KR20160007744A - Composition for anti-hangover and anti-obesity containing extract of Mytilus edulis - Google Patents

Abstract

The present invention relates to a functional composition for hangover cures and antiobesity including an extract of Mytilus edulis as an active ingredient. The purpose of the present invention is to provide the functional composition for hangover cures including the extract of Mytilus edulis. Other purpose of the present invention is to provide a composition for preventing and treating obesity including the extract of Mytilus edulis. A food composition for hangover cures contains a whole blue mussel extract or a blue mussel flesh extract of Mytilus edulis for hangover cures. The present invention is prepared by the following steps: producing the extract of Mytilus edulis; testing the antioxidative activity, ACE inhibition activity, and ADH and ALDH acitvities of the extract of Mytilus edulis obtained from the previous step; analyzing the alcoholysis ability and antioxidative enzyme of liver tissues of an animal which has taken the extract of Mytilus edulis; and comparing and evaluating the adipocytic differentiation suppressive activity of the extract of Mytilus edulis. The present invention has an effect of providing a new health supplement food for hangover cures, and an excellent effect of providing a composition having excellent obesity preventing and treating effects at the same time.

Description

[0001] The present invention relates to an anti-hangover and anti-obesity containing extract of Mytilus edulis,

The present invention relates to a hangover resolution and an anti-obesity functional composition comprising a mushroom extract as an active ingredient.

Mussel, Mytilus , which belongs to the mussel family, edulis ) is a Korean wave. It secures a sticky substance to dozens of beards called 足 糸 on rocks between 20m depths in the intertidal zone and fixes on the seashore, Eat the nutrients. The inside of the shell is called "Pearl mussel" because it is said to be pearly light, and it is also called "Marsh mussel" or "Mediterranean mussel" by the Mediterranean.

In general, mussels are known to be good for relieving hangovers, but the effect of mussels on alcohololysis enzymes has not been actually disclosed. Korean Patent No. 10-1122586 discloses a hangover-eliminating beverage containing abalone freeze-dried meat, and a food composition for improving and preventing liver disease containing shellfish extract is disclosed in Korean Patent Registration No. 10-0841476, Korean Patent Laid-Open No. 10-2009-0007676 discloses a health food for hangover seafood using or added with soybean sprouts, sweet potatoes, glutinous rice, barley, and plum extract. The hangover resolution activity of the mussel of the present invention is not implied or disclosed anywhere in the known literature.

It is therefore an object of the present invention to provide a hangover resolution functional composition comprising a mushroom extract.

Another object of the present invention is to provide a composition for preventing and treating obesity, which comprises a mushroom extract.

The above object of the present invention can be achieved by a method for producing a mussels extract of the present invention; Testing the antioxidant activity, ACE inhibitory activity, ADH and ALDH activity of the pearl mussel extract obtained in the above step; Analyzing the alcohol degrading ability and antioxidant enzyme of an animal liver tissue ingested with the mushroom extract of the present invention; And comparing and evaluating the activity of inhibiting adipocyte differentiation of pearl mussel extract.

INDUSTRIAL APPLICABILITY The present invention has an effect of providing a novel hangover safe health supplement, and at the same time, it has an excellent effect of being able to supply a composition excellent in prevention and treatment of obesity.

1 is a graph showing DPPH radical scavenging activity of a manganese hot water extract (BMF: mussel, BMW: p. Mussel).
2 is a graph showing an ACE (Angiotensin converting enzyme) inhibitory activity of the manganese hot-water extract.
FIG. 3 is a graph showing the effect of pearl mussel hot-water extract on ALDH (A) and ADH (B) activity.
Fig. 4 is a graph showing the activity of liver tissue alcoholase in experimental animals ingested with rhizome. Fig. NC: Negative control, A: Alcohol intake, A-BM: Alcohol + 5% boiled mussels, A-MS: Alcohol + 1% mussels.
FIG. 5 is a graph showing the activities of hepatic antioxidant enzymes catalase (A) and GSH-Px (B) in an experimental animal ingested with rhizome.
FIG. 6 is a graph showing DNA damage of hepatic tissue cells of an experimental animal ingesting a rhizome.
FIG. 7 is a graph showing cytotoxicity of 3T3-L1 cells of pearl moxibustion extract. FIG.
FIG. 8 is a flow chart showing the treatment period of pearl-marijuana extract and 3T3-L1 adipocyte differentiation.
Fig. 9 is a graph showing the activity of inhibiting adipocyte differentiation of pearl-marijuana extract.
Fig. 10 is a photograph showing changes in the number and size of lipid sphere according to the treatment of pearl-marijuana extract.
FIG. 11 is a flowchart showing a treatment period of pearl-marijuana extract and 3T3-L1 adipocyte differentiation.
FIG. 12 is a graph showing the effect of the treatment of the mulberry juice extract on adipocyte differentiation.
Fig. 13 is a photograph showing the effect of pearl mugwort water extract treatment on the number and size of lipid sphere.
14 is a graph showing a mechanism of inhibiting adipocyte differentiation of pearl moxibustion extract.

