KR101908862B1 - Method for manufacturing fermented Cordyceps militaris extract grown on Rhynchosia Nulubilis Soybean and its use for prevention and treatment of non-alcoholic fatty liver disease - Google Patents

Abstract

The present invention relates to a manufacturing method of a fermented extract of Cordyceps militaris grown on Rhynchosia nulubilis, and a pharmaceutical composition for fatty liver suppression comprising the same. The pharmaceutical composition of the present invention does not show cytotoxicity while increasing the expression of fatty acid oxidized genes, and thus can be usefully used for the purpose of treatment of fatty liver.

Description

TECHNICAL FIELD [0001] The present invention relates to a method for preparing a fermented extract of Cordyceps mellon L., and a pharmaceutical composition for inhibiting fatty liver comprising the same,

The present invention relates to a method for producing a fermented extract of Cordyceps mellitus Worticulture having the effect of inhibiting the accumulation of triglycerides by fatty acid oxidation activation against fatty liver disease caused by fat accumulation in the liver and a pharmaceutical composition for inhibiting fatty liver comprising the active ingredient More particularly, the present invention relates to a method for producing a fermented extract of Cordyceps mellitus Wort, which causes increase of sphingosine monophosphate and the increase of sphingosine monophosphate through the activation of sphingophospholipase, and a pharmaceutical composition for inhibiting fatty liver comprising the same .

In the modern society, due to the improvement of living standards, excessive nutrient intake occurs. However, due to lack of exercise and low calorie consumption, accumulation of excess nutrients in the body increases the incidence of various metabolic disorders such as obesity, fatty liver, insulin resistance and cardiovascular disease. Among the adult diseases caused by overeating, fatty liver is caused by accumulation of triglyceride in the liver due to excessive fat and alcohol consumption, increase of triglyceride biosynthesis in the liver, decrease of triglyceride from the liver, Is diagnosed.

Fatty liver disease is classified as alcoholic fatty liver disease caused by excessive alcohol consumption and nonalcoholic fatty liver disease caused by obesity, diabetes, hyperlipidemia, or drug consumption. Nonalcoholic fatty liver disease is an accumulation of triglycerides in the liver, regardless of whether it is alcoholic or not. It appears as steatosis in the early stage and develops as steatohepatitis during development.

Nonalcoholic lipidemia is classified into giant bacilli lipid and small bacilli lipid according to the size of lipid droplet which occurs in hepatocyte pathologically. Most nonalcoholic lipid bacilli are giant bacillary, There is no unusual symptom, but it progresses chronically and the liver function is gradually deteriorated. Nonalcoholic lipidemia is caused by an increase in fatty acid transported from adipose tissue to the liver due to unbalanced energy consumption, and is caused by a decrease in fatty acid oxidation ability and an accumulation of fat in the liver due to an increase in triglyceride biosynthesis. Thus, changes in nuclear receptors such as PPARα and SREBP-1, which regulate the expression of fatty acid oxidation and biosynthesis-related enzymes, play an important role. Fatty fat accumulation in the province of lipid hepatitis develops as the inflammatory response to the deepening of fibrosis (fibrosis) and cirrhosis (cirrhosis) will develop. Therefore, in order to prevent the development of liver cirrhosis from the initial prognosis, lipid-lowering, there is no proper treatment to prevent accumulation in the liver. Given that 50% of patients with alcoholic lipidemia and 30% of those diagnosed with nonalcoholic fatty liver develop into cirrhosis, lipidemia can be considered a very serious liver disease.

Currently, there are few effective medicines that specifically treat fatty liver disease, and exercise and diet therapy are recommended. However, treatment efficiency is very low, and the development of therapeutic agents for fatty liver disease by multinational pharmaceutical companies is continuing. As a result of the reported correlation between insulin resistance and fatty liver observed in diabetes and obesity, metformin or fenofibrate, a PPARα activator, is prescribed for the treatment of fatty liver.

