KR101602540B1 - Composition of anti-obesity containing extract of myelophycus simplex and method of making the same - Google Patents

Abstract

The present invention relates to an anti-obesity composition comprising a rock beard extract and a preparation method thereof. More particularly, the present invention relates to an anti-obesity composition comprising a rock beard extract and a rock beard extract which has no side effects and has a high anti-obesity effect, and a process for producing the same.
The composition for anti-obesity comprising the extract of rock beard according to the present invention may contain an extract of rock beard (Myelophycus simplex) and have an anti-obesity effect.

Description

TECHNICAL FIELD The present invention relates to an anti-obesity composition comprising an extract of rosacea and a method for producing the same. BACKGROUND ART [0002]

The present invention relates to an anti-obesity composition comprising a rock beard extract and a preparation method thereof. More particularly, the present invention relates to an anti-obesity composition comprising a rock beard extract and a rock beard extract which has no side effects and has a high anti-obesity effect, and a process for producing the same.

Obesity is a phenomenon in which excess energy is accumulated in body fat due to an imbalance between the energy consumed by food and the energy consumed by physical activity. Generally, obesity means excessive fat tissue in the body. If such obesity persists for a long time, various metabolic diseases such as diabetes, hyperlipemia, heart disease, stroke, arteriosclerosis, fatty liver and adult diseases are induced. Due to excessive caloric intake and lack of exercise, the obesity population is rapidly increasing not only in developed countries but also in Korea, which is becoming a serious social problem.

Obesity is known to be the main cause of excess energy supply, which is caused by the accumulation of fat in the body by inducing an increase in fat cell size and number. In addition, genetic factors, environmental factors by Westernized diet, psychological factors, It is known that various causes such as abnormalities are acting.

There is a multidisciplinary study on the development of obesity drugs worldwide. Drugs for the treatment of obesity can be broadly divided into the inhibition of fat absorption, the promotion of fat decomposition and heat generation, the regulation of appetite and satiety, the inhibition of protein metabolism, and the emotional regulation associated with the ingestion of food. Typical treatments for obesity include Xenical ™, which inhibits fat absorption with orlistat, and Reductil ™, which stimulates the sympathetic nervous system to sibutramine.

However, Xenical ™ has been reported to have adverse effects such as nausea, abdominal pain, vomiting, itching, and liver damage. Reductin ™ has serious side effects such as headache, anorexia, insomnia and constipation as well as serious cardiovascular side effects There has been controversy such as the recent use standard being strengthened. In addition to pharmacotherapy through these obesity drugs, there are also dietary therapies to limit the intake of foods and exercises to increase energy consumption, psychotherapy, behavioral therapy, and surgical therapy.

The preferred method of treating obesity is to promote energy consumption through exercise and to treat obesity drugs with few side effects as the most safe and effective method. However, in the case of obesity treatment drugs, severe side effects such as the above examples of Xenical ™ and Reductil ™ have been reported, and there is no clear reliance on safety.

Therefore, it is required to develop a substance which shows excellent efficacy of anti-obesity against the human body while ensuring 100% safety. Recently, various studies have been made on substances for treating obesity through in vivo fat burning / degradation mechanism and energy metabolism promotion mechanism, and in particular, functional foods based on a material capable of ensuring safety to human body And so on.

Therefore, anti-obesity compositions based on natural extracts having few side effects have attracted attention, and various studies related thereto have been conducted.

In the related art, Prior Art 1 (KR 10-2011-0127454 A, published on Nov. 25, 2011) discloses that an effective ingredient of corn beard extract, shiitake mushroom extract, The present invention relates to a composition for anti-obesity characterized by an increase in body weight due to ingestion, an increase in weight of liver tissue and epididymal adipose tissue, an increase in total cholesterol concentration in plasma, an increase in triglyceride and glucose concentration, Inhibiting the increase of the amount of the antioxidant effect, thereby exhibiting a substantial antiobestic effect.

Prior art 2 (KR 10-2012-0090514A, public date: August 17, 2012) is a spinach freeze-dried powder or extract; A lyophilized powder or extract of at least one of lyophilized powder or extract of at least one of anthocyanin or black garlic, which is a purple sweet potato pigment extract; And a green tea extract powder or an extract. The present invention relates to an antioxidant and anti-obesity composite composition containing a spinach powder or an extract.

However, research on a wide variety of natural products is needed to develop a composition for anti-obesity that has high anti-obesity effect and no side effects.