The present invention provides a hangover decomposing composition comprising 1 to 10% by weight of a mussel-free mussel or mussel extract of pearl mussel.

The present invention provides a pharmaceutical composition containing the above-described hangover remedy composition as an active ingredient and having a hangover resolution effect due to drinking.

The present invention provides a health supplement containing the above-described hangover remedy composition as an active ingredient and having a hangover resolution effect due to drinking.

The spoilage composition of the present invention comprises 1 to 10% by weight, preferably 5% by weight, of a mussel-free mussel or mussel extract.

The pharmaceutical composition of the present invention can be prepared into a conventional pharmaceutical formulation by selecting one or two or more pharmaceutically acceptable conventional carriers and one or more additives to the effective amount of the hangover decomposing composition as a main component .

The carrier may be selected from one or more selected from among diluents, lubricants, binders, disintegrants, sweeteners, stabilizers and preservatives. Examples of the additives include one or more of flavorings, vitamins and antioxidants The pharmaceutically acceptable composition of the present invention may be formulated into tablets, pills, tablets, capsules, soft capsules, solutions, powders, granules and the like and then taken orally.

In the present invention, any of the pharmaceutically acceptable carriers and additives may be used. Specific examples of the diluent include lactose monohydrate, cornstarch, soybean oil, microcrystalline cellulose, Magnesium stearate or talc is preferable as a lubricant and polyvinypyrrolidone (PVP) or hydroxypropylcellulose (HPC) is used as a binder. Is preferable. The disintegrant may be selected from carboxymethylcellulose calcium (Ca-CMC), sodium starch glycolate, polacrylin potassium or cross-linked polyvinylpyrrolidone And the sweetener is selected from the group consisting of white sugar, fructose, sorbitol or aspartame. Examples of the stabilizer include sodium carboxymethylcellulose sodium (Na-CMC), beta-cryclodextrin, White bee's wax or xanthan gum and preservatives include methyl p-hyroxy benzoate, methylparaben, propyl p-hyroxy benzoate, propylparaben or potassium sorbate (potassium sorbate).

The health supplement food containing the composition for hangover of the present invention can be used as a formulation for folk remedies such as tea, jelly, juice, juice, The health supplements of the invention are excellent in detoxification action without adverse effects on the human body, and are easy to take, which can be useful for relieving hangover before and after drinking.

Hereinafter, the present invention will be described in detail with reference to examples.

<Pearl mussel extract in vitro  Hangover elimination test>

Disclosure material

We received the fresh juice from the Masan Fisheries Cooperative in May 2013. L-histidine-L-leucine (HHL), captopril, alcohol dehydrogenase (ADH), acetaldehyde dehydrogenase (ALDH) ascorbic acid, and NAD + were purchased from Sigma Chemical Co. (St. Louis, Mo., USA), and Hepos was purchased from commercial pharmacy.

Example : Manufacture of Pearl Mushroom Extract According to the Invention

The tap water was removed from the shell of the pearl mussels and the mussels were divided into a mussel and a mussel with only the edible part (meat) removed from the skin. Three times more purified water was added based on the weight of whole mussel (skin, meat) and extracted at 100 ℃ for 30 minutes. 1.00 kg of mussel was obtained from whole mussel 3.75 kg. Thus, hot water extract was prepared by adding purified water based on the weight of whole mussel using this ratio. Then, the extract was filtered with Whatman No.3 filter paper and lyophilized. The dried powder was diluted to 10, 20 and 30 mg / mL with distilled water and used as a test material in the experiment.