Cordyceps militaris (Cordyceps militaris) is a medicinal mushroom grown by insect infestation, a kind of filamentous fungus, distributed in Korea, Japan, China and Nepal. Cordyceps extracts are effective for anti-inflammatory and lymphatic, leukemic, bladder cancer, anti-cancer activity and immunity control. However, because the wild Chinese caterpillar fungus is rare and it is difficult to collect large quantities in nature, it was cultivated with Rhizopus nulubilis Soybean as a host. Cordyceps militaris grown on Rhynchosia Nulubilis Soybean cultivated in different host cultivars has not yet been clarified in terms of physiological activity as compared with natural harvesting caterpillar fungus growing on insect host.

Korea is consuming a lot of fermented foods such as miso, soy sauce, and kimchi, and traditional fermented foods have been known to have many health effects. In particular, the fermentation of food by lactic acid bacteria increases the physiological activities such as digestibility and natural metabolites, as well as preservation, taste, aroma and texture. Foods fermented with lactic acid bacteria increase the physiologically active substances and increase immunity There are research results. However, the nonalcoholic fatty liver alleviation and therapeutic efficacy of extracts fermented with Lactobacillus acidophilus were not known.

As related related art, U.S. Published Patent Application No. 2013-0259894 (Feb. 21, 2014) discloses a composition for preventing hepatic fibrosis and non-alcoholic fatty liver disease diseases on Cordyceps synensis powder, a Chinese lichen- .

U.S. Published Patent Application No. 2014-08658151 (Feb. 21, 2014)

Food Chem Toxicol. 2013; 60: 52-7 Chin Med. 2009; 4: 12

The present invention has been made in order to meet the above-mentioned demand of the prior art. The present invention relates to a fermented extract of Cordyceps sinensis microflora, which increases the expression of sphingosine phosphorylase and increases fatty acid oxidation in hepatocytes, And a food composition for preventing and alleviating the non-alcoholic fatty liver of the extract and a process for producing the same.

A food composition for prevention and improvement of nonalcoholic fatty liver disease and a method for producing the same according to the first embodiment of the present invention is characterized by a) a method of hot water extraction of caterpillar fungus ciliaris extracts at 105 DEG C for 2 hours with distilled water, Pediococcus pentosaceus ON188), fermented for 24 hours, heated for 10 minutes at a heating temperature of 100 ° C, and sonicated for 3 minutes; And b) extracting the fermented Pseudomonas aeruginosa fermented extract of fermented West Indies with hot water.

According to the method of the present invention for producing a fermented extract of Cordyceps mellitus, the present invention has an effect of increasing the expression of a fatty acid oxidizing gene and showing no cytotoxicity, and thus being useful for the purpose of treatment of fatty liver.

Brief Description of the Drawings Fig. 1 is a bar graph showing the cell survival rate of (A) wild-type Cordyceps mellifera, (B) Seomotai caterpillar fungus (GSC) hot water extract or (C) Seedlings caterpillar fungus extract (GSC-ON188) ego,
2 is a graph showing changes in the expression of the fatty acid oxidized genes of (A) wild-type C. militaris, (B) seaweed, or (GSC) hydrothermal extract or (C) fermented extract of caterpillar fungus (GSC-ON188) A bar graph is shown,
FIG. 3 is a graph showing changes in expression of biosynthetic genes of (A) wild-type C. militaris, (B) Pseudomonas aeruginosa (GSC) hot water extract or (C) Pseudomonas aeruginosa extract (GSC-ON188) (D) a change in S1P concentration in a cell, and (E) a change in protein expression in a cell,
FIG. 4 is a photograph showing fatty acid in the hepatocyte of the present invention to generate intracellular lipid, treating lipid peroxidized microorganism fermented extract, and observing accumulation of lipid by dyeing.
FIG. 5 is a graph showing changes in body weight of a mouse (A) and an intake rate of a dietary composition (B) when the fermented extract of the present invention was mixed with a high fat diet for 4 weeks,
FIG. 6 is a graph showing the results of (A) liver lipid change and (B) fat fat mass in the fat tissue obtained by mixing the fermented extract of the present invention with the high fat diet for 4 weeks, This is the picture shown.