KR 10-2011-0127454A KR 10-2012-0090514A

The object of the present invention is to provide a method for inhibiting the differentiation of adipocytes by inhibiting the expression of transcription factors in the process of adipogenesis of adipocytes differentiating into adipocytes from adipocytes, Or a pharmaceutically acceptable salt thereof.

Another object of the present invention is to provide a functional product such as an anti-obesity food prepared using the composition for anti-obesity comprising the rock beard extract.

In order to achieve the above object, the composition for anti-obesity including the extract of rock beard according to an embodiment of the present invention includes an extract of Myelophycus simplex and has an anti-obesity effect.

The rock beard extract may be one selected from the group consisting of water, alcohols having 1 to 10 carbon atoms, hexane, chloroform, ethyl acetate, and mixtures thereof.

The rock beard extract may inhibit the differentiation of 3T3-L1 adipose precursor cells into adipocytes.

And the rock beard extract may be contained in an amount of 0.01 to 30% by weight.

The anti-obesity composition containing the rock beard extract may have a viscosity of 1.2 to 100 cP.

The food composition according to another embodiment of the present invention may comprise an anti-obesity composition comprising the rock beard extract.

The pharmaceutical composition according to another embodiment of the present invention may include an anti-obesity composition containing a rock beard extract.

A method for preparing an anti-obesity composition comprising a rock beard extract according to another embodiment of the present invention comprises: drying a rock beard; Any one selected from the group consisting of rock salt and water, the alcohol having 1 to 10 carbon atoms, hexane, chloroform, ethyl acetate, and a mixture thereof is mixed at a weight ratio of 1:10 to 10: 1, An extracting step of extracting from an image; Repeating the extraction step twice to five times with respect to the extract obtained through the extraction step; And concentrating the extract obtained through the repeated extraction step at 35 to 65 DEG C and lyophilizing the concentrated extract to powderize the powdered rock beard extract, wherein the anti-obesity effect Lt; / RTI >

Hereinafter, the present invention will be described in more detail.

An anti-obesity composition comprising a rock beard extract according to an embodiment of the present invention includes an extract of Myelophycus simplex and has an anti-obesity effect.

The scientific name of the rock beard is Myelophycus Simplex Papenfuss , and it belongs to the broad seaweed family. It is a brownish or blackish brown needles and several bundles are gathered to form a bundle. The size is 5 to 15 cm in height and 1 to 2 mm in thickness. The lower part is thinner and shorter.

The shape is brown, needle-shaped, stratified, several bundles are formed, the cortex has long and thin cells arranged in length, the outermost part is small and slender bead-shaped cells, and thick-layered cells contain dark brown matter. The male tissue is rod-shaped, and the innermost male tissue cell is broken into an empty space. Dansil sporangs are short spatulate and buried in a long, stalked paraphysis.

The anti-obesity effect inhibits the differentiation of adipocytes, promotes lipolysis and absorption, inhibits amylase activity, inhibits glucosidase activity, inhibits lipase activity, promotes activity of ediportin, And the like.

The rock beard extract may be prepared by inoculating a rock beard with at least one microorganism such as Bacillus lichenifomis or Aspergillus oryzae.

When the fermented rock beard is used as the rock beard extract according to the fermentation process by inoculation of the microorganisms, the effective ingredient of the anti-obesity activity is increased and the anti-obesity effect can be further enhanced.

The at least one microorganism among the Bacillus lichenifomis or Aspergillus oryzae may be inoculated at 0.0001 to 0.05% (w / v) with respect to the mass of the dried rock beard.

The rock beard extract may be one selected from the group consisting of water, alcohols having 1 to 10 carbon atoms, hexane, chloroform, ethyl acetate, and mixtures thereof.

Preferably, the extract may be an alcohol extract having 1 to 10 carbon atoms, and the rock salt and the alcohol having 1 to 10 carbon atoms may be mixed at a weight ratio of 1:10 to 10: 1. When the alcohol is used, it is useful for extracting an active ingredient of an anti-obesity. When the alcohol is used, it may be suitable for extracting the anti-obesity active ingredient contained in the rock beard by the above-mentioned range.

More preferably, the rock salt and the alcohol having 1 to 10 carbon atoms may be mixed at a weight ratio of 1:10 to 1: 1. In the above range, the yield of the anti-obesity effective ingredient contained in the rock salt must be increased.

The rock beard extract may inhibit the differentiation of 3T3-L1 adipose precursor cells into adipocytes.

The rock beard extract has a high anti-obesity effect because it has excellent effect of inhibiting the differentiation of 3T3-L1 adipocytes into adipocytes.