Experimental Example  1: Antioxidant activity of the extract of the mushroom of the present invention

The DPPH radical scavenging activity was measured by using the method of Jeong et al., And 0.9 mL of 0.041 mM DPPH solution was added to 0.1 mL of the sample, followed by reaction at room temperature for 30 minutes, and the absorbance was measured at 517 nm. The DPPH radical scavenging activity of each sample was calculated by the following formula and expressed as a percentage.

1, DPPH radical scavenging activity of blue mussel flesh (BMF) and whole blue mussel (BMW) hydrothermal extract was 36.81% and 30.81% at 10 mg / mL of pear mussels, respectively Respectively. The mussel extract showed higher antioxidant activity than the mussel hydrothermal extract and increased in concentration dependent manner.

Experimental Example  2: Invention of pearl mussel extract of the present invention ACE  Inhibitory activity

ACE inhibitory activity was measured according to the method of Cushman and Cheung. To the 20 mL of 50 mM sodium borate buffer (pH 8.3), 1 g of rabbit lung acetone powder was stirred at 4 ° C for 24 hours and then centrifuged for 10 minutes at 4 ° C and 5,000 × g to obtain an ACE enzyme solution. 50 μL of each diluted sample was added, and preliminary reaction was carried out at 37 ° C. for 10 minutes. Then, 100 μL of 25 mM HHL was added thereto and reacted at 37 ° C. for 60 minutes. The reaction was stopped by adding 250 μL of 1 M HCl. The reaction was stopped by adding 1.5 mL of ethyl acetate, stirring for 30 seconds, and centrifugation (5,000 × g, 10 minutes, 4 ° C.) to obtain 1 mL of supernatant. The supernatant was completely dried at 80 ° C for 30 minutes, and 1 ml of distilled water was added thereto, and the absorbance was measured at 228 nm. As a control, 50 μL of extraction solvent was used and the ACE inhibitory activity was calculated using the following equation.

As shown in FIG. 2, the mussel and mussel extracts of the mussel and mussel showed 50.27% and 28.99% inhibitory activity at 30 mg / mL, respectively. As the concentration increased, ACE inhibitory activity also increased. The mussel extract showed higher ACE inhibitory activity than the mussel extract.

Experimental Example  3: Invention of Pearl mussel extract of the present invention ADH ( alcohol dehydrogenase ) activation

The effects of pearl mussel extract on ADH activity were measured by Bergmeyer 's method. The reaction solution was prepared by adding 1.4 mL of distilled water, 0.75 mL of 1.0 M Tris-HCl buffer (pH 8.8), 0.3 mL of 20 mM NAD +, 0.3 mL of 0.2 M ethanol, and 0.1 mL of the sample, followed by reaction at 25 ° C in a constant temperature water bath for 10 minutes Add 0.15 mL of ADH (5 units / mL) to adjust the reaction solution to 3 mL, and preliminarily react at 25 ° C for 5 minutes. Absorbance of the resultant NADH was measured at 340 nm for 5 minutes using a spectrophotometer. The relative activities of the compounds were compared. The negative control was the addition of distilled water instead of the sample, and the positive control was Hepos. The ADH activity was expressed as the ratio of the maximum absorbance at the end of the reaction to the maximum absorbance of the control, and was calculated by the following equation.

The reaction mixture consisted of 0.1 mL of distilled water, 0.3 mL of 1.0 M Tris-HCl buffer (pH 8.0), 0.1 mL of 20 mM NAD +, 0.1 mL of 0.1 M acetaldehyde, 0.1 mL of 3.0 M KCl, 0.1 mL of 0.33 M 2-mercaptoethanol, mL, and reacted at 25 ° C for 10 minutes. After adding 0.1 mL of ALDH (1 unit / mL), the reaction solution was adjusted to 3 mL, placed in a cuvette, preliminarily reacted at 25 ° C for 5 minutes, Formation was measured by a change in absorbance at 340 nm. Negative control and positive control activities were calculated using the same method as ADH activity measurement.

