasing. In 2004, 24.3 deaths per 100,000 population were the cause of death due to diabetes. In particular, recent dietary changes and various stresses have increased the number of obese patients. Thus, the excessively accumulated triglycerides are stored not only in adipose tissue but also in liver and muscle, inducing insulin resistance, causing metabolic diseases including diabetes As a result, the number of diabetic patients in Korea is increasing year by year, and it is emerging as a social problem.

Therefore, there is a growing interest in the development of new therapeutic agents to improve the quality of life suffered from metabolic diseases including diabetes. Metformin, a type of biguanide, Thiazolidinedione, Pioglitazone and Rosiglitazone, which activates PPAR-γ, α-glucosidase inhibitor, which prevents the rapid increase of postprandial blood glucose level by delaying carbohydrate absorption in the small intestine, and pancreatic Sulfonylureas and Non-sulfonylureas are used to promote insulin secretion in β-cells. Metformin, which is the most widely used in the world, lowers blood glucose level but does not induce hypoglycemia, promote insulin secretion, and has side effects of digestive system. The Thiazolidinedione family reduces insulin resistance, increases insulin-induced blood glucose consumption, increases blood glucose levels in muscle and fat, and regulates blood glucose levels in the liver. However, liver toxicity, weight gain, and edema occur. α-Glucosidase inhibitors delay digestion of carbohydrate digestion to prevent rapid increases in postprandial blood glucose and insulin levels, but adverse gastrointestinal effects occur.

Accordingly, development of a therapeutic agent for diabetes mellitus with improved side effects of such conventional therapeutic agents has been the subject of major researches, and studies have been conducted (Korean Patent Laid-open No. 10-2015-0027648).

Disclosure of the Invention The present invention has been conceived to solve the above-mentioned problems. The present inventors have found that the extract of Artemisia or Artactylenolide III, which is an active ingredient thereof, can control the differentiation of adipose tissue, decrease fat accumulation and promote energy metabolism or sugar consumption The effect of improving insulin resistance has been confirmed in detail and the present invention has been completed on the basis thereof.

Accordingly, an object of the present invention is to provide a pharmaceutical composition / food composition for improving or treating obesity or lipid-related metabolic diseases, which comprises extract of Atractylodis macrocephala rhizoma as an active ingredient.

Another object of the present invention is to provide a pharmaceutical composition / remedy composition for preventing or treating obesity or lipid-related metabolic diseases, which comprises atractylenolide (III) or a pharmaceutically acceptable salt thereof as an active ingredient .

However, the technical problem to be solved by the present invention is not limited to the above-mentioned problems, and other matters not mentioned can be clearly understood by those skilled in the art from the following description.

In order to achieve the object of the present invention, the present invention provides a pharmaceutical composition / improvement food composition for preventing or treating obesity or lipid-related metabolic diseases comprising extract (Atractylodis macrocephala rhizoma) as an active ingredient .

The present invention provides a pharmaceutical composition / improvement food composition for preventing or treating obesity or lipid-related metabolic diseases comprising atractylenolide (III) or a pharmaceutically acceptable salt thereof as an active ingredient.

In one embodiment of the present invention, the atactylenolide III may be separated from the extract.

In another embodiment of the present invention, the metabolic disease may be any one or more selected from the group consisting of insulin resistance syndrome, fatty liver, type 2 diabetes, hyperlipidemia, cardiovascular disease, and arteriosclerosis.

The present invention provides a method for treating obesity or lipid-related metabolic diseases comprising administering the above pharmaceutical composition to a subject.

The present invention provides a therapeutic use of a composition comprising atractylodis macrocephala rhizoma extract or atractylenolide Ⅲ for obesity or lipid-related metabolic diseases.

The composition according to the present invention comprises atractylodis macrocephala rhizoma extract or atractylenolide Ⅲ as an active ingredient and has an effect of improving the insulin resistance and controlling glucose metabolism of the extract or compound in fat and muscle cells It is expected that the composition can be useful as a pharmaceutical composition for preventing or treating obesity or obesity type 2 diabetes.