And the rock beard extract may be contained in an amount of 0.01 to 30% by weight. When the rock beard extract is contained in an amount of less than 0.01% by weight, the anti-obesity effect of the rock beard extract is lowered. When the rock beard extract is contained in an amount of more than 30% by weight, the anti- Therefore, the above range may be suitable for use as a composition having a high anti-obesity effect without side effects according to the present invention.

Preferably, the rock beard extract is contained in an amount of 0.1 to 5% by weight. This is because the anti-obesity effective substance contained in the rock beard extract may have an excellent anti-obesity effect within the above range.

More preferably, the rock beard extract may be contained in an amount of 0.1 to 0.5% by weight. By the above-mentioned range, a high anti-obesity effect can be obtained without side effects. In particular, the above range has an advantage of exhibiting an excellent anti-obesity effect even in a relatively small concentration range.

The anti-obesity composition containing the rock beard extract may have a viscosity of 1.2 to 100 cP. Originally, rock beard is a brown algae plant which is generally not used for edible purposes. When the viscosity range is used, the flavor and texture can be enhanced because of the unique scent of the rock beard. The anti-obesity composition containing the rock beard extract must have a high texture and flavor for the steady-state administration of the composition for anti-obesity. The composition for anti-obesity comprising the rock beard extract may further comprise a thickening agent to adjust the viscosity. The thickening agent is considered to be a food or pharmacologically acceptable thickening agent that can be used by those skilled in the art.

Preferably, the anti-obesity composition comprising the rock beard extract may have a viscosity of 1.2 to 10 cP. According to the above range, the texture and flavor are very good, which can be helpful for steady consumption for a certain period of time.

The food composition according to another embodiment of the present invention may comprise an anti-obesity composition comprising the rock beard extract.

The pharmaceutical composition according to another embodiment of the present invention may include an anti-obesity composition containing a rock beard extract. The pharmaceutical composition may include an anti-obesity composition containing the rock beard extract in a pharmaceutically acceptable range.

A method for preparing an anti-obesity composition comprising a rock beard extract according to another embodiment of the present invention comprises: drying a rock beard; Any one selected from the group consisting of rock salt and water, the alcohol having 1 to 10 carbon atoms, hexane, chloroform, ethyl acetate, and a mixture thereof is mixed at a weight ratio of 1:10 to 10: 1, An extracting step of extracting from an image; Repeating the extraction step twice to five times with respect to the extract obtained through the extraction step; And concentrating the extract obtained through the repeated extraction step at 35 to 65 DEG C and lyophilizing the concentrated extract to powderize the powdered rock beard extract, wherein the anti-obesity effect Lt; / RTI >

When the extract contains an extraction step at 10 to 35 ° C and a concentration step to concentrate at 35 to 65 ° C according to the production method of the rock beard extract, the effective substance against the anti-obesity contained in the rock beard is not destroyed, . ≪ / RTI >

The composition for anti-obesity including the extract of rock beard according to the present invention inhibits the expression of transcription factors in the process of adipogenesis of adipocytes differentiating into adipocytes from adipose precursor cells without adverse effect due to no cytotoxicity, It is possible to inhibit cell differentiation. Thus, a composition having an anti-obesity effect without side effects can be provided.

In addition, a functional product such as an anti-obesity food prepared by using the composition for anti-obesity including the rock beard extract may be provided based on the effect of the composition for anti-obesity including the rock beard extract.

FIG. 1 shows the results of MTT assay for the effect of the rock beard extract on the growth of 3T3-L1 cells.
Figure 2 relates to the inhibitory effect of the rock beard extract on the formation of lipid spheres.
Fig. 3 relates to the effect of inhibiting the differentiation of 3T3-L1 lipo-precursor cells into adipocytes in the rock beard extract.
FIG. 4 shows the inhibitory effect of the rock beard extract on the expression of PPARγ, C / EBPα and C / EBPβ, which are adipogenic transcription factors, in 3T3-L1 adipose precursor cells.

Hereinafter, embodiments of the present invention will be described in detail so that those skilled in the art can easily carry out the present invention. The present invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein.

[Production Example: Method for producing rock beard extract]

According to the method for preparing an anti-obesity composition containing the rock beard extract, methanol extract of rock beard was prepared using dried rock beard and methanol as a solvent.