Hereinafter, preferred embodiments of the present invention will be described with reference to the accompanying drawings so that those skilled in the art can easily carry out the present invention. It should be noted that the drawings denoted by the same reference numerals in the drawings denote the same reference numerals whenever possible, in other drawings. In the following description of the present invention, a detailed description of known functions and configurations incorporated herein will be omitted when it may make the subject matter of the present invention rather unclear. And certain features shown in the drawings are to be enlarged or reduced or simplified for ease of explanation, and the drawings and their components are not necessarily drawn to scale. However, those skilled in the art will readily understand these details.

Unless defined otherwise, all terms used herein, including technical or scientific terms, have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Terms such as those defined in commonly used dictionaries are to be interpreted as having a meaning consistent with the contextual meaning of the related art and are to be interpreted as either ideal or overly formal in the sense of the present application Do not.

With reference to the accompanying drawings, preferred embodiments of the present invention will be described in detail. In the following description of the present invention, detailed description of known functions and configurations incorporated herein will be omitted when it may make the subject matter of the present invention rather unclear.

[First Embodiment]

Figure 1 shows the results obtained after 24 hours after treatment of AML-12 hepatocytes with (A) wild-caught C. militaris, (B) hot-water extract of Seomotai caterpillar fungus (GSC) or (C) And the cell viability measured. Results were expressed as a percentage relative to the vehicle-treated control and expressed as mean ㅁ standard error.

FIG. 2 shows the results of treatment of AML-12 hepatocyte with (A) wild-type C. militaris, (B) hot water extract of Seomotai caterpillar fungus (GSC) or (C) fermented caterpillar fungus extract (GSC-ON188) The change in mRNA expression of the oxidized gene, CPT1, was measured by real-time PCR. WY-14643, a PPARα activator, was treated as a positive control. Data were expressed as mean ㅁ standard error ( ^* p <0.05 vs. vehicle control).

FIG. 3 is a graph showing the effect of sphingosine (A) on the AML-12 hepatocyte after 24 hours after (A) wild-caught C. militaris treatment, (B) hot water extract of Seok-Tae Cordyceps (GSC) 1-phosphate (S1P) biosynthesis gene SPHK2 mRNA expression was measured by real-time PCR. GSC-ON188 (D) Changes in S1P concentration in cells after 24-hour treatment and (E) Changes in protein expression of intracellular CPT1, SPHK2 and PPARα. WY-14643, a PPARα activator, was treated as a positive control. Data were expressed as mean ㅁ standard error ( ^* p <0.05 vs. vehicle control).

FIG. 4 shows lipid accumulation of AML-12 hepatocyte treated with oleic acid, which is a fatty acid, and lipid accumulation was observed by Oil Red O staining. WY-14643, a PPARα activator, was treated as a positive control.

FIG. 5 shows (A) mice body weight change and (B) dietary intake when 8 weeks old C57BL6 mice were fed the fermented extracts of P. japonicum microorganism fermented in a high fat diet for 4 weeks. Data were expressed as mean ㅁ standard error ( ^* p <0.05 vs. high-fat diet control group).

FIG. 6 shows the results of (A) liver lipid changes and (B) fat cells in the adipose tissues after 8 weeks old C57BL6 mice, Eosin staining.

According to a first embodiment of the present invention, there is provided a method for producing a fermented soybean curd according to the first embodiment of the present invention, comprising the steps of: a) Heating for 10 minutes and ultrasonication for 3 minutes; And b) extracting the fermented endemic caterpillar fungus fermentation extract with hot water.

The fermentation of step a) may be carried out for 2 to 10 hours, and the extraction of step b) may be carried out for 30 minutes to 5 hours.

According to one aspect of the present invention, there is provided a food composition comprising a fermented hot-water extract of the above-mentioned Cordyceps mellon.

According to one aspect of the present invention, there is provided a food and pharmaceutical composition for the treatment of fatty liver comprising a hydrothermal extract of a fermented extract of Cichoraceae Cordyceps.