As shown in FIG. 3, ADH activity was increased in a concentration-dependent manner in the manganese hydrothermal extract and 220.25% and 219.10% at 30 mg / mL of mussel and mussel extracts, respectively. ALDH activity also increased in a concentration dependent manner. At 30 mg / mL concentration, mussel and mussel hot water extracts induced 142.96% and 162.10%, respectively.

Statistical analysis

All experiments were repeated 3 times or more, and the mean values were processed according to the method of General Linear Model using SPSS software (Ver. 12). One-way ANOVA was performed to determine the significance of the difference between the mean and the mean ± standard deviation. Differences in all treatment values were analyzed by comparing 95% confidence level (p <0.05).

<Pearl mussel extract in vivo  Hangover elimination test>

Disclosure material

The samples used for the diet were 11.25 liters of water and 1 hour of heating at 100 ° C for 30 minutes to 1 kg of live pear mussels. After solely freezing the solids except for the shell, 5% of the amount was added to the diet, The pearl mackerel was prepared by adding 30 L of water to 10 kg of shell mussel, heating it at 100 ° C for 30 minutes, filtering it with Whatman no. 3, and lyophilizing it to add 1% to the diet.

Experimental animal

Five weeks old Wistar rats were allowed to adapt to the laboratory environment for one week, and then divided into four groups according to the nude method. Each group was divided into two groups: an isocaloric dextrin (NC), an alcohol-treated group (A), an alcohol + 5% pearl mushroom solid-treated group (A-BM) and an alcohol + 1% pearl mine-fed group (A-MS). The diets were mixed with mushroom solid and broth powder according to AIN-93. In the alcohol-treated group, 6 g / kg / day of alcohol was mixed with drinking water. All feeds were kept at 4 ° C and fed for 6 weeks. The animals were sacrificed after fasting for 12 hours, and blood and liver were collected and used for analysis. The dietary composition intake for 6 weeks is shown in [Table 2].

Experimental Example  4: Change of Animal by Pearl Mushroom Extract According to the Invention

The effects of pearl mussel solids and broth on body weight gain, dietary intake, and dietary efficiency after 6 weeks of alcohol consumption and diet supplemented with pearl mussel solids and broth were investigated. In all groups, weight gain, dietary intake, There was no significant difference between the two groups.

Table 6 shows the changes in long-term weights after ingestion of 6-week alcoholic beverages and dietary supplementation of pearl mussel solids and broth. As a result, there was no significant difference between groups in all organs.

Experimental Example  5: Pearl mussel extract in vivo  Alcohololytic activity

The liver was mixed with 0.25M sucrose and homogenized at 4 ° C using a homogenizer. Then, the cytosol portion of the liver was separated from the supernatant by centrifugation at 600g for 10 minutes and then at 10,000g for 20 minutes. Subsequently, 0.1 mL of the supernatant was mixed with 0.2 mL of ethanol, 0.02 mL of 0.5 M semicarbazide, and 2.0 mL of preincubated reaction solution containing 0.1 M NAD and reacted at 37 ° C to measure absorbance at 340 nm. The activity was expressed in nmoles of NADH produced by 1 mg of protein.

As shown in FIG. 4, the ADH of the liver was the highest in group A, and that of the two groups in which the mussel solid and mussel broth were fed Similar to NC, ADH was decreased.

Experimental Example  6: Pearl mussel extract in vivo  Antioxidant activity

A. Catalase Enzyme Activity: Liver tissues were homogenized using a homogenizer, centrifuged at 600 g for 10 min, centrifuged at 1000 g for 20 min, centrifuged at 1000 g for 20 min, treated with 1 × RBC Washed and used in the experiment. For the experiment, 0.05 mL of pellet and 0.3 mL of 30 mM hydrogen peroxide were added to 0.6 mL of 50 mM Na-K phosphate buffer (pH 7), and the decrease in hydrogen peroxide was measured at 240 nm for 30 seconds. Activity was expressed in nmoles of hydrogen peroxide per hour in 1 mg of protein.