FIG. 1 shows the results of RT-PCR for the increased expression of differentiation and adipogenic transcription factors in the adipocyte by atractylenolide Ⅲ (A) treatment with extracts of the extracts (ARA) and its active ingredient, atractylenolide Ⅲ (A).
FIG. 2 shows the result of Western blot analysis of the increase in PPAR-γ expression and the increase of phosphorylation of AMPK in adipocytes by atractylenolide Ⅲ treatment, which is an active ingredient of extract (ARA).
FIG. 3 shows the result of RT-PCR analysis of increase in glucose uptake and mitochondrial energy metabolism-related gene expression in muscle cells by extractive extract (ARA) treatment.
FIG. 4 shows Western blot analysis of glucose uptake and mitochondrial energy metabolism-related protein expression in muscular cells treated with atractylenolide III (A), an extract of Paeonia sinensis extract (ARA) and its active ingredient.
FIG. 5 shows the results of the reduction of the accumulation of fatty acid in the muscle cells by the extract of atractylenolide III (A) and its active ingredient (ARA) ( ^* P <0.05, ^** P <0.01 vs control (a) or metfromin-treated group (b) Data present mean SD of three independent experiments Met, metformin 2.5 mM-treated cells, A, atractylenolide III 100 μM-treated cells.

The inventors of the present invention have found that atractylodis macrocephala rhizoma extract or atractylenolide Ⅲ which is an active ingredient thereof can be used to control the differentiation of adipose tissue, inhibit fat accumulation, and improve insulin resistance by promoting energy metabolism or sugar consumption And the present invention has been completed on the basis thereof.

Hereinafter, the present invention will be described in detail.

The present invention relates to a pharmaceutical composition for preventing or treating obesity or lipid-related metabolic diseases, which comprises atractylodis macrocephala rhizoma extract or atractylenolide III or a pharmaceutically acceptable salt thereof as an active ingredient to provide.

Particularly, since the present invention has an effect of controlling the increase of blood glucose and the increase of insulin resistance due to obesity or lipid-related metabolic diseases, Type 2 diabetes and its complications can be prevented or treated.

In the present invention, the extract can be extracted using a conventional solvent known in the art for extracting an extract from a natural product, that is, under ordinary temperature and pressure conditions. For example, in the present invention, the extract can be extracted using at least one solvent selected from the group consisting of water, an alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof, preferably water. Water refers to water suitable for drinking standards on the basis of the Ministry of Health. Generally, water such as purified water, sterilized water, anion water, tap water and mineral water filtered with a filter or a water purifier can be used. The method for extracting the extract can be carried out by various methods such as hot water extraction, cold extraction, reflux extraction, ultrasonic extraction and the like, but is not limited thereto.

The extract thus prepared may be filtered, concentrated or dried to remove the solvent, and may be subjected to both filtration, concentration and drying. For example, the filtration can be performed using a filter paper or a vacuum filter, the concentration can be carried out using a vacuum concentrator, and the lyophilization can be carried out, but the present invention is not limited thereto.

Further, the extract extracted with the solvent may be further fractionated with a solvent selected from the group consisting of hexane, methylene chloride, acetone, ethyl acetate, ethyl ether, chloroform, water, and mixtures thereof. The fractionation temperature may be 4 占 폚 to 120 占 폚, but is not limited thereto.

In the present invention, Atractylenolide III represented by the following general formula (1) may be isolated from the extract, but is not limited thereto.

As salts, acid addition salts formed by pharmaceutically acceptable free acids are useful. Acid addition salts include those derived from inorganic acids such as hydrochloric acid, nitric acid, phosphoric acid, sulfuric acid, hydrobromic acid, hydroiodic acid, nitrous acid or phosphorous acid, and aliphatic mono- and dicarboxylates, phenyl-substituted alkanoates, hydroxyalkanoates, Dioleate, aromatic acid, aliphatic and aromatic sulfonic acids. Such pharmaceutically innocuous salts include, but are not limited to, sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, nitrate, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate chloride, bromide, Butyrate, caprate, heptanoate, propiolate, oxalate, malonate, succinate, succinate, maleic anhydride, maleic anhydride, , Sebacate, fumarate, maleate, butyne-1,4-dioate, hexane-1,6-dioate, benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, hydroxybenzoate, Methoxybenzoate, phthalate, terephthalate, benzene sulfonate, toluene sulfonate, chlorobenzene sulfide Propyl sulphonate, naphthalene-1-yne, xylenesulfonate, phenylsulfate, phenylbutyrate, citrate, lactate,? -Hydroxybutyrate, glycolate, maleate, Sulfonate, naphthalene-2-sulfonate or mandelate.

The acid addition salt according to the present invention can be obtained by a conventional method, for example, by dissolving the compound represented by the formula (1) in an excess amount of an acid aqueous solution, and then mixing the salt with a water-miscible organic solvent such as methanol, ethanol, acetone or acetonitrile &Lt; / RTI &gt; It may also be prepared by evaporating a solvent or excess acid in this mixture and then drying or by suction filtration of the precipitated salt.

In addition, the base may be used to make a pharmaceutically acceptable metal salt. The alkali metal or alkaline earth metal salt is obtained, for example, by dissolving the compound in an excess amount of an alkali metal hydroxide or alkaline earth metal hydroxide solution, filtering the insoluble compound salt, and evaporating and drying the filtrate. At this time, it is preferable for the metal salt to produce sodium, potassium or calcium salt. The corresponding silver salt is obtained by reacting an alkali metal or alkaline earth metal salt with a suitable salt (such as silver nitrate).

The term "obesity " as a disease to be improved, prevented or treated by the composition of the present invention means a state in which the fat cells multiply and differentiate in the body due to metabolic disorders and the fat is accumulated excessively, , The number and volume of adipocytes are increased, resulting in an increase in the mass of adipose tissue. Obesity at the cellular level means an increase in the number and volume of adipocytes due to promotion of proliferation and differentiation of adipocytes. In addition, it accelerates glucose uptake in muscle skeleton, thereby increasing sugar consumption and improving energy synthesis in mitochondria, resulting in an obesity-suppressing effect through energy consumption. Increased energy synthesis at the cellular level implies improved insulin resistance due to accelerated sugar consumption in muscle cells and improved diabetes through energy expenditure.

Obesity is closely associated with increased insulin resistance, a major pathophysiological feature of type 2 diabetes. Insulin resistance means a decrease in insulin sensitivity with no blood glucose lowering even with a large amount of insulin injections. This reduction in insulin sensitivity is due to the irregular secretion of adipokines and free fatty acids, which cause fatty acids to accumulate in insulin-sensitive tissues such as the beta cells, kidney, liver, and heart, resulting in lipotoxicity It is known to occur. Therefore, the "lipid-related metabolic disease" of the present invention is also subject to prevention or treatment, and may be any one or more selected from the group consisting of fatty liver, type 2 diabetes, hyperlipidemia, cardiovascular disease, and arteriosclerosis , But is not limited thereto.

As used herein, the term "prophylactic " means any action that inhibits or delays the onset of obesity or lipid-related metabolic diseases by administration of the pharmaceutical composition according to the present invention.

As used herein, the term "treatment" means any action that improves or alters the symptoms of obesity or lipid-related metabolic diseases by the administration of the pharmaceutical composition according to the present invention.

The pharmaceutical composition according to the present invention may contain, in addition to the active ingredient, a pharmaceutically acceptable carrier. Herein, pharmaceutically acceptable carriers are those conventionally used at the time of formulation, such as lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia rubber, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose But are not limited to, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. Further, in addition to the above components, a lubricant, a wetting agent, a sweetener, a flavoring agent, an emulsifying agent, a suspending agent, a preservative, and the like may be further included.

The pharmaceutical composition of the present invention may be administered orally or parenterally (for example, intravenously, subcutaneously, intraperitoneally or topically) depending on the intended method, and the dose may vary depending on the condition and weight of the patient, The mode of administration, the route of administration, and the time, but may be appropriately selected by those skilled in the art.

The pharmaceutical composition of the present invention is administered in a pharmaceutically effective amount. In the present invention, the term "pharmaceutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment, and the effective dose level will depend on the type of disease, severity, The sensitivity to the drug, the time of administration, the route of administration and the rate of release, the duration of the treatment, factors including co-administered drugs, and other factors well known in the medical arts. The pharmaceutical composition according to the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, sequentially or concurrently with conventional therapeutic agents, and may be administered singly or in multiple doses. It is important to take into account all of the above factors and to administer the amount in which the maximum effect can be obtained in a minimal amount without side effects, which can be easily determined by those skilled in the art.

Specifically, the effective amount of the pharmaceutical composition of the present invention may vary depending on the age, sex, condition, body weight, absorbency, inactivation rate and excretion rate of the active ingredient in the body, type of disease, 100 to 500 mg / kg of body weight may be administered daily or every other day, or one to three times a day. However, the dosage may be varied depending on the route of administration, the severity of obesity, sex, weight, age, etc. Therefore, the dosage is not limited to the scope of the present invention by any means.

In one embodiment of the present invention, atractylenolide Ⅲ (Atractylenolide Ⅲ) was prepared and prepared (Example 1), and the insulin resistance-improving transcription factor-related gene (PPAR- (PGC1?, Tfam,?,?,?, C / EBP ?, SREBP 1c, FAS, ACS, perillipin, Adiponectin) NRF1, Cpt1?, UCP3, SIRT1) and proteins (AMPK, PGC1?, Tfam). Thus, it has been confirmed that the composition of the present invention can be effectively used for prevention or treatment of obesity or lipid-related metabolic diseases.

Accordingly, the present invention provides a food composition for preventing or ameliorating obesity or lipid-related metabolic diseases, which comprises Atractylodis macrocephala rhizoma extract or Atractylenolide III as an active ingredient.

As used herein, the term "improvement" means all actions that at least reduce the degree of symptom associated with the condition being treated. At this time, the functional food composition may be used simultaneously or separately with the medicament for treatment before or after the onset of the disease for the prevention or improvement of obesity or lipid-related metabolic diseases.

The term "food composition" as used in the present invention means a food composition containing at least one of a carrier, diluent, excipient and additives and being formulated into one selected from the group consisting of tablets, pills, powders, granules, powders, . Examples of foods that can be added to the present invention include various foods, powders, granules, tablets, capsules, syrups, beverages, gums, tea, vitamin complexes, and health functional foods. Examples of the additive that can be further included in the present invention include natural carbohydrates, flavors, nutrients, vitamins, minerals (electrolytes), flavors (synthetic flavors, natural flavors and the like), colorants, fillers, , At least one component selected from the group consisting of an inorganic acid and a salt thereof, an organic acid, a protective colloid thickening agent, a pH adjusting agent, a stabilizer, a preservative, an antioxidant, glycerin, an alcohol, a carbonating agent and a pulp. Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And polysaccharides, for example, conventional sugars such as dextrin, cyclodextrin and the like, and sugar alcohols such as xylitol, sorbitol and erythritol. As the above-mentioned flavors, natural flavors (tautatin, stevia extract (for example, rebaudioside A and glycyrrhizin) and synthetic flavors (saccharin, aspartame, etc.) can be advantageously used. The composition according to the present invention can be used in various forms such as flavorings such as various nutrients, vitamins, minerals (electrolytes), synthetic flavors and natural flavors, colorants and heavies, factic acid and its salts, alginic acid and its salts, , A pH adjusting agent, a stabilizer, a preservative, a carbonating agent used in glycerin, an alcohol, a carbonated beverage, etc. In addition, the composition according to the present invention may contain flesh for the preparation of fruit juice and vegetable drinks . These ingredients may be used independently or in combination. Specific examples of the carrier, excipient, diluent and additives include, but are not limited to, lactose, dextrose, But are not limited to, sucrose, sorbitol, mannitol, erythritol, starch, acacia, calcium phosphate, alginate, gelatin, calcium phosphate, calcium silicate, microcrystalline cellulose, polyvinylquilolidone, cellulose, polyvinylpyrrolidone, methylcellulose, It is preferable to use at least one selected from the group consisting of sugar syrup, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.

In another aspect of the present invention, the present invention provides a method for treating obesity or lipid-related metabolic diseases comprising administering the above pharmaceutical composition to a subject. The term " individual "as used herein refers to a subject in need of treatment for a disease, and more specifically refers to a mammal such as a human or non-human primate, mouse, dog, cat, horse and cattle .

Hereinafter, preferred embodiments of the present invention will be described in order to facilitate understanding of the present invention. However, the following examples are provided only for the purpose of easier understanding of the present invention, and the present invention is not limited by the following examples.

Example 1: Preparation of dried extract

In the present invention, as the active ingredient, rhizome of Atractylodis macrocephala KOIDZ (Atractylodis macrocephala rhizoma, ARA) was added, and 2 L of purified water was added to 210 g of extract, which was heated for 3 hours using a hot water extractor. Thereafter, the mixture was filtered with 3-layer gauze and Watman filter paper, concentrated using a rotary vacuum concentrator, and lyophilized to prepare a dried extract of the present invention. The yield of the extract was 26%, and the extract was stored at 4 ° C and used as an efficacy test. In addition, Atractylenolide III, one of the active ingredients of the extract, was used for the test.

Hereinafter, the extract obtained from Example 1 was named ARA extract.

Example 2 Confirmation of Improvement of Insulin Resistance in Adipocytes

1. Fat cell modeling

The adipocytes were treated with 3T3-L1 mouse preadipocytes and 0.5 mM 3-isobutyl-1-methylxanthine (IBMX), 1 μM dexamethasone, and 5 μg / For 8 days to induce differentiation into adipocytes. In addition, ARA extract or Atractylenolide Ⅲ was treated while inducing adipocyte differentiation, and treated for 24 hours for RT-PCR and 45 minutes for Western blot.

2. Improvement of insulin resistance.

(PPAR-γ, C / EBPα, SREBP 1c, FAS, ACS, perillipin, and Adiponectin) associated with the improvement of insulin resistance by ARA extract or Atractylenolide Ⅲ treatment in the differentiation of adipocytes was confirmed by RT-PCR . Specifically, PPAR-γ (peroxidase proliferator-activated receptor-γ) is a member of the nuclear hormone receptor family and is a receptor for the thiazolidinedione (TZD) family that increases insulin sensitivity in energy metabolism. The gene enhances insulin resistance, which is a key pathology of diabetes by increasing the body's glucose disposal rate, and is highly expressed in adipose tissue and regulates the activation of transcription factors involved in lipid differentiation. Expression of the transcription factor C / EBPα (CCAAT / enhancer binding proteins), PPAR-γ and SREBP 1c (sterol regulatory element binding protein-1c) FAS (fatty acid synthase), ACS (acyl-CoA synthase), and perillipin are associated with fat transport, synthesis, and accumulation. Increased expression of these transcription factors increases glucose and fatty acid uptake in adipocytes, thereby improving insulin resistance by inducing lipid synthesis. In addition, Adiponectin has been evaluated as the most important factor of diabetes, and it is known that it is an adipocyte-specific expression protein that increases insulin sensitivity and prevents atherosclerosis. Therefore, it is clinically applicable to obesity and diabetes mellitus Critical targets are being recognized.

In this example, the control group (C) differentiated into immature lipoprotein (P) and adipocytes and the ARA extract (250, 500 / / ml) or Atractylenolide Ⅲ (50, 100 袖 M) RNA was isolated from the treated group by Trizol solution and then incubated with RT mixture [1 × reverse transcriptase buffer, 0.5 mM dNTP (deoxynucleotide triphosphate), 3 mM MgCl ₂ , 5 U RNase inhibitor, 0.5 μM oligo-dT primer, 5 U Superscript Reverse Transcriptase (PPAR-γ, C / EBPα, SREBP-1c, FAS, and ACS1) were confirmed by PCR. GAPDH was used as a house keeping gene to determine the synthesis of RT-PCR, and triglytazone (T) was used as a control drug to compare insulin resistance improving effects.

As a result, as shown in Fig. 1, the expression of PPAR-γ in adipocytes was increased in a concentration-dependent manner when ARA extract was treated, and PPAR-γ expression was also increased by treatment with Atractylenolide Ⅲ (A). In addition, the expression of C / EBPα, SREBP-1c, FAS, ACS1, and perillipin was also increased by ARA extract or Atractylenolide Ⅲ treatment, and the expression of Adiponectin associated with the enhancement of insulin sensitivity was also increased. This increase in gene expression was similar to or superior to that of the reference drug triglytidase, and the above results can be effectively used to improve insulin resistance through the control of adipocyte differentiation and inhibition of fat accumulation through ARA extract or atractylenolide Ⅲ, an active ingredient thereof .

3. Improvement of insulin resistance. Confirmation of transcription factor-related protein expression level

Activation of AMPK and PPAR-γ by ARA extract or Atractylenolide Ⅲ (A) treatment in the differentiation of adipocytes was confirmed by Western blot. Specifically, AMP-activated protein kinase (AMPK) is an enzyme protein that mediates the synthesis and degradation of fatty acids. Activation of AMPK in adipocytes leads to the formation and degradation of fat through inhibition of ACC and HSL (hormone sensitive lipase) Inhibiting insulin resistance through obesity inhibition. Therefore, the regulation of AMPK activation plays an important role in the prevention and treatment of various metabolic diseases such as lipid metabolism abnormality, obesity, fatty liver, type 2 diabetes, atherosclerosis, hyperlipidemia, and cardiovascular diseases.

In this example, the control group (C) differentiated into immature lipoprotein (P) and adipocytes and the ARA extract (250, 500 / / ml) or Atractylenolide Ⅲ (50, 100 袖 M) The protein was isolated from the treated group (ARA-250, ARA-500, A50, A100) and then treated with PPAR-γ and AMPK specific antibody (anti-PPAR-γ monoclonal antibody, anti- AMPK monoclonal antibody) blot. We used β-actin as the internal protein of Western blot and triglitazone (T) as the reference drug to compare insulin resistance improvement.

As a result, as shown in FIG. 2, protein expression of PPAR-γ and AMPK was promoted by treatment with ARA extract or Atractylenolide III. This increase in protein expression was similar to or better than that of the reference drug triglytidase. The results showed that, similar to Example 2-2, the results of the ARA extract or Atractylenolide III, an active ingredient thereof, can improve insulin resistance, And type 2 diabetes due to insulin resistance.

Example 3: Obesity suppression and insulin resistance improvement effect in muscle cells

1. Muscle cell modeling

The muscle cells were cultured in C2C12 mouse undifferentiated muscle cells. After culturing the muscle cells in an amount of 90% or more, the medium containing 2% horse serum was treated at intervals of 1 day for 5 days to induce differentiation into mature muscle cells . On the 6th day of differentiation, ARA extract or Atractylenolide Ⅲ was treated with the same medium and treated for 24 hours for RT-PCR and 45 minutes for Western blot.

2. Identification of genes involved in glucose uptake and energy metabolism-regulated transcription factors

Tfam, NRF1, Cpt1β, UCP3, or SIRT1, which increases gene expression associated with mitochondrial energy synthesis, by RT-PCR method to confirm the effect of glucose uptake and fatty acid oxidation in muscle cells Respectively. Specifically, PGC1α, Tfam, NRF1, CPT1, UCP3, and SIRT1 not only increase glycemic control by increasing glycemic control and regulation of function in cell respiration and mitochondria, ie, energy biosynthesis, It is associated with obesity treatment. In particular, SIRT1 is linked to insulin action, which reduces insulin resistance in muscle and is therefore attracting attention as a new target for the treatment of type 2 diabetes.

In this example, RNA was isolated from the control group (C) differentiated into muscle fat cells and the control group with the Trizol solution from the experimental group treated with the ARA extract (250, 500, 1000 / / ml) RT-PCR was performed to synthesize cDNA through transcriptase buffer, 0.5 mM of dNTP (deoxynucleotide triphosphate), 3 mM MgCl ₂ , 5 U RNase inhibitor, 0.5 μM oligo-dT primer and 5 U Superscript Reverse Transcriptase] Gene expression was confirmed. GAPDH was used as a control gene to determine the synthesis of RT-PCR, and metformin (M), the most widely used diabetes drug currently used as an AMPK activator, was used as a reference drug.

As a result, as shown in FIG. 3, it was confirmed that the expression of PGC1α, Tfam, NRF1, Cpt1β, UCP3 and SIRT1 was increased by the ARA extract treatment. This increase in gene expression is similar to or better than that of the reference drug, metformin, which means that by controlling blood glucose and promoting energy metabolism through the treatment of ARA extract, insulin resistance due to obesity in muscle can be improved.

3. Identification of protein-related and fatty acid consumption effects related to glucose uptake and energy metabolism

The changes of protein expression of AMPK, PGC1α, and Tfam were confirmed by Western blotting and the concentration of fatty acids in the cytoplasm was measured to confirm the effect of glucose consumption and fatty acid oxidation in muscle cells. Specifically, AMPK is an enzyme that acts as a sensor to maintain intracellular energy homeostasis. In muscle cells, when intracellular energy decreases due to metabolic stress or exercise, it accelerates the process of energy synthesis (fatty acid oxidation and the corresponding process) It inhibits obesity and promotes energy consumption through inhibition of lipolysis and fat production.

In this example, the control group (C) differentiated into muscle cells and the control group (ARA, A) treated with ARA extract (250, 500 ㎍ / ml) or Atractylenolide III and the control group (M) And then subjected to Western blotting using a specific antibody for each target protein. Β-actin was used as a control protein in Western blot, metformin was used as a comparative drug for comparison, and a fatty acid measurement kit (Abcam ^® ) was used to measure the concentration of fatty acids in the cytoplasm.

As a result, as shown in FIG. 4 and FIG. 5, when ARA extract or Atractylenolide III treatment was performed, phosphorylation of AMPK (p-AMPK) was increased and expression of PGC1α and Tfam was similarly increased similarly to the control drug, Especially, atractylenolide III and Tfam were significantly increased in PGC1α treated with ARA extract. In addition, it was confirmed that the accumulation of cytoplasm was decreased by further promoting fatty acid oxidation compared with the reference drug. The above results indicate that atractylenolide Ⅲ treatment of ARA extract or its active ingredient can promote metabolism of glucose in muscles and increase energy metabolism in mitochondria to treat metabolic diseases such as type 2 diabetes due to obesity and insulin resistance do.

It will be understood by those skilled in the art that the foregoing description of the present invention is for illustrative purposes only and that those of ordinary skill in the art can readily understand that various changes and modifications may be made without departing from the spirit or essential characteristics of the present invention. will be. It is therefore to be understood that the above-described embodiments are illustrative in all aspects and not restrictive.

Claims (7)

delete A pharmaceutical composition for the prevention or treatment of obesity comprising atractylenolide (III) or a pharmaceutically acceptable salt thereof as an active ingredient
3. The method of claim 2,
Wherein said atactylenolide (III) is isolated from a dried extract.
delete delete A food composition for preventing or improving obesity comprising atractylenolide (III) or a pharmaceutically acceptable salt thereof as an active ingredient.
The method according to claim 6,
Wherein said atactylenolide (III) is isolated from a dried extract.

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