The hot-water extract may increase expression of one or more selected from the group consisting of SPHK1, SPHK2, CPT1, ACOX1, and PPARa, may lead to a decrease in lipid droplet in liver tissue, , And may not exhibit cytotoxicity.

The present invention relates to a method for the fermentation of caterpillar fungus; And b) extracting the above-mentioned fermented caterpillar fungus extract with hot water. The fermentation temperature is preferably 37 ° C and the germination time is preferably 48 hours.

The fermentation of step a) can be carried out for 24 to 48 hours and can be preferably carried out by fermentation for 48 hours. The extraction of step b) may be carried out for from 6 hours to 24 hours and may preferably be carried out by heating for 1 hour.

In addition, the present invention provides a food composition comprising the above-mentioned fermented water-extract of Cordyceps mellon.

In addition, the present invention provides a pharmaceutical composition for the treatment of fatty liver comprising fermented extract of Fermented Cordyceps mellifera from Fermented West Indies.

In one embodiment, the pharmaceutical composition may further comprise one or more pharmaceutically acceptable carriers or additives. That is, the pharmaceutical composition of the present invention can be formulated in the form of oral, granule, tablet, capsule, suspension, emulsion, syrup, aerosol or the like oral preparation, external preparation, suppository and sterilized injection solution according to a conventional method . The pharmaceutically acceptable carrier may be selected from the group consisting of lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, Cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil, and the like. It also includes diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrants, surfactants, and the like. Solid form preparations for oral use include tablets, pills, powders, granules, capsules and the like, which may contain at least one excipient such as starch, calcium carbonate, sucrose or lactose lactose, gelatin and the like, and may include lubricants such as magnesium stearate and talc. Oral liquid preparations include suspensions, solutions, emulsions, syrups, and the like, and may contain diluents such as water and liquid paraffin, wetting agents, sweetening agents, fragrances, preservatives and the like. Examples of the non-aqueous solution include sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, freeze-dried preparations and suppositories. Non-aqueous solvents and suspensions include vegetable oils such as propylene glycol, polyethylene glycol and olive oil, ethyl And injectable esters such as oleate. As a suppository base, witepsol, macrogol, tween, cacao butter, laurin, glycerogelatin and the like can be used.

The dosage of the Fusarium oxyspora fermented extract contained in the pharmaceutical composition of the present invention varies depending on the condition and body weight of the patient, the degree of disease, the drug form, the administration route and the period, but can be appropriately selected by those skilled in the art. For example, the fermented extract of Cordyceps mellitus may be administered at a dose of 0.0001 to 1000 mg / kg per day, preferably 0.01 to 1000 mg / kg per day, and the administration may be administered once or several times a day . In addition, the pharmaceutical composition of the present invention may contain 0.001 to 90% by weight of the above-mentioned Cordyceps mellitus fermented extract of the present invention for the total weight of the composition.

The pharmaceutical compositions of the present invention can be administered to mammals such as rats, mice, livestock, humans, and the like in a variety of routes, such as oral, intraperitoneal, rectal or intravenous, intramuscular, subcutaneous, intracerebral, or intracerebroventricular &Lt; / RTI &gt;

In one embodiment, the hot-water extract can increase expression of one or more selected from the group consisting of TNF [alpha], IL-1 [beta] and iNOS and increase expression of one or more selected from the group consisting of pJNK, pERK and pp38 , And may not exhibit cytotoxicity.

Hereinafter, the present invention will be described in more detail through examples and test examples. However, the following examples and test examples are provided for illustrating the present invention, and the scope of the present invention is not limited thereto.

<Examples>

1. Preparation of Seomyeongtae Cordyceps Extract

Cordyceps militaris grown on germinated Rhynchosia nulubilisa, Kucati: 0093) was cultivated in the Cell Activity Research Institute (Seongnam City, Gyeonggi Province).

For the preparation of hot water extract, Seomyeongtae caterpillar fungus was sterilized at 121 ℃ for 15 minutes and used for fermentation. For fermentation, Pediococcus pentosaceus ON188, a lactic acid bacteria derived from onion, was inoculated and fermented at 37 ° C for 48 hours. Next, 100 g of the fermented product of Tae-kwanghaek, Seomyeongtaek was added to 500 ml of hot water and extracted for 24 hours. After the lyophilization, the final extracted powder was stored at -20 ° C until analysis.

2. Cell Culture and Cell Survival Analysis

AML-12 hepatocytes were obtained from ATCC (American Type Culture Collection, Manassas, Va.). Cells is 10% FBS (fetal bovine serum) , 100 U / mL of penicillin and 100 μg / in mL DMEM medium in a high content of glucose (glucose) supplemented with streptomycin (Dulbecco's Modified Eagle's Medium) 5% CO 2 conditions under The cells were inoculated to a 6-well plate at 8 ° C to 10 ⁵ cells under a humid temperature condition of 37 ° C and cultured for 24 hours. Cells were treated with various concentrations of Fusarium oxysporum fenugreek extract and cells were treated with XTT (2,3-Bis- (2-Methoxy-4-Nitro-5-Sulfophenyl) -2H-Tetrazolium-5-Carboxanilide) The absorbance at 450 nm was measured on a spectrophotometer to measure cell viability.

3. RNA isolation and real-time RT-PCR method

Total RNA was isolated from AML-12 hepatocytes using an RNA extraction kit (intron Biotechnology, Korea). cDNA (first-strand cDNA) was synthesized with PCR cyclase using an iScript cDNA biosynthesis kit (BioRad, Hercules, Calif.). The PCR conditions were 5 min at 25 ° C, 30 min at 42 ° C, 5 min at 8 ° C and then kept at 4 ° C. RT-PCR analysis was performed using a primer and a SYBR Green Master Mix (TaKaRa, Japan) in a PCR system (Step-One Plus Real-Time PCR System, Applied Biosystems Ins, Carlsbad, CA). RT-PCR was performed at 95 ° C for 5 minutes, followed by 5 cycles of 95 ° C for 15 minutes, 60 ° C for 15 minutes, 72 ° C for 45 minutes, 95 ° C for 15 minutes, and 60 ° C for 1 minute for 40 cycles Respectively. The sequences of the primers used in the present invention are summarized in Table 1 below. Expression of these genes was normalized on the basis of β-actin.

gene Primer sequence CPT1 Forward CTT CCA AGG CAG AAG AGT GGG Reverse GAA CCT TGG CTG CGG TAA GAC PPARα Forward ATC CAC GAA GCC TAC C Reverse CAC ACC GTA CTT TAG CAA G SPHK2 Forward AGA CGG GCT GCT TTA CGA G Reverse CCT GCT CAA ACC CGC CAT β-actin Forward GAC GTT GAC ATC CGT AAA G Reverse CAG TAA CAG TCC GCC T

4. Sphingosine Monophosphate  How to measure

AML-12 cells were treated with various concentrations of Fusarium oxysporum miltiorrhizae extract and cultured for 24 hours. After the cells were pulverized, the sphingolipids were separated and quantified using a high performance liquid chromatography-mass spectrometer (LC-MS / MS) Respectively.

5. Western blotting analysis method

Cells were homogenized in a lysis buffer containing 10 mM Tris (pH 7.8), 1 mM EDTA, 150 mM NaCl, phosphatase inhibitors and protease inhibitors, and lysates ) Were centrifuged to remove cell lysates and non-lysed cells. The protein concentration was quantified by measuring the absorbance at 750 nm using a protein assay kit (BioRad, Hercules, CA). 30 [mu] g of the protein was separated using a 10% SDS-PAGE gel, and proteins in the gel were transferred to the PDVF membrane. A primary antibody specific for the protein was ligated and a mouse-specific or rabbit-specific secondary antibody was incubated. A view of the protein was visualized using an enhanced chemiluminescence kit (BioRad, Hercules, CA) and a chemiluminescent imaging device (Vilber Lourmat, Sint-Denijs-Westrem, Belgique). Primary antibodies to CPT1, SPHK2, PPARα, β-actin (Cell signaling, Danvers, MA) were used.

6. Mouse animal experiment

8 weeks old C57BL6 mice were fed a 60% kcal high fat diet for 2 weeks and then mixed with 60% high fat diet (50, 100, 200 mg / kg / day, n = 7) 4 weeks. At this time, a 60% kcal high-fat diet was fed with a combination of a normal diet (NC) and fenofibrate (20 mg / kg / day) as a PPARα activator. Body weight and dietary intake were measured weekly during 4 weeks feeding. Liver and adipose tissues were separated from mice after 4 weeks feeding and used for pathological analysis.

7. Pathology Analysis

Liver and adipose tissues from mouse were fixed in formalin for 24 hours. Paraffin blocks were prepared and 5 μm thick sections were stained with hematoxylin and eosin. At this time, lipid accumulation of liver tissue and cell size of adipose tissue were observed under a microscope.

8. Statistical processing

All results were expressed as mean ㅁ standard error (SEM). The test was performed at least three times independently. Results were analyzed using a Student's t-test and were considered statistically significant if the p-value was less than 0.05.

<Test Example>

1. Cell survival rate of Fermented Extract of Cordyceps sinensis

To investigate the cytotoxicity of GSC and GSC-ON188, we investigated whether GSC and ON188 affect cell viability. AML-12 hepatocytes were cultured with 0, 5, 10, 25, 50 and 100 μg / ml of Cordyceps sinensis extract for 24 hours. Cell viability was measured by XTT analysis using a cell proliferation assay kit (Wel gene Inc, Korea). The control group, C. militaris extract and GSC extract, showed no cytotoxicity (Fig. 1, A and B).

The GSC-ON188 extract also did not show cytotoxicity (FIG. 1C). As a result, it can be suggested that the extracts of Seomotai mushroom extract and Seomyeongtae mushroom extract do not show cytotoxicity at all.

2. Increase of expression of fatty acid oxidized gene in fermented extract of Cordyceps mellon

To investigate the effect of fermented extracts of P. vannamei on cell viability and fatty acid oxidation in hepatocytes. AML-12 hepatocytes were treated with C. militaris, GSC and GSC-ON188 extracts at various concentrations, and mRNA was isolated after 24 hours. Control group (control) (Con) and positive control group PPARα activator WY-14643 (WY) were used as the control group. It was observed that CPT1, a fatty acid oxidizing gene, was increased in hepatocytes treated with WY-14643 (FIG. 2).

CPT1 mRNA expression was not increased in C. militaris and GSC-treated hepatocytes (Fig. 2, A and B), whereas GSC-ON188-treated hepatocytes increased CPT1 expression 4-5 times 2C). These results demonstrate that the fermented extracts of P. vannamei, unlike the wild - harvested caterpillar fungus, do not significantly increase fatty acid oxidation in hepatocytes.

Expression of SPHK2, a synthetic enzyme for sphingosine 1-phosphate (S1P), was measured in order to investigate the mechanism by which GSC-ON188 increases fatty acid oxidizing ability.

In the hepatocytes treated with C. militaris and GSC, mRNA expression of SPHK2 was not increased (Fig. 3, A and B), whereas expression of SPHK2 was increased 5-fold in hepatocytes treated with GSC-ON188 3 of C).

In addition, S1P in the cells was observed to be increased by 72% when treated with GSC-ON188 (FIG. 3 D). It was also observed that increased expression of CPT1, SPHK2 and PPARα was induced by GSC-ON188. These results demonstrate that GSC-ON188 increases S1P, a signal transducer that increases fatty acid oxidation in the liver through increased expression of SPHK2, and increases expression of the fatty acid oxidant CPT1 (Hepatology. 2015.62 (1): 135-46).

3. Effect of Fermented Extract of Cordyceps sinensis on the Fat Storage Accumulation in Hepatocytes

In order to investigate whether the increased expression of fatty acid oxidase actually inhibits intracellular fat accumulation, AML-12 hepatocytes were treated with oleic acid fatty acid. In the case of olive acid treatment (+ FA (oleic acid)), olive oil accumulated in the cells was observed as reddish color due to fatty staining rather than control (control). On the other hand, in the cells treated with positive control WY-14643 (+ FA + WY), the lipid was markedly decreased and the lipid was significantly decreased even in the GSC-ON188 (+ FA + ON188) treated cells there was. Therefore, GSC-ON188 contributes to the decrease of intracellular lipid by increasing expression of fatty acid oxidized gene.

4. Effect of Fermented Extract of Cordyceps militaris on the Fatty Liver

To investigate the increase in fatty acid oxidation observed in hepatocytes, male C57BL6 mice (male rats) were fed high-fat diet (HFD) for 2 weeks and then mixed with GSC-ON188 at 50, 100 and 200 mg / kg / day And fed for 4 weeks. (NCD) and a positive control group (PPARα activator, fenofibrate 20 mg / kg / day) were used in the control group. The 4-week feeding diets (NCD) showed significantly lower body weight (100 mg / kg / day) showed no significant difference in body weight between the high fat diet group and the high fat diet group, but 200 mg / kg / day Showed a significant decrease in body weight (Fig. 5A).

The mice receiving fenofibrate showed a significant decrease in body weight, presumably due to a decrease in food intake (Fig. 5B). There was no significant difference in other groups of mice on food intake (Fig. 5B).

When mouse liver tissue was isolated and stained with Hematoxylin & Eosin, high - fat diet group showed fatty accumulation in liver tissue unlike normal diet group, and hepatic cell showed pathological form accumulating fat. However, the control group, the general diet and the fenofibrate group, showed a general hepatic pathology (Fig. 6A).

The liver tissue of GSC-ON188-treated mice showed accumulation in some lipid tissues at the dose of 50 mg / kg / day, but the size was smaller than that of the high-fat diet. As the dose of GSC-ON188 increased, the liver tissue morphology similar to that of the normal diabetic group was observed, and the disappearance of the lipid was observed (Fig. 6A).

On the other hand, when the abdominal adipose tissue was separated and stained with Hematoxylin & Eosin, it was observed that the fat cells of the high fat diet group were significantly larger than that of the normal diet group, and the fat cells of the positive control group, fenofibrate, . GSC-ON188 mice were significantly reduced in size as the adipocyte size increased, suggesting that accumulation of adipocyte fat was significantly inhibited by GSC-ON188. Therefore, it is concluded that the extract of Fermented Cordyceps sinensis extract inhibits fatty liver and differentiation and growth of adipocytes.

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is evident that many alternatives, modifications and variations will be apparent to those skilled in the art. And will be included in the described technical idea.

Claims (8)

a) Pseudomonas aeruginosa was extracted with distilled water at 105 ° C for 2 hours, and the extract was inoculated with lactic acid bacteria (Pediococcus pentosaceus ON188), fermented for 24 hours, heated at 100 ° C for 10 minutes, and sonicated for 3 minutes step; And
and b) extracting the fermented extract from the fermented Cordyceps mellitus fermentation extract with hot water. The method according to claim 1,
Wherein the fermentation of step (a) is carried out for 24 to 48 hours. The method according to claim 1,
Wherein the extraction of step b) is carried out for 6 to 24 hours. A food composition comprising a fermented extract of Cordyceps mellitus according to any one of claims 1 to 3. A pharmaceutical composition for inhibiting fatty liver comprising the fermented extract of Cordyceps mellon extract of any one of claims 1 to 3. [Claim 6] The pharmaceutical composition according to claim 5, wherein the Fusarium oxysporum fungus extract is one or more selected from the group consisting of CPT1, SPHK2 and PPARa. [Claim 6] The pharmaceutical composition according to claim 5, wherein the fermented extract of Cordyceps mellitus enhances sphingosine 1-phosphate biosynthesis. [Claim 6] The pharmaceutical composition according to claim 5, wherein the fermented extract of Cordyceps mellitus does not exhibit cytotoxicity.

Images