[Experimental Example: Cytotoxicity and anti-obesity effect test]

1. Experimental material

The PPARγ, C / EBPα, C / EBPβ and actin antibodies used for protein analysis in this experiment were obtained from Santa Cruz Biotechnology Inc. (Santa Cruz, CA, USA) and Cell Signaling Technology (Beverly, MA, USA). Peroxidase-labeled donkey anti-rabbit and peroxidase-labeled sheep anti-mouse immunoglobulin used as secondary antibodies for immunoblotting were purchased from Amersham Life Science Corp. (Arlington Heights, Ill., USA). In addition, insulin, dexamethasone, and IBMX used to differentiate 3T3-L1 adipose precursor cells and Oil Red O used to identify triglyceride production in adipocytes were purchased from Sigma-Aldrich (St. Louis, MO, USA).

2. Cell culture

The 3T3-L1 adipose precursor cells used in the experiments were 90% Dulbecco's Modified EaDCRT Media (DMEM, Gibco-BRL, Grand Island, NY, USA), 10% bovine calf serum (BCS, Gibco BRL) and 1% penicillin And streptomycin (Gibco BRL) at 37 ° C and 5% CO 2. FBS was dissolved at room temperature to inactivate remaining complement components and then heat inactivation (heating in a 50 ° C water bath for 30 minutes). In order to overcome the overcorrection caused by cell proliferation, the growth medium was exchanged every 48 hours to maintain a proper number of cells.

3. MTT assay

MTT assays were performed to determine the range of concentrations to be used in the experiments as well as the cytotoxicity of each substance. 3T3-L1 adipose precursor cells were seeded in 6 well plates for cell culture and cultured to confluent state. MSME was treated at 100, 300 and 500 μg / ml concentration. After 72 hours, 200 μl of tetrazolium bromide salt (MTT, Amresco, Solon, OH, USA) diluted at a concentration of 0.5 mg / ml was added and cultured in a CO2incubator for 2 hours. Then, the medium and the MTT reagent were cleanly removed. DMSO was dispensed in 1 ml aliquots. All of the generated formazan was dissolved in 200 μl aliquots in a 96-well plate. Absorbance was measured at 540 nm using an ELISA reader (Molecular Devices, Sunnyvale, CA, USA). The toxicity of each cell was determined by the average absorbance value of each control and expressed as a percentage of the average absorbance value.

4. induction of adipocyte differentiation

3T3-L1 adipose precursor cells were cultured in confluent 6-well plates to confluent and then replaced with differentiation medium containing 10 μg / ml insulin, 10 μM dexamethasone and 0.5 mM 3-isobutyl-1-methylxanthine (IBMX) Induced differentiation of lipid precursor cells. After 2 days of induction of differentiation, the cells were replaced with medium containing 10 μg / ml insulin every 2 days to promote the differentiation of adipose precursor cells. To confirm the effect of MSME in the course of adipogenesis, MSME was treated together at the beginning of differentiation.

5. Oil Red O staining

Oil Red O staining was performed to confirm the content of fat formed during lipid progenitor cell differentiation. The prepared 3T3-L1 adipose precursor cells were washed with PBS, fixed with 3.7% formalin for 10 minutes, washed with 60% isopropanol, treated with Oil Red O solution, and stained at room temperature for 20 minutes. After dyeing, Oil Red O solution was removed and washed 4 times with PBS. The stained cells were observed using a phase contrast microscope. For quantitative analysis after oil red O staining, fat was extracted with 100% isopropanol, and transferred to a 96-well plate in 200 μl increments. Absorbance was measured at 500 nm using an ELISA reader and expressed as a percentage of the absorbance value of the control group.

6. Western blot analysis

To the 3T3-L1 adipose precursor cells prepared in the same manner, an appropriate amount of lysis buffer (25 mM Tris-Cl (pH 7.5), 250 mM NaCl, 5 mM EDTA, 1% NP-40, 1 mM phenymethylsulfonyl fluoride dithiothreitol (DTT)] was added, reacted at 4 ° C for 1 hour, and centrifuged at 14,000 rpm for 30 minutes to isolate the total protein in the supernatant. The protein concentration of the supernatant was quantitated according to the Bio-Rad protein assay reagent (Bio-Rad, Hercules, Calif., USA) and the method of use, and then mixed with an equal volume of Laemmli sample buffer (Bio-Rad). Samples of the same amount were separated by electrophoresis using sodium dodecyl sulphate (SDS) -polyacrylamide gel and electroblotted with PVDF membrane (Bio-Rad, USA). The PVDF membrane was treated with 5% skim milk to block non-specific proteins, and the primary antibody was treated for 2 hours at room temperature or over night at 4 ° C., and washed with PBS-T (4 times for 10 minutes) and reacted for 1 hour at room temperature using a secondary antibody (diluted 1: 1000 with PBS-T) that corresponds to the treated primary antibody. After the reaction was completed, the Enhanced Chemiluminoesence (ECL) solution (Amersham Life Science Corp.) was applied to the dark room and then exposed to X-ray film to analyze the amount of specific protein expressed.

7. Statistical analysis

All the results were expressed as mean ± standard deviation (SD) using Statistical Package for the Social Sciences (SPSS) statistical program. Statistical significance of the analytical items was verified at the p <0.05 level using Student t-test and Duncan's multiple range test.

[Experimental result: cytotoxicity and anti-obesity effect test]

1. Effect of methanol extract of rock beard on 3T3-L1 cell survival

To investigate the cytotoxicity of methanol extract of rock beard in 3T3-L1 adipocytes, MTT assay experiments were carried out at a concentration of 100 to 700 μg / ml. When methanol extract of rock beard was treated, the degree of change of cell proliferation was calculated as percentage compared with the control, and the methanol extract of rock beard was examined for cytotoxicity. The effect of methanol extract of rock beard on the growth of 3T3-L1 cells was analyzed by MTT assay as shown in Fig. At a concentration of 100 to 500 μg / ml, cell viability of 80% or more was similar. Therefore, the experimental conditions were set up to a concentration of 500 μg / ml which does not significantly affect the survival rate and proliferation (see FIG. 1 below).

2. Effects of methanol extract of rock beard on inhibition of 3T3-L1 cell differentiation

In order to investigate the effect of methanol extract of rock beard on the adipocyte differentiation and triglyceride accumulation of lipid precursor cells, differentiation of lipid precursor cells into mature adipocytes was induced by treatment with rock salt methanol extract. After induction of differentiation, differentiation was made before and after dyeing with Oil Red O and the degree of lipogenesis was observed with a phase contrast microscope. The results are shown in Fig. 2 below.

As shown in FIG. 2, in contrast to the control without induction of differentiation, when the differentiation was induced without treatment of the methanol extract of the rock salt, it was observed that the formation of lipids in the cytoplasm was actively induced, It was confirmed that the formation of spheres was suppressed depending on the treatment concentration.

In addition, the amount of triglyceride was 92%, 80%, and 50%, respectively, when methanol extract of rock beard was treated with methanol extract of rock beard, The results are shown in Fig. 3 below.

Therefore, referring to FIG. 3, it was found that the methanol extract of rock beard was highly effective in inhibiting the differentiation of 3T3-L1 adipocytes into adipocytes.

The adipogenesis process, which is differentiated from adipocyte precursor cells into adipocytes, involves a number of adipogenic transcription factors. C / EBPα is a major transcription factor that regulates C / EBPα and PPARγ. PPARγ and C / EBPα are key transcription factors that regulate adipogenesis. They are highly expressed in the late phase of differentiation and are a terminal marker of adipogenesis including adiponectin and GLUT4 Lt; RTI ID = 0.0 &gt; expression &lt; / RTI &gt; These differentiated cells have characteristic features of adipocytes by inducing specific gene expression along with morphological features such as lipid droplet generation and cell size increase. Therefore, we could confirm at the protein level how rock beard extracts affect the expression of adipogenic transcription factors.

As shown in FIG. 4, when the differentiation inducing factors insulin, dexamethasone and IBMX were treated to induce differentiation, the expression of PPARγ, C / EBPα and C / EBPβ was markedly increased, , The expression of PPARγ, C / EBPα and C / EBPβ decreased at the protein level in the treated group of 500 μg / ml compared to the concentration of 100 μg / ml in the case of treatment with the rock beard extract, and adipogenic transcription inhibited the differentiation into adipocytes by effectively inhibiting the expression of PPARγ, C / EBPα and C / EBPβ, which are factors of lipid droplet and TG.

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the invention is not limited to the disclosed exemplary embodiments, Of the right.

Claims (8)

delete delete delete delete delete delete delete Dry stage to dry rock beard;
Any one selected from the group consisting of rock salt and water, the alcohol having 1 to 10 carbon atoms, hexane, chloroform, ethyl acetate, and a mixture thereof is mixed at a weight ratio of 1:10 to 10: 1, An extracting step of extracting from an image;
Repeating the extraction step twice to five times with respect to the extract obtained through the extraction step;
Concentrating the extract obtained through the repeated extraction step at 35 to 65 占 폚;
Lyophilizing the concentrated extract to powderize the pulverized rock mustard extract; and
And adjusting the viscosity to 1.2 to 100 cP by mixing the powdered extract with water.
A method for preparing an obesity-improving food composition comprising a rock beard extract.

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