B. GSH-Px (glutathione peroxidase) enzyme activity: The liver tissue was homogenized using a homogenizer (IKA® T10 basic, IKA Laboratory Equipment, Staufen, Germany) and centrifuged at 10,000 g for 20 min. Respectively. To 0.605 mL of sample solution (0.15 mL of 250 mM KP buffer (pH 7), 0.075 mL of 10 mM EDTA, 0.075 mL of 10 mM NaN, 0.075 mL of 10 mM GSH, 0.075 mL of 2 mM NADPH, 0.15 mL of DW, and 0.00501 mL) 0.025 mL was added, and the mixture was allowed to stand at room temperature for 5 minutes. After 5 minutes, 0.025 mL of 2.5 mM H 2 O 2 was added and the decrease in NADPH was measured at 340 nm for 70 minutes. Activity was expressed in nmole / min / protein of NADPH produced per 1 mg of protein per hour.

As shown in FIG. 5, the antioxidant enzyme activities in the liver tissues were significantly increased after ingesting the diets supplemented with the pearl mussel solids and the pearl mussels in the catalase than in the alcohol groups. However, GSH-Px, which plays a role in converting H ₂ O ₂ to H ₂ O in liver tissues, was more active than the alcohol group in the group fed the pearl-soaked solid and the pearl-marinated meats.

Experimental Example  7: Expression of Hepatocyte Cells by Pearl Mushroom Extract DNA  Damage degree

0.5 g of liver tissue was extracted and treated with 300 units of collagenase in HBSS buffer, followed by treatment at 37 ° C for 30 minutes. After centrifugation at 40 g for 5 minutes, the supernatant was separated and centrifuged at 700 g for 10 minutes to separate the cell lysate. The cell lysate was separated from the supernatant by a fully frosted slide precoated with 1% normal melting agarose (NMA) And LMA were dispersed evenly and then covered with cover glass and stored in a refrigerator at 4 ° C.

When the gel stiffened for electrophoresis, one double layer of LMA was loosened and DNA double strand was loosened during 1 hour immersion in cold Lysis solution. The slides are then placed in an electrophoresis bath, unwound in electrophoresis buffer for 20 minutes, and electrophoresed at 25 V / 300 mA for 20 minutes. After electrophoresis, the cells were washed thoroughly with Tris buffer, stained with ethidium bromide (20 μL / mL), observed with a fluorescence microscope, and analyzed by computer.

As shown in Figure 6, DNA damage is the% Tail DNA of all alcohol groups with significantly be treated as not treated with _{H 2 O 2 200μM H 2 O} 2 was high. In addition, DNA damage of hepatic tissues was significantly lower than that of alcohol group, as% Tail DNA of pearl mussel solid and broth group was significantly lower than NC level.

Statistical analysis

The mean ± standard deviation (SE) was calculated according to each item, and the significance difference between the two groups was verified using Tukey's multiple range test.

<Test for suppressing obesity of pearl mussel extract>

Disclosure material

Fetal bovine serum (FBS) and Fetal calf serum (FCS) were purchased from Gibco BRL. Dulbecco's modified Eagle's medium (DMEM) and penicillin-streptomycin cocktail were purchased from WELGENE Inc. in Carlsbad, CA, USA. (Daegu, Korea). TRIzol® and SuperScriptII kit were purchased from Oil Red O, 3- (4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT), U0126, SP600125, LY294002, SB431542, The thermal cycler for the polymerase chain reaction (PCR) was a Bio-Rad (Tokyo, Japan) microplate reader (Molecular Devices, Sunnyvale, CA, USA) and the IWR-1 was purchased from Sigma Japan) were used.

Experimental Example  8: Cytotoxicity of the extract of the mussel of the present invention

3T3-L1 preadipocytes were pre-cultured in 12-well culture plates and cultured in 3T3-L1 pre-adipocytes with 10, 50, 100, 200, 400, 600 and 800 μg / And treated with 0.2 mg / mL of MTT solution for 1 hour. The DMEM medium was then removed and the treated 3T3-L1 preadipocytes were washed with 1 mL of phosphate buffered saline (PBS, pH 7.4). MTT formazan, which is not soluble in aqueous solution, was dissolved in 400 μL of dimethyl sulfoxide (DMSO), and the absorbance was measured at 595 nm using a microplate reader.

As shown in FIG. 7, significant cell death was observed at a concentration of 600 and 800 μg / mL compared to the control (control) without any treatment, but concentrations of 10, 50, 100, 200 and 400 μg / mL The extract of pearl mackerel was not toxic to 3T3-L1 preadipocytes. In order to exclude the effect on cytotoxicity in the future, the pearl mushroom extract of 400 μg / mL or less was selectively used.

Experimental Example  9: Inhibition activity of adipocyte differentiation of pearl mackerel juice extract of the present invention

Mouse-derived 3T3-L1 cells were purchased from the American Type Culture Collection (ATCC; Rockville, Md., USA). 3T3-L1 preadipocytes were cultured in DMEM medium containing 10% FCS and 100 unit / mL of penicillin-streptomycin at 37 ° C and 5% CO ₂ . To differentiate 3T3-L1 preadipocytes into adipocytes, the adipocytes were replaced with differentiation induction medium two days after becoming confluent, and the day of substitution with differentiation medium (DM) was designated as Day 0 . The differentiation induction medium was prepared to include 500 μM 3-isobutyl-1-methylxanthine (IBMX), 5.2 μM dexamethasone, and 167 nM insulin in DMEM medium containing 10% FBS and 100 unit / mL penicillin-streptomycin Respectively. Differentiation induction medium was used from Day 0 to Day 2 for 48 hours. Post-differentiation medium (Post-DM) containing only 167 nM of insulin was used for DMEM from day 2 to day 4, and DMEM was used for Day 7 in Day 4 after that. Adipocyte differentiation was terminated on Day 7 (Fig. 8) and ORO staining was performed. The differentiated 3T3-L1 adipocytes (Day 7) were fixed with 3.7% (v / v) formaldehyde for 30 minutes at room temperature, stained with 3 mg / mL ORO for 1 hour at room temperature, I washed it. Images were obtained from a stained 12-well plate using a scanner. For ORO quantitative analysis, 200 μL of DMSO was added to the stained 12-well plate, and the intracellular ORO was dissolved. The ORO was transferred to a 96-well plate at a rate of 50 μL / well and the absorbance was measured at 517 nm using a microplate reader. Lipid accumulation (% of control) was calculated by using differentiated fat cells not treated with pearl marijuana extract as a control.

As shown in Fig. 9, the pearl marijuana extract significantly inhibited adipocyte differentiation of 3T3-L1 cells in a concentration-dependent manner. The inhibition of adipocyte differentiation by 22.61%, 44.42% and 84.20%, respectively, compared with the differentiation rate (100%) of the untreated adipocytes in the 3T3-L1 cells treated with 100, 200 and 400 μg / .

Experimental Example  10: by pearl mackerel juice extract Local district  change

The number and size of lipid droplets in differentiated 3T3-L1 adipocytes were qualitatively analyzed by microscopic observation. In the same manner as above, 3T3-L1 preadipocytes were treated with 100, 200, and 400 μg / mL of pearl marijuana extracts from Day -2 to Day 7, and adipocyte differentiation was performed in the same manner as above Respectively. 3T3-L1 adipocytes of differentiated day 7 were fixed in 3.7% (v / v) formaldehyde for 1 hour at room temperature, and then filled with PBS to prevent drying and observed with a microscope.

As shown in Fig. 9, the pear mussels extract decreased the size and number of fat spheres in a concentration-dependent manner compared with 3T3-L1 adipose cells that were not treated with anything.

Experimental Example  11: Inhibitory activity of adipocyte differentiation according to differentiation time

In order to investigate the mechanism of inhibition of adipocyte differentiation by 3T3-L1 cells, the adipocyte differentiation period (pre-A differentiation, early stage of B adipocyte differentiation, mid-C adipocyte differentiation stage, The mugwort extract was treated at concentrations of 100, 200 and 400 μg / mL. Adipocyte differentiation of 3T3-L1 cells was carried out in the same manner as described above and ended on Day 7 (Fig. 10).

Experimental results showed that the treatment of pearl-marijuana extract before induction of differentiation did not significantly affect adipocyte differentiation of 3T3-L1 cells. In addition, the extract of pearl mackerel showed inhibitory activity in early, middle and late stages of differentiation of adipocytes. Especially, in the early and middle stage of adipocyte differentiation, (Figs. 11 and 12).

Experimental Example  12: According to the present invention, Anti-obesity  Mechanism

On Day 7, total RNA was extracted with 200 μL of TRIzol®. 100 μL of chloroform was added to the extracted total RNA, shaken, allowed to stand at room temperature for 15 seconds, and centrifuged at 17,000 rpm for 20 minutes to separate the supernatant. 250 μL of isopropanol was added to the obtained supernatant, and the mixture was further incubated at room temperature for 10 minutes. Then, RNA was isolated by centrifugation at 17,000 rpm for 15 minutes. The isolated RNA was quantified and cDNA was synthesized at 48 ° C for 1 hour using SuperScriptII kit (Invitrogen). The synthesized cDNA was subjected to PCR using a thermal cycler (Bio-Rad) and electrophoresed on a 1% (w / v) agarose gel to analyze mRNA gene expression levels. The primer sequences of C / EBPβ, PPARγ, C / EPBα, fatty acid synthase (FAS) and β-actin used in this experiment are as follows. C / EBP? Forward: 5'-GCAAGAGCCGCGACAAG-3 ', reverse: 5'-GGCTCGGGCAGCTGCTT-3'; PPAR? Forward: 5'-CCATTCTGGCCCACCAACTT-3 ', reverse: 5'-CCTTCTCGGCCTGTCGATCC-3'; C / EPB? Forward: 5'-CCAGAGGATGGTTTCGGGTC-3 ', reverse: 5'-TCCCCAACACCTAAGTCCCT-3'; FAS forward: 5'-GTGCACACAGTGCTCAAAGG-3 ', reverese: 5'-GGTATAGACGACGGGCACAG-3'; β-actin forward: 5'-AGGGAAATCGTGCGTGACAT-3 ', reverse: 5'-AGCTCAGTAACAGTCCGCCT-3'

As shown in Fig. 13, the specific inhibitor used in this experiment is as follows. U0126 (ERK signaling system inhibitor), SP600125 (JNK signaling system inhibitor), LY294002 (PI3K signaling system inhibitor), SB431542 (TGF-β signaling system inhibitor), IWR-1 inhibitor). As a result of treatment with 100 and 400 μg / mL of mulberry juice extract, 25.61% and 84.20% of adipocyte differentiation was observed, respectively. The result of the simultaneous treatment of 400 μg / mL of pear mussels and U0126 (10 μM), SP600125 (5 μM), LY294002 (5 μM) and SB431542 (5 μM) specific signaling system specific inhibitors, The inhibitory activity of inhibition of differentiation was 84.20%, 81.14%, 88.99% and 94.59%, respectively, but no significant difference was observed. In contrast, the inhibition rate of 3T3-L1 cells in adipocyte differentiation was 63.63%, compared with 400 μg / mL of pearl-marijuana extract and IWR-1 (10 μM) The inhibitory activity of about 21% was decreased.

Statistical analysis

All data were expressed as mean ± SD. Data were analyzed statistically using Statistical Package for Social Science (SPSS, Chicago, IL, USA). The significance of the mean values was verified at the confidence level p <.05. Each item was subjected to one-way ANOVA and Duncan's test and t-test were used to verify the significance of each interval. p <.05, and significance was expressed as * or # <. 05, ** or ## <.01, and *** or ### <.001.

Claims (8)

Hangover pearl mussel mussel or mussel skin extract. A food composition for hangover use containing the extract of claim 1 as an active ingredient. Mixing mussel mussel or mussel with pearled mussel and purified water, respectively;
Filtering the hot water extract obtained in the above step to separate the supernatant;
Lyophilizing the supernatant obtained in the above step; And
Wherein the dried pearl mite dry powder obtained in the above step is diluted with a carrier or an additive acceptable for food processing. 4. The method according to claim 3, wherein the purified water is mixed with three times the weight of mussel mortar or mussel. 4. The method according to claim 3, wherein the hot water extraction is performed at 90 to 100 占 폚. A dietary supplement as claimed in any one of claims 1 to 3, wherein the active ingredient is selected from the group consisting of: 7. The health supplement according to claim 6, wherein the health supplement is selected from the group consisting of tea, jelly, juice, juice, extract, beverage, and preference food. A pharmaceutical composition for the prevention and treatment of obesity, which comprises as an active ingredient an extract of mussel mussel or mussel.

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