A pharmaceutical composition for preventing or treating hyperlipidemia comprising fermented product of Cudrania tricuspidate branch and a health-functional food comprising the same

Abstract

The present invention relates to a pharmaceutical composition for hyperlipidemia comprising a fermented extract of Cudrania tricuspidata branches and a health functional food comprising the same. Specifically, by including a fermented extract of Cudrania tricuspidata branches as an active ingredient, it is possible to reduce total cholesterol, triglycerides, LDL, and VLDL levels and increase HDL-cholesterol levels. Through this, it is possible to achieve the effect of lowering the arteriosclerosis index, arteriosclerosis correlation coefficient, heart risk index, coronary artery index, etc.

Description

Technology Field

[0001] The present invention relates to a pharmaceutical composition for hyperlipidemia comprising a fermented branch of Cudrania tricuspidata and a health functional food comprising the same. Background Technology

[0002] Cudrania tricuspidate is a deciduous tree belonging to the Moraceae family. According to the *Donguibogam* (a classic Korean medical text), it is known to have tonic, nourishing, and hangover-relieving effects. The stems were used to treat gynecological diseases, the leaves for eczema, pulmonary tuberculosis, bruises, and arthritis, and the fruit for clearing heat and nourishing the blood. The stems of the Cudrania tree are rich in polyphenols, and various benefits have been reported, including antioxidant, antibacterial, anticancer, antidiabetic, and hepatoprotective effects.

[0003] Ganoderma lucidum is distributed worldwide and has been used for various therapeutic purposes, including tonic, diuretic, detoxification, antibacterial, immune, antitussive, analgesic, and hypotensive effects, particularly in Korea, Japan, and China. Various physiological activities, such as anti-inflammatory, anti-diabetic, immunomodulatory, anticancer, antitumor, and antihypertensive effects, have been reported for the fruiting bodies and mycelia of Ganoderma lucidum.

[0004] Hyperlipidemia refers to a condition in which lipids such as free cholesterol, cholesterol esters, phospholipids, and triglycerides are abnormally increased in the blood. Hyperlipidemia can manifest in three forms: (1) hypercholesterolemia, (2) hypertriglyceridemia, and (3) combination hyperlipidemia (hypercholesterolemia and hypertriglyceridemia). Hyperlipidemia is typically classified as primary or secondary hyperlipidemia. Primary hyperlipidemia is generally caused by genetic defects, whereas secondary hyperlipidemia is induced by various disease states, medications, and dietary habits. Additionally, hyperlipidemia can be induced by a combination of primary and secondary causes.

[0005] To date, methods used to reduce blood lipid levels include dietary therapy involving reducing the intake of foods high in cholesterol or fat, exercise therapy, and drug therapy. However, strict management and implementation of dietary therapy and exercise therapy are difficult, and there are limitations in their therapeutic effects. Therefore, in order to efficiently manage and treat hyperlipidemia, there is a need to develop new therapeutic agents capable of treating various symptoms simultaneously. The problem to be solved

[0006] The technical problem to be solved by the present invention is to provide a composition that prevents or treats hyperlipidemia by including a fermented extract of Cudrania tricuspidata branches as an active ingredient, and a health functional food containing the same.

[0007] However, the problems that the present invention aims to solve are not limited to those mentioned above, and other unmentioned problems will be clearly understood by those skilled in the art from the description below. means of solving the problem

[0008] One embodiment of the present invention is a Cudrania tricuspidata tree ( Cudrania tricuspidata Provides a pharmaceutical composition for the prevention or treatment of hyperlipidemia comprising a fermented eggplant extract as an active ingredient.

[0009] According to one embodiment of the present invention, the Cudrania tricuspidata tree ( Cudrania tricuspidata ) Fermented eggplant is Cudrania tricuspidata ( Cudrania tricuspidata ) branches Ganoderma( Ganoderma It may have been fermented with strains of the genus )

[0010] According to one embodiment of the present invention, Ganoderma ( Ganoderma It may further contain Cudrania tricuspidata fruit puree fermented with strains of the genus )

[0011] According to one embodiment of the present invention, the strains of the genus *Ganoderma* include *G. alba*, *G. annularis*, *G. atrum*, *G. aurea*, *G. australe*, *G. amboinense*, *G. applanatum*, *G. boninense*, *G. brownie*, *G. colossus*, *G. cupreum*, *G. curtisii*, *G. formosanum*, *G. incrassatum*, *G. lobatum*, *G. lingzhi*, *G. lucidum*, *G. megaloma*, *G. meredithiae*, *G. miniatocinctum*, *G. multipileum*, *G. neo-japonicum*, *G. nigrolucidum*, *G. orbiforme*, *G. oregonense*, *G. purpurea*, *G. pfeifferi*, *G. philippii*, *G. pseudoferreum*, *G. resinaceum*, *G. rubra*, *G. sessile*, *G. sichuanense*, *G. sinense*, and *G. It may be one selected from the group consisting of steyaertanum, G. tornatum, G. tsugae, G. viridis, G. weberianum, G. zonatum and combinations thereof.

[0012] According to one embodiment of the present invention, the Ganoderma strain used was obtained from a Ganoderma lucidum (G. lucidum KACC 42231) strain, but a strain distributed as Ganoderma lucidum in the market may also be used. Although Ganoderma lucidum generally refers to G. lucidum, in the field of traditional medicine, G. lingzhi, G. multipileum, G. sichuanense, and G. sinense are also referred to as Ganoderma lucidum, so these strains may also be used.

[0013] According to one embodiment of the present invention, the Cudrania tricuspidata branch may be a twig including bark.

[0014] According to one embodiment of the present invention, Cudrania tricuspidata ( Cudrania tricuspidata ) The fermented eggplant product may be an extract obtained by immersing the fermented eggplant of *Cudrania tricuspidata*, fermented with strains of the genus *Ganoderma*, in a solvent.

[0015] According to one embodiment of the present invention, Cudrania tricuspidata ( Cudrania tricuspidata ) Eggplant fermented extract and Ganoderma( Ganoderma It may be extracted by immersing a mixture of Cudrania tricuspidata fruit puree fermented with strains of the genus ) in a solvent.

[0016] According to one embodiment of the present invention, the Cudrania fruit puree may be obtained by adding water to Cudrania fruit and grinding it.

[0017] According to one embodiment of the present invention, the Cudrania tricuspidata fruit may be crushed after adding hot water at room temperature (25℃) to 90℃.

[0018] According to one embodiment of the present invention, the fermented product may include a fermented product, a filtered product obtained after fermentation, and a fermented extract obtained by extracting the fermented product with a solvent.

[0019] According to one embodiment of the present invention, the fermentation can be carried out by inoculating a strain of the genus *Ganoderma* in an amount of 1% to 10% by weight, 1% to 8% by weight, 1% to 6% by weight, 1% to 5% by weight, 2% to 10% by weight, 2% to 8% by weight, 2% to 6% by weight, 2% to 5% by weight, 3% to 10% by weight, 3% to 8% by weight, 3% to 6% by weight, 3% to 5% by weight, 4% to 10% by weight, 4% to 8% by weight, 4% to 6% by weight, or 5% by weight.

[0020] According to one embodiment of the present invention, the fermentation may be carried out at 15°C to 35°C, 20°C to 30°C, or room temperature.

[0021] According to one embodiment of the present invention, the fermentation may be carried out for 1 to 14 days, 1 to 10 days, 1 to 8 days, 1 to 7 days, 2 to 14 days, 2 to 10 days, 2 to 8 days, 2 to 7 days, 3 to 14 days, 3 to 10 days, 3 to 8 days, 3 to 7 days, 4 to 14 days, 4 to 10 days, 4 to 8 days, 4 to 7 days, 5 to 14 days, 5 to 10 days, 5 to 8 days, 5 to 7 days, 6 to 14 days, 6 to 10 days, 6 to 8 days, 6 to 7 days, or 7 days.

[0022] According to one embodiment of the present invention, the solvent may be water, an alcohol having 1 to 3 carbon atoms, or a mixed solvent thereof.

[0023] According to one embodiment of the present invention, the extraction may be performed at 15 to 35°C, 20 to 30°C, or room temperature.

[0024] According to one embodiment of the present invention, the extraction may be performed by immersing in a solvent for 1 to 14 days, 1 to 10 days, 1 to 8 days, 1 to 7 days, 2 to 14 days, 2 to 10 days, 2 to 8 days, 2 to 7 days, 3 to 14 days, 3 to 10 days, 3 to 8 days, 3 to 7 days, 4 to 14 days, 4 to 10 days, 4 to 8 days, 4 to 7 days, 5 to 14 days, 5 to 10 days, 5 to 8 days, 5 to 7 days, 6 to 14 days, 6 to 10 days, 6 to 8 days, 6 to 7 days, or 7 days.

[0025] According to one embodiment of the present invention, the Cudrania tricuspidata branches used in the fermentation and extraction process may be ground to a size of 1 mm to 5 mm, 1 mm to 4 mm, 1 mm to 3 mm, 2 mm to 5 mm, 2 mm to 4 mm, or 2 mm to 3 mm.

[0026] According to one embodiment of the present invention, the hyperlipidemia may be hypercholesterolemia, hypertriglyceridemia, or combined hyperlipidemia (hypercholesterolemia and hypertriglyceridemia).

[0027] Cudrania tricuspidata according to one embodiment of the present invention ( Cudrania tricuspidata A pharmaceutical composition for the prevention or treatment of hyperlipidemia containing a fermented eggplant extract as an active ingredient can reduce levels of total cholesterol, triglycerides, LDL-cholesterol, and VLDL-cholesterol, and increase levels of HDL-cholesterol. Through this, it is possible to achieve the effect of lowering the atherogenic index (AI), atherogenic coefficient (AC), cardiac risk ratio (CRR), and coronary artery index (CAI). Each value can be calculated using the following formulas.

[0028] Arteriosclerosis Index = [TG / HDL-C]

[0029] Atherosclerosis correlation coefficient = [(TC-HDL-C) / HDL-C]

[0030] Cardiac Risk Index = [TC / HDL-C]

[0031] Coronary artery index = [LDL-C / HDL-C]

[0032] According to one embodiment of the present invention, in a hyperlipidemia model, the arteriosclerosis index may be 2.50 or less after the end of a 50-day diet. The arteriosclerosis index may be 2.40, 2.30, 2.20, 2.10, 2.00, 1.90, 1.80, 1.70, 1.60, or 1.50 or less.

[0033] According to one embodiment of the present invention, the arteriosclerosis correlation coefficient may be 0.60 or less after the end of a diet for 50 days in a hyperlipidemia model. The arteriosclerosis correlation coefficient may be 0.55, 0.50, 0.45, 0.40, 0.35, or 0.30 or less.

[0034] According to one embodiment of the present invention, in a hyperlipidemia model, the cardiac risk index may be 1.50 or less after the end of a 50-day diet. The cardiac risk index may be 1.45, 1.40, 1.35, 1.30, 1.25, or 1.20 or less.

[0035] According to one embodiment of the present invention, in a hyperlipidemia model, the coronary artery index may be 0.90 or less after the end of a 50-day diet. The coronary artery index may be 0.85, 0.80, 0.75, 0.70, 0.65, or 0.60 or less.

[0036] According to one embodiment of the present invention, the composition may include a fermented Cudrania tricuspidata branch in an effective amount or as an active ingredient. The effective amount may be appropriately selected depending on the individual. It may be determined based on factors including the severity of the disease or condition, the individual's age, body weight, health, gender, sensitivity to the fermented product, time of administration, route of administration and elimination rate, duration of administration, other compositions combined with or used simultaneously with the composition, and other factors well known in the physiological or medical field.

[0037] According to one embodiment of the present invention, the pharmaceutical composition may be administered by a method known in the art. For example, it may be administered directly to an individual by any means, such as by intravenous, intramuscular, oral, transdermal, mucosal, intranasal, intratracheal, or subcutaneous administration. The administration may be systemic or local.

[0038] According to one embodiment of the present invention, the dosage of the pharmaceutical composition may be 0.1 mg to 1,000 mg per person per day, for example, 0.1 mg to 500 mg, 0.1 mg to 100 mg, 0.1 mg to 50 mg, 0.1 mg to 25 mg, 1 mg to 1,000 mg, 1 mg to 500 mg, 1 mg to 100 mg, 1 mg to 50 mg, 1 mg to 25 mg, 5 mg to 1,000 mg, 5 mg to 500 mg, 5 mg to 100 mg, 5 mg to 50 mg, 5 mg to 25 mg, 10 mg to 1,000 mg, 10 mg to 500 mg, 10 mg to 100 mg, 10 mg to 50 mg, or 10 mg to 25 mg. However, the dosage may vary depending on factors such as the formulation method, method of administration, patient's age, body weight, gender, pathological condition, food, time of administration, route of administration, excretion rate, and response sensitivity, and a person skilled in the art may appropriately adjust the dosage by considering these factors. The frequency of administration may be once a day or two or more times within the range of clinically acceptable side effects, and the administration site may be one or two or more sites, and the total duration of administration may be from 1 to 30 days per treatment, administered daily or at intervals of 2 to 5 days. If necessary, the same treatment may be repeated after the appropriate time. For animals other than humans, the dosage may be the same as that for humans per kg, or an amount calculated by converting the above dosage based on, for example, the ratio of organ (e.g., heart) volume between the target animal and humans (e.g., average value).

[0039] One embodiment of the present invention provides a health functional food for hyperlipidemia comprising a pharmaceutical composition for the prevention or treatment of hyperlipidemia.

[0040] According to one embodiment of the present invention, the health functional food may be formulated into a conventional health functional food formulation known in the art. The health functional food composition may be manufactured into a general formulation such as, for example, a powder, granule, tablet, pill, capsule, suspension, emulsion, syrup, infusion, liquid, or extract, or it may be manufactured into any health food form such as meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, chewing gum, jelly, ice cream, dairy products, various soups, beverages, tea, drinks, alcoholic beverages, and vitamin complexes. Food science-acceptable carriers or additives may be used for the formulation of the health food, and any carrier or additive known to be available in the art may be used for the manufacture of the formulation to be manufactured. The above additives may contain various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, etc. In addition, it may contain fruit pulp for the production of natural fruit juices, fruit juice beverages, and vegetable beverages.

[0041] According to one embodiment of the present invention, the pharmaceutical composition for the prevention or treatment of hyperlipidemia in the health functional food composition may be appropriately determined according to the purpose of use (prevention or improvement). Generally, it may be included in an amount of 0.01% to 15% by weight of the total food weight, and when prepared as a beverage, it may be included in an amount of 0.02 g to 10 g, specifically 0.3 g to 1 g, based on 100 mL. The beverage may further include other ingredients in addition to the composition, and may further include various flavoring agents or natural carbohydrates commonly used in beverages. The natural carbohydrates may include conventional sugars such as monosaccharides (e.g., glucose, fructose, etc.), disaccharides (e.g., maltose, sucrose, etc.), and polysaccharides (e.g., dextrin, cyclodextrin, etc.), and sugar alcohols such as xylitol, sorbitol, and erythritol. In addition, natural flavoring agents (e.g., taumatin, stevia extract, etc.) and synthetic flavoring agents (e.g., saccharin, aspartame, etc.) may be included as flavoring agents. The proportion of the natural carbohydrate may generally be about 1 g to 20 g, specifically about 5 g to 12 g per 100 mL of beverage. These additive components may be used independently or in combination, and the proportion of the additive may be 0.001 g to 5 weight%, specifically 0.01 g to 3 weight%, based on the total weight of the composition. Effects of the invention

[0042] Cudrania tricuspidata according to one embodiment of the present invention ( Cudrania tricuspidata A pharmaceutical composition for the prevention or treatment of hyperlipidemia containing a fermented eggplant extract as an active ingredient and a health functional food containing the same can reduce total cholesterol, triglyceride, LDL, and VLDL levels and increase HDL-cholesterol levels. Through this, it is possible to achieve the effect of lowering the arteriosclerosis index, arteriosclerosis correlation coefficient, heart risk index, coronary artery index, etc. Brief explanation of the drawing

[0043] Fig. 1 ...demonstrates the effect of fermented Cudrania tricuspidata branches on glucose uptake in TNF-α-treated 3T3-L1 adipocytes. Fig. 2 This shows the effect of fermented Cudrania tricuspidata branches on adiponectin levels in TNF-α-treated 3T3-L1 adipocytes. Fig. 3 It shows the effect of fermented Cudrania tricuspidata branches on inflammatory cytokine levels in TNF-α-treated 3T3-L1 adipocytes. Fig. 4 This shows the effect of fermented Cudrania tricuspidata branches on PPARγ expression in TNF-α-treated 3T3-L1 adipocytes. Fig. 5 This shows the effect of Cudrania tricuspidata fermentation on GLUT expression in TNF-α-treated 3T3-L1 adipocytes. Fig. 6 This shows the effects of fermented CTSB extract and fenofibrate on the body weight of mice with hyperlipidemia and obesity. Fig. 7 This shows the effects of fermented CTSB extract and fenofibrate on the weight gain rate of mice with hyperlipidemia and obesity. Fig. 8 This shows the effects of fermented CTSB extract and fenofibrate on the diet of mice with hyperlipidemia and obesity. Fig. 9 This shows the effects of fermented CTSB extract and fenofibrate on the dietary efficiency of mice with hyperlipidemia and obesity. Fig. 10 This shows the effects of fermented CTSB extract and fenofibrate on liver tissue weight, kidney weight, and spleen weight in mice with hyperlipidemia and obesity. Fig. 11 This shows the effects of fermented CTSB extract and fenofibrate on inhibiting epididymal fat and liver fat accumulation in mice with hyperlipidemia and obesity. Fig. 12 This shows the effects of fermented CTSB extract and fenofibrate on serum glucose levels, serum insulin levels, and insulin resistance (HOMA-IR) in mice with hyperlipidemia and obesity. Fig. 13This shows the effects of fermented CTSB extract and fenofibrate on total protein, total cholesterol, triglycerides, HDL-cholesterol, LDL-cholesterol, and VLDL-cholesterol in mice with hyperlipidemia and obesity. Fig. 14 This shows the effects of fermented CTSB extract and fenofibrate on the arteriosclerosis index (AI), arteriosclerosis coefficient (AC), cardiac risk ratio (CRR), and coronary artery index (CAI) in mice with hyperlipidemia and obesity. Specific details for implementing the invention

[0044] In this specification, when a part is described as "comprising" a certain component, this means that, unless specifically stated otherwise, it does not exclude other components but may include additional components.

[0046] Hereinafter, the present invention will be described in detail with reference to examples to specifically explain the invention. However, the embodiments according to the present invention may be modified in various different forms, and the scope of the present invention is not to be interpreted as being limited to the embodiments described below. The embodiments of this specification are provided to more completely explain the present invention to those with average knowledge in the art.

[0047] Experimental method

[0048] 1. Samples and Strains

[0049] The Cudrania tricuspidata branches used were harvested directly in Gimje, dried, and stored at room temperature. Reishi mushrooms ( G. lucidum The strain KACC 42231) was obtained from the Korean Agricultural Culture Collection (KACC) of the Rural Development Administration and used.

[0051] 2. Preparation of fermented extract of Cudrania tricuspidata branches

[0052] A Ganoderma lucidum strain cultured in malt extract broth (Kisan Biotech, Seoul, Korea) at 25°C for one week was inoculated onto a Cudrania tricuspidata stem and cultured at 25°C for 20 days to be used as a Ganoderma lucidum spawn for the fermentation of Cudrania tricuspidata branches.

[0053] Sawdust was prepared by crushing dried Cudrania tricuspidata branches, specifically Cudrania tricuspidata twigs, into pieces 2 mm to 3 mm in size using a crusher. 3 liters of hot water (90 ℃) were added to 1 kg of frozen Cudrania tricuspidata fruit, and the mixture was homogenized using a mixer to prepare Cudrania tricuspidata fruit puree. A mixture was prepared by mixing the Cudrania tricuspidata branch sawdust and the Cudrania tricuspidata fruit puree, and the moisture content of the mixture was adjusted to approximately 65% ​​by weight. The mixture was placed in a container and sterilized at 121 ℃ for 45 minutes. The sterilized mixture was inoculated with approximately 5% by weight of the Reishi mushroom spawn relative to 100% by weight of the mixture, fermented at 25 ℃ for one week, and then freeze-dried.

[0054] Subsequently, 300 ml of an aqueous ethanol solution (concentration: 70 wt%) was added to 100 g each of the fermented and unfermented sawdust of Cudrania tricuspidata branches, and the mixture was immersed at room temperature for one week to prepare an extract. The extract was filtered using filter paper (Adantec, Tokyo, Japan), concentrated using a rotary vacuum concentrator (Tokyo Rikakikai, Tokyo, