KR20210099482A - Obesity prevention or treatment composition using soybean processing by-product as an active ingredient - Google Patents

Abstract

The present invention relates to a composition for preventing or treating obesity, including a soybean processing byproduct as an active ingredient. Particularly, the present invention relates to a composition for preventing or treating obesity, including water strained off coagulating bean curd, brew soybean in water or a mixture thereof as an active ingredient. The composition according to the present invention uses an extract of natural food and is not harmful to the human body, shows an excellent effect of reducing fat accumulation in the body and inhibits acceleration of fatty acid oxidation, and thus can be used effectively for preventing and treating obesity caused by high-fat intake with no side effect. In addition, the composition according to the present invention has an advantage in that it can contribute to prevention of various diseases, such as metabolic, cardiovascular and cerebrovascular diseases and cancer, caused by obesity.

Description

Obesity prevention or treatment composition using soybean processing by-product as an active ingredient {Obesity prevention or treatment composition using soybean processing by-product as an active ingredient}

The present invention relates to a composition for preventing or treating obesity using the by-product of soybean processing as an active ingredient, and more particularly, a composition for preventing or treating obesity comprising pure soybean, soybean immersion water, or a mixture thereof as an active ingredient. is about

In modern society, along with the westernization of dietary life due to the improvement of living standards, the intake of plant foods gradually decreases due to changes in eating patterns, while the intake of animal foods containing a lot of saturated fatty acids or cholesterol increases. It is emerging as a problem (Kim et al, 1993).

Obesity is a type of metabolic disease caused by excessive accumulation of body fat due to an imbalance in energy intake and consumption (Song et al, 2009). It is a complex syndrome involving various causes such as genetic, environmental and social structural changes. As a result (Grundy, 1998; Albu et al, 1997), excess energy is consumed for metabolic activity in the body and the remaining nutrients are accumulated as triglycerides in adipose tissue. It affects biochemical functions (Huh, 1990) and the exact mechanism of obesity is not known, but it is known that the higher the degree of obesity, the higher the risk of cardiovascular disease. Obesity is attracting a lot of attention worldwide because it can cause diseases such as cardiovascular disease, diabetes, respiratory disease, and osteoarthritis as well as its own problems (Antipatis and Gil, 2001).

Obesity is a combination of endocrine factors, drug causes, and genetic factors as well as excessive energy intake and lack of exercise, so diet, exercise, behavioral therapy, or drug treatment are used as methods to improve obesity. As drug therapy, drugs such as Reductil, which has an appetite suppressant effect, and Xenical, which exhibits an effect of inhibiting digestion and absorption of fat, are used in clinical practice depending on the mechanism of action that causes obesity (Kim, 2004). As it is known that it can cause side effects such as dry mouth, constipation, and increased blood pressure (Reddy and Chow, 1998), the development of safer and more effective natural functional substances is required. the current situation.

In Korea, many natural substances are marketed as health foods that are effective in preventing obesity, but most of them have not been scientifically verified, and they are rather a cause of health food abuse, which is a social problem.

On the other hand, soybean pure water and soybean soaked water are by-products generated during the tofu manufacturing process, and currently, pure soybean water and soaked water are discarded during the manufacturing process of tofu. A study was conducted on the antioxidant activity (Kim and Lee, 2007) and functional properties (Kim et al, 2006) of some pure soybeans and soybean soaked water, and changes in the physical properties of tofu according to the soaking stock (Jang et al, 1995). However, there are very few studies on the physiological effects of pure water and soaking water, which are by-products of soybean processing.

Therefore, there is an urgent need to objectively verify natural resources that can be eaten as food to develop natural substances that have therapeutic and preventive effects on obesity.

Therefore, while the present inventors continued research on the development of natural substances having therapeutic and preventive effects on obesity, soybean pure water and soybean steep water, which are by-products of soybean processing, improved lipid metabolism in mice by a high-fat diet and excellent natural anti-obesity The physiological activity efficacy was confirmed, and the present invention was completed.

An object of the present invention is to provide a natural food extract that is harmless to the human body and can reduce fat accumulation in the body due to a high-fat diet.

Another object of the present invention is to provide a soybean processing by-product having an effect of preventing and treating obesity.

Another object of the present invention is to provide a composition for preventing or treating obesity comprising the by-product of soybean processing as an active ingredient.

In order to achieve the above object, the present invention provides a composition for preventing or treating obesity, and more particularly, to provide a composition for preventing or treating obesity using a by-product of soybean processing as an active ingredient.

The present invention also provides a health supplement comprising the composition.

Hereinafter, the present invention will be described in detail.

The composition according to the present invention is characterized in that it contains a by-product of processing soybean, and more particularly, it is intended to provide a by-product of processing soybean, which is used for preventing and treating obesity due to a high-fat diet, and a composition for preventing or treating obesity containing the same. characterized.

The soybean processing by-product of the present invention is characterized in that it is soybean soaking water, soybean curd whey, or a mixture thereof.

As used herein, the term “prevention” refers to inhibiting the occurrence of a disease or disease in an animal that has never been diagnosed as possessing a disease or disease, but is prone to such disease or disease. As used herein, the term “treatment” refers to (a) inhibiting the development of a disease or disorder; (b) alleviation of the disease or condition; and (c) the disease or elimination of the disease.

The present invention provides a composition for preventing or treating obesity, which contains, as an active ingredient, a by-product of soybean processing.

The soybean processing by-product is soybean soaking water, soybean curd whey, or a mixture thereof, and more preferably, soybean soaking water is used.

In the present invention, the pure tofu (soybean curd whey) is a supernatant obtained by centrifuging at 10,000 to 14,000 rpm of pure liquid generated during tofu production, and then removing the solids using filter paper, It includes a concentrate of the supernatant, or a dried product prepared by freeze-drying the concentrate.

In the present invention, the soybean soaking water (soybean soaking water) is obtained by immersing soybeans in water at room temperature. It includes a filtrate obtained after immersion, a concentrate of the filtrate, or a dried product prepared by freeze-drying the concentrate.

The composition of the present invention may be prepared by including the active ingredient, which is the soybean soaking water, soybean curd whey, or a mixture thereof, in an amount of 0.001 to 30% by weight of the total composition.

The active ingredient inhibits fat degeneration and fat accumulation in the liver and epididymal tissues; inhibition of the accumulation of cholesterol in the body; inhibition of triglyceride accumulation, differentiation of pre-adipocytes into adipocytes, and proliferation of adipocytes; inhibition of adipogenesis; inhibition of the activity of acetyl-CoA carboxylase (ACC); and increased activity of AMP-induced activated protein kinase (AMPK, AMP-activated protein kinase); characterized in that it controls the underlying cause of obesity through at least one action selected from the group consisting of:

More specifically, the active ingredient is characterized in that it has an anti-obesity effect by inhibiting the activity of the peroxisome proliferator activated receptor gamma (PPARγ, peroxisome proliferator activated receptor).

The composition of the present invention is prepared in unit dosage form by formulating using a pharmaceutically acceptable carrier and/or excipient according to a method that can be easily carried out by a person skilled in the art to which the present invention pertains, or It can be prepared by pouring into a multi-dose container. In this case, the formulation may be in the form of a solution, suspension or emulsion in oil or an aqueous medium, or may be in the form of a powder, granule, tablet, or capsule, and may additionally contain a dispersing agent or a stabilizer.

The composition of the present invention can be developed as a dietary supplement. The health supplement is at least one selected from drinks, caramel, chocolate, diet bar formulations, and confectionery, and includes ingredients commonly added during food production, for example, protein, carbohydrate, fat, nutrients, seasoning and flavoring agents. Examples of the above-mentioned carbohydrates include monosaccharides such as glucose, fructose and the like; disaccharides such as maltose, sucrose, oligosaccharides 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 flavoring agent, natural flavoring agents [taumatine, stevia extract (eg, rebaudioside A, glycyrrhizin, etc.)] and synthetic flavoring agents (saccharin, aspartame, etc.) can be used. For example, when manufactured as a drink, citric acid, high fructose, sugar, glucose, acetic acid, malic acid, fruit juice, cephalothorax extract, jujube, in addition to stannyocalcin 2 of the present invention

Extract, licorice extract, etc. may be additionally included. Considering the easy access to food, the food of the present invention is very useful for the treatment or prevention of obesity, and improvement of cognitive function.

The composition according to the present invention is harmless to the human body by using an extract of natural food, has an excellent effect of reducing the accumulation of fat in the body, and inhibits the promotion of fatty acid oxidation, thereby causing obesity caused by high fat intake can be effectively used for prevention and treatment without side effects, and has the advantage of greatly contributing to the prevention of various diseases such as metabolic and cardiovascular, cerebrovascular diseases, and cancer caused by obesity.

1 is a result of confirming the histological changes of hepatocytes in the soybean processing by-product intake diet group according to the present invention, (A: normal diet group, B: high-fat diet group, C: soybean pure water diet group, D: soybean steepness index Dietary group) FIG. 2 is the result of confirming the histological changes of fat cells in the diet group consuming soybean processing by-products according to the present invention, (A: normal diet group, B: high-fat diet group, C: soybean pure product diet group, D : Soybean steep water diet group)
carboxylase), (ND: normal diet group, HFD: high-fat diet group, H-SCW: soybean pure water diet group, H-SSW: soybean steep water diet group) FIG. 4 shows soybean processing according to the present invention It is a result of confirming the distribution of mouse abdominal adipose tissue for the by-product intake diet group, (A: normal diet group, B: high-fat diet group, C: soybean pure water diet group, D: soybean steep water diet group) The cell viability of whole adipocytes was confirmed according to the concentration of soybean processing by-product treatment of ) and after differentiation (B) are confirmed through a microscope, FIG. 8 is the result of confirming the differentiation ability of whole adipocytes according to the concentration of soybean processing by-product of the present invention, and FIG. It is the result of confirming the expression of peroxisome growth factor activation receptor gamma (PPARγ) according to the concentration, and FIG. 11 shows the results of quantification of the expression of fat metabolism-related genes according to the concentration of soybean processing by-products of the present invention using ImageQuant program, (A: APOE, B: adipsin, C: Fabp4, D: Scd1)
12 is a result of confirming the expression of adipokine-related genes according to the treatment concentration of soybean processing by-products of the present invention.

The present invention will be described in more detail by the following examples. However, the following examples are only for helping understanding of the present invention, and the scope of the present invention is not limited by these examples in any sense.

At this time, if there is no other definition in the technical terms and scientific terms used, it has the meaning commonly understood by those of ordinary skill in the technical field to which this invention belongs, and the gist of the present invention in the following description and accompanying drawings Descriptions of known functions and configurations that may be unnecessarily obscure will be omitted.

[Production Example]

(1) Soybean curd whey

The pure water sample used in the experiment was centrifuged at 12,000 rpm for liquid pure water generated during tofu production, and then the solid content was removed using filter paper (whatman No1) to obtain a supernatant, and the obtained supernatant was concentrated , and freeze-dried to use dry samples. On the other hand, as the liquid sample, liquid pure water generated during the production of tofu was used as the sample.

(2) soybean soaking water

Soybeans were immersed in water at 15° C. for 8 hours without hot water heating, followed by filtration, concentration, and freeze-drying before use.

[Experimental Example 1]

Three-week-old male mice (C57BL/6) used in this experiment were purchased from Samtaco and bred in a breeding room under constant conditions (temperature 214±005° C., humidity 61±1%, light and dark cycle 12 hours). The diet was acclimatized for 1 week with sufficient intake of water (distilled water) and feed [Dyets' feed (AIN-93G Purified Rodent Diet) purchased from Central Laboratory Animals Co., Ltd.).

After acclimatization, each experimental animal ingested a high-fat feed according to each experimental group (40% Beef Tallow+AIN-76A Diet), and the induction of obesity and the effect of each sample on obesity induction were investigated for about 7 weeks. Food and drinking water were freely ingested, and all experimental diets were refrigerated during the breeding period and used in the experiment.

The experimental group consisted of four groups: a normal diet group (Normal diet (fat: 117%), ND), a high fat diet group [High fat diet (fat: 40%), HFD], and a 10% soybean pure plant diet group [high fat ( fat: 40%) + 10% soybean pure water prepared in Preparation Example, H-SCW], 10% soybean steep water diet group [high fat (fat: 40%) + 10% soybean soaked in Preparation Example) , H-SSW] and fed with feeds of different compositions as shown in Table 1 below for about 7 weeks. In addition, in order to observe the change in body weight during the experiment, the body weight was measured at a certain time every day, and the amount of change in feed and drinking water was checked.

In this experimental example, all animal experiments were performed according to the Principle of Laboratory Animal Care of the National Institutes of Health (NIH) and the guidelines for the management and use of laboratory animals of Andong University.

On the last day of the experiment, the animals were fasted for about 12 hours, anesthetized with ether, and blood was collected from the carotid artery. Blood was collected in a serum tube and centrifuged at 1600 x g for 15 minutes to obtain serum, and stored frozen at -80°C until analysis. After blood collection, epididymal adipose tissue and liver tissue from the laparotomy animals were removed and washed with physiological saline.

After washing, the water was wiped off with a filter paper. Live tissue was weighed with an electronic balance and stored frozen at -80°C until analysis.

(1) Adipose tissue mass and histological changes in liver and epididymal adipose tissue

In order to investigate the effect of soybean stock and soybean steep water on obesity in mice caused by a high-fat diet, the weights of liver and epididymal adipose tissue were measured and compared.

To measure the weight of adipose tissue, liver tissue and epididymal adipose tissue were extracted immediately after laparotomy, washed with a phosphate buffer solution, water was removed with filter paper, and the weight was measured.

The weight of liver tissue per each experimental group was higher in the high-fat diet group compared to the normal diet group.

832% higher. On the other hand, the soybean pure water diet group and the soybean steep water diet group decreased by 248% and 683%, respectively, compared to the high fat diet group. Also, as a result of examining the weight of epididymal fat, the high-fat diet group was 210% higher than the normal diet group, and the soybean pure water diet group and the soybean steep water diet group decreased to 243% and 769%, respectively, compared to the high-fat diet group.

In addition, in order to check the size of adipocytes in the adipose tissue, the adipose tissue after weight measurement was fixed with a 4% formaldehyde solution. After fixation, water washing and dehydration were performed, and then paraffin solution was treated to make paraffin blocks, and then sectioned to a thickness of 4 to 6 μm. The sectioned tissue contains hematoxylin and eosin.

(eosin), observed under an optical microscope (Olympus BX50, Japan), and photographed using Olympus DP-71 to observe the adipose tissue of each experimental group (Heo etal, 2009; Choietal, 2006)

As a result, in the high-fat diet group, excess fat and cholesterol were not normally excreted from the body, so fat accumulation in the liver was induced, resulting in an enlarged liver. However, it was found that fat accumulation was decreased by the diet of pure water and soaked water. In the high-fat diet group, excess fat and cholesterol were not normally excreted from the body, leading to fat accumulation in the liver, resulting in enlarged liver and epididymal adipose tissue. It is also the result of confirming that it is effective in reducing.

In particular, it was found that soybean soaking water significantly inhibited the accumulation of fat in the liver and epididymal adipose tissue caused by a high-fat diet. could

(2) Histopathological changes in liver and epididymal adipose tissue

The liver is a digestive organ that synthesizes glucose into glycogen and stores it. Glycogen produced by anabolic action is used to control blood sugar, but if it is too much, it is converted into fat and stored. In addition, since obesity is characterized by an increase in adipose tissue weight and an increase in blood fat content (Friedman et al, 1991), the investigation of epididymal fat is an important indicator to investigate changes in body weight and obesity. When diet-induced obesity is induced, both weight and size of epididymal fat increase (Fujioka, 2002).

In this respect, the effect of pure water and immersion water on the histological changes of the liver and epididymal fat of mice by a high-fat diet was investigated. It was confirmed that Kupffer cells between the liver cells were evenly spread throughout. On the other hand, in the case of obesity-induced mice with a high-fat diet, severe fatty degeneration and vacuolation were observed around the nucleus.

In contrast, in the pure soybean diet group, hepatocytes were enlarged due to intracellular fat accumulation compared to the normal diet group, but the phenomenon of lipophagy in the hepatocyte volume was not evident as in the high fat diet group. It was confirmed that only the fat phobia was confirmed, but the majority of hepatocytes and Kupffer cells were similar to normal hepatocytes, confirming that serious liver tissue abnormalities such as those in the high-fat diet group did not appear.

In addition, as a result of examining the effect of soybean pure water and soybean soaking water on the morphological changes of epididymal fat, which is white fat, as can be seen in FIG. I could see that it had grown.

On the other hand, the soybean pure water diet group and the soybean soaked diet group had smaller adipocytes compared to the high fat diet group. It was confirmed that it was almost similar to fat cells. According to the above results, in a recent study, Moon et al. (2003) reported that a natural material composition composed of herbal medicines was effective in anti-obesity by reducing the amount of epididymal adipose tissue and reducing the size of adipocytes. As reported to have an effect of reducing the accumulation of epididymal fat, it is the result of confirming that the immersion water of soybean puree and soybean of the above example is effective in suppressing fatty degeneration of the liver and epididymal tissue due to the high-fat diet, and furthermore, the anti-obesity effect It is also the result of confirming that there is

(3) Biochemical analysis of blood components

Obesity often accompanies various dyslipidemias due to abnormal flow of fat metabolism and glucose metabolism. (Lee et al, 2006)

In addition, the change in plasma GOT and GPT enzyme activity is an indicator of liver tissue damage as hepatocyte degeneration and necrosis progress and transaminase is released and increased in the blood.

In this experiment, the collected blood was centrifuged at 3,000 rpm for 10 minutes and plasma was used for analysis. The separated plasma contains amino acid transferase (GOT, GPT), total cholesterol, high density lipoprotein, low density lipoprotein, total lipid and triglyceride. For the investigation of the content, it was analyzed in a hematology analyzer (Hitachi 7600).

As a result, the GOT activity in the serum was compared with the normal diet group and each experimental group.

The high fat diet group showed an increase rate of 278%, the soybean pure product diet group increased by 571%, and the soybean soaked water diet group showed an increase rate of about 206%. In the case of the immersion water diet group, the values were similar to those of the normal diet group.

In addition, the serum total cholesterol level was 936% in the high-fat diet group, 712% in the pure soybean diet group, and 132% in the soybean steep water diet group, compared to the normal diet group. In particular, the triglyceride content was normal in the high-fat diet group. It increased by 2527% compared to the diet group, showing a very large numerical change.

As a result, it was confirmed that the soybean steep water diet group was effective in reducing the increase rate of cholesterol and triglycerides in the body due to the high-fat diet. In addition, HDL-cholesterol level was highest in the high-fat diet group compared to the normal diet group, and the LDL-cholesterol level showed the lowest decrease in the soybean soaked diet group compared to the high-fat diet group and the soybean pure soybean diet group.

Based on the results of several studies (Park et al, 2005; Wat et al, 2009; Davignon and Cohn, 1996) that a reduction in blood lipid content reduces the risk of coronary heart disease, the It is also the result of confirming that it is effective in reducing the cardiovascular risk induced by obesity.

(4) phosphorylation ratio of obesity target protein

In this experiment, the expression of p-AMPK and p-ACC was investigated in liver tissue to investigate the activity of AMPK and ACC, target proteins of obesity.

In liver tissue, phospho-AMPK and phospho-ACC were confirmed by western blot. The liver tissue was homogenized using a lysis buffer (50 mM Tris-HCl pH 75, 1 mM EDTA, 025% sucrose, 04 mg/ml digonin and 15 mM PMSF), and protein quantification was performed with Bio-Rad assay reagent (Bio- Rad, USA). extracted from the liver

After electrophoresis of 40 μg of protein, primary antibody (p-AMPK-alpha, p-Acetyl-CoA) (Cell signaling Technology, Beverly, USA) was treated, and secondary antibody (anti-rabbit IgG-HRP) (Santa) cruz biotechnology, Santa Cruz, USA) to determine the phosphorylation ratio of AMPK and ACC.

As can be seen in FIG. 3, as a result of investigating the phosphorylation ratio of the experimental group relative to the expression of p-ACC in the normal diet group, p-ACC in the high-fat diet group was 230%, whereas in the pure soybean diet group, 836%, and soybean saliva The exponential diet group was 1642%.

From the above results, it was confirmed that soybean pure water and soybean steepwater effectively acted on phosphorylation of ACC compared to a high-fat diet. By inhibiting the promotion of oxidation, it was confirmed that the anti-obesity effect was very high.

(5) Distribution of adipose tissue in the abdomen

To measure abdominal fat volume in 12-week-old male C57BL/6 mice, lumbar vertebrae 2 to 5 were used using an in vivo micro-tomography system (In-vivo Micro-CT, Skyscan 1076, SKYSCAN NV, Belgium). No. (L2~L5) was taken. For anesthesia of mice during imaging, ketamine (Ketamine, Huons, 15 ml/kg) and xylazine (Xylazine, Bayer, 05)

ml/kg) was used. To acquire abdominal fat images, the voxel size of the in vivo micro tomography system was set to 35 , the filter was aluminum 10 mm, the exposure time was 790 ms, the voltage was 100 kV, the current was 100 , and the rotation angle was set to 12 ° (deg). did.

Using the Mimics 130 (materialise NV, Belgium) program, gray values corresponding to each tissue inside the abdomen of a male C57BL/6 mouse were obtained from the images obtained using the in vivo micro tomography system. did. Each area is divided into lumbar vertebral, lean tissue, adipose tissue, and skin in the image based on the gray value of each tissue measured using the threshold method. Separated, the extracted abdominal adipose tissue was reconstructed into a three-dimensional structure, and the volume was measured therefrom.

As can be seen in FIG. 4, the distribution of abdominal adipose tissue in mice was confirmed through an in vivo micro tomography system. As a result, it was confirmed that a significant amount of subcutaneous fat and visceral fat occupied a high area in the abdomen in the high-fat diet group compared to the normal diet group. became However, the 10% pure soybean meal group had more abdominal fat than the high fat diet group, and it was confirmed that the 10% soybean soaked water diet group was reduced by about 17 to 27 times compared to the soybean pure water group and the high fat diet group.

Also, when comparing body weight and abdominal fat according to each experimental group, the high-fat diet group showed the highest body weight (405±05 g), but the volume of visceral fat and subcutaneous fat was the volume of total abdominal fat in the soybean pure-grain diet group. showed the highest value at 42057±5112 ㎣. On the other hand, the soybean soaked water diet group showed a significant decrease in the volume of abdominal fat compared to the high fat diet group and the soybean pure water diet group. It showed a similar value to 1297 ㎣.

From the above results, it was confirmed that the soybean steeped diet group was effective in suppressing the accumulation of abdominal fat due to the high fat diet compared to the soybean pure water diet group.

[Experimental Example 2]

(1) Cell viability of whole adipocytes according to the concentration of soybean processing by-product treatment

3T3-L1 pre-adipocyte cells (pre-adipocyte cells) used in the experiment were purchased from the American Type Culture Collection (ATCC) and added to Dulbecco's modified Eagle's medium (DMEM, Gibco Inc, USA) with 10% fetal bovine serum and 1 % antibiotics-antimycotic was added and subcultured once every 2-3 days in a cell incubator at 37°C in the presence of 5% CO2 and used in the experiment.

The concentration-dependent cell viability by soybean by-product treatment was measured by MTT assay. The cultured 3T3-L1 whole adipocytes were aliquoted in a 96-well plate to 5×103 cells/well, the supernatant was removed after culturing for 24 hours, and the soybean by-product extraction sample was treated with the medium at concentrations of 10 ppm, 25 ppm, and 100 ppm. did.

The soybean by-products were treated by concentration and incubated for 24 hours, then 10 μl of MTT reagent (5 mg/ml) was added per well and further cultured for 3 hours. Thereafter, the supernatant was removed, and 150 μl of DMSO was dispensed into each well, stirred for 30 minutes, and then absorbance was measured at 570 nm using a microplate reader (Infinite200, Tecan Trading AG, Switzerland). This absorbance indicates the amount of MTT reduced by the cells and is proportional to the number of viable cells in each well.

Cell viability (%) = (absorbance of sample treatment group / absorbance of control group) × 100

As a result, as can be seen in FIG. 5, it was confirmed that the soybean steep water showed a cell viability of more than 95% when treated with a high concentration of 100 ppm, and 9521% at 10 ppm, 9212% at 25 ppm, and 100% when treated with pure soybean water. Cell viability was 8865% (P<005) at ppm, and it was confirmed that the cell viability was partially reduced when a high concentration of soybean processing by-products was treated.

(2) Effect on the efficacy of inhibiting adipogenesis

To analyze the effect on the differentiation inhibitory efficacy of 3T3-L1 adipocyte cells (adipocytes), cultured cells were treated with soybean steep water and soybean pure water at concentrations of 10 ppm, 25 ppm, and 100 ppm, respectively, together with a differentiation inducer. . After the differentiation induction process was completed, the cells were stained with Oil Red O and observed under a microscope, and then the fat content was quantified. Cells not treated with differentiation inducing substances and cells treated with differentiation inducing substances were used as controls.

As a result, as can be seen in FIG. 6 , compared with the cells treated with the differentiation inducing substance, the group treated with the soybean extract showed 9442% at 10 ppm, 8709% at 25 ppm, and 7914% at 100 ppm, increasing the treatment concentration. As a result, it was confirmed that the fat content in adipocytes was significantly reduced.

In addition, it was confirmed that the dyed fat content decreased as the concentrations of soybean pure water and soybean steepwater increased, as can be seen in FIG.

From the above results, it was confirmed that in the case of soybean pure water, the adipocyte differentiation inhibitory effect appeared as the treatment concentration increased, and in the case of soybean steep water, it was confirmed that 8554% of the adipocyte differentiation inhibitory effect was exhibited even at a low concentration of 10 ppm treatment. This is also the result of confirming that soybean processing by-products have the effect of inhibiting the differentiation of pre-adipocytes into adipocytes and the formation of adipocytes.

[Experimental Example 3]

(1) Effect on expression of differentiation-inducing final transcription factor PPARγ

In order to examine the effect on the expression of genes involved in the induction of differentiation of 3T3-L1 adipocyte cells (adipocytes), differentiation-induced control was treated with 10 ppm, 25 ppm and 100 ppm of soybean pure water and soybean steep water, respectively. The induced experimental groups were compared.

It is known that preadipocytes induce the expression of the final transcription factor, PPARγ, by appropriate differentiation stimulation and expression of adipogenesis-inducing regulators. Peroxisome proliferator activated receptor gamma (PPARγ, peroxisome proliferator activated receptor) belongs to the nuclear hormone receptor superfamily, and members of the PPAR family include PPARα,

PPARγ and PPARζ are known. These PPAR transcription factors form a heterodimer with Retinoid X receptor α to recognize the peroxisome proliferator response element (PPRE) of the target gene and regulate the expression of adipocyte differentiation-related genes involved in adipogenesis. Therefore, in order to confirm this, the expression of PPARγ, the final transcription factor for adipocyte formation following differentiation induction and treatment of soybean by-products, was confirmed.

First, total RNA was extracted using TRIzol (Gibco Inc, USA), and then purified using a Total RNA Clean up kit (Ambion, USA). Single using the SuperScript™ kit (Invitrogen, USA) using purified RNA as a template

Stranded cDNA was prepared. The following primers were prepared to specifically amplify genes related to adipocyte differentiation, and the expression level of each gene was confirmed by comparing it with the expression of the GAPDH gene. The expression level of the gene was analyzed using the ImageQuant 52 (GE Healthcare, USA) program after electrophoresis.

As can be seen in FIG. 9, as a result of confirming the expression pattern of PPARγ mRNA in the treatment group treated with soybean pure water and soybean steep water, PPAR γ in the treatment group treated with soybean pure water and soybean steep water compared with the control induced to differentiate adipocytes. It was confirmed that the mRNA expression of In the case of soybean steepwater treatment, the expression decreased by less than 50% at all treatment concentrations, but some concentration-independent phenomena were confirmed. In addition, in the case of soybean stock, as the treatment concentration increased, the expression of PPARγ decreased in a concentration-dependent manner, which was consistent with the results of cell observation through Oil Red O staining.

From the above results, it could be confirmed that the inhibition of adipocyte differentiation of soybean stock and soybean steeped water was achieved through inhibition of PPARγ expression.

(2) Effect of soybean processing by-products on fat metabolism

When the differentiation induction process of adipocytes progresses, the expression of fat metabolism-related genes is increased by the regulation of PPARγ, a transcriptional regulator. It is known that an increase in the expression of PPARγ increases the absorption of sugar in the body and induces differentiation of adipocytes. In addition, apolipoprotein E (APOE) and fatty acid binding involved in fat metabolism during adipocyte differentiation.

Protein 4 (Fabp 4), adipsin and stearoyl-coenzyme A desaturase 1 (Scd 1) are known to be regulated by transcriptional regulators such as PPARγ. In order to do this, the expression level of these genes was analyzed.

As a result, as can be seen in FIGS. 10 and 11 , in the case of APOE, it was confirmed that the expression was reduced to less than 30 to 40% when treated with pure soybean water and soybean soaked water compared to the control group. The expression of Scd1 gene was increased when differentiation was induced, compared to undifferentiated, but after treatment with soybean pure water and soybean steep water, the expression level of the gene was confirmed to be 50 to 85%, respectively, depending on the treatment concentration compared to the control group.

On the other hand, it was confirmed that mRNA expression of Fabp 4 did not significantly change gene expression when treated with soybean pure water and soybean steep water compared with the control group. In addition, adipsin is mainly identified in cultured adipocytes as a serine protease that promotes complement peptide by stimulating triglyceride acylation. It is known that expression is induced in relation to energy storage signals during differentiation induction. . It was confirmed that the expression of adipsin in adipogenic cells was inhibited in a concentration-dependent manner when treated with soybean pure water and soybean steep water. decreased. When the soybean steepwater was treated, the expression of adipsin was decreased to 45% and less than 28% at 100 ppm concentration, respectively, in a concentration-dependent manner.

The above result of suppressing the mRNA expression of some genes among the fat metabolism-related genes is also the result of confirming that soybean pure water and soybean steep water regulate the fat metabolism process to regulate the differentiation of adipocytes.

(3) Effect of soybean processing by-products on adipokine

It has been found that adipocytes can induce various adult diseases such as obesity and diabetes through the synthesis and storage of triglycerides as well as the secretion of various hormones and cytokines. As adipokines secreted from these adipocytes, resistin (resistin, RETN), adiponectin (ADIPOQ), and the like are known.

[0106] The resistin (resistin, RETN) is present in the cytoplasm of fat and is only expressed in adipose tissue, and it is known that the expression is reduced by thialidinedione (TZD), a synthetic ligand of PPARγ. Therefore, the expression level of these adipokines was analyzed.

As a result, as can be seen in FIG. 12 , the expression of resistin (RETN) expressed upon induction of adipocyte differentiation did not show a significant difference in expression when treated with soybean by-products compared to the adipocyte differentiation induction control.

On the other hand, when adiponectin (ADIPOQ) was treated with soybean steep water and soybean pure water, it was confirmed that mRNA expression was rapidly decreased in all treatment concentration conditions.

From the above results, it was confirmed that soybean steep water and soybean puree were involved in the regulation of energy metabolism related to adipogenesis in adipocytes by decreasing the secretion of adiponectin (ADIPOQ) among adipokines, and inducing adipogenesis. Suppression and support of adipocyte differentiation during differentiation from pre-adipocytes to adipocytes by substance treatment

It was confirmed that the release generation was suppressed.

In addition, it suppressed the expression of genes such as adipsin, Fabp4, and adiponectin (ADIPOQ) that regulate fat metabolism and energy metabolism during adipocyte differentiation. The treatment of soybean processing by-products is consistent with the results of Oil-Red O dyeing according to the content level.

It was confirmed that it is involved in some regulation in the lipogenesis step.

In addition, in view of the results of soybean processing by-products decreasing the expression level of PPARγ, the inhibition of adipocyte differentiation induction of soybean processing by-products is mainly through inhibition of differentiation-inducing transcription factors, and at the same time has an effect of inhibiting adipogenesis in adipocytes. could check

In conclusion, from the above results, adipocyte differentiation and adipogenesis inhibition by soybean processing by-products are regulated through the expression of genes related to fat differentiation and fat metabolism regulation. It was confirmed that it could be used.

Claims (9)

A composition for preventing or treating obesity, comprising, as an active ingredient, a by-product of soybean processing. The method of claim 1,
The soybean processing by-product is a composition for preventing or treating obesity, characterized in that it is soybean soaking water, soybean curd whey, or a mixture thereof. 3. The method of claim 2,
The soybean immersion water is a composition for preventing or treating obesity, characterized in that it is obtained by immersing soybeans in water at a temperature of 10 to 30° C. for 3 to 24 hours. 3. The method of claim 1 or 2,
The active ingredient is a composition for preventing or treating obesity, characterized in that 0001 to 30% by weight of the total composition. 5. The method of claim 4,
The active ingredient inhibits fat degeneration and fat accumulation in the liver and epididymal tissue; inhibition of the accumulation of cholesterol in the body; inhibition of accumulation of triglycerides, differentiation of pre-adipocytes into adipocytes, proliferation of adipocytes; inhibition of adipogenesis; inhibition of the activity of acetyl-CoA carboxylase (ACC); and an increase in activity of AMP-induced activated protein kinase (AMPK, AMP-activated protein kinase); a composition for preventing or treating obesity, characterized in that it is performed by at least one action selected from the group consisting of: 6. The method of claim 5,
The active ingredient is a composition for preventing or treating obesity, characterized in that it inhibits the activity of peroxisome proliferator activated receptor gamma (PPARγ, peroxisome proliferator activated receptor). 3. The method of claim 1 or 2,
The composition is a composition for preventing or treating obesity, characterized in that it comprises one or more pharmaceutically acceptable carriers or excipients. A health supplement for preventing or treating obesity comprising the composition according to claim 1 or 2. 9. The method of claim 8,
The health supplement is a health supplement for the prevention or treatment of obesity, characterized in that at least one selected from drinks, caramel, chocolate, diet bar formulation, and confectionery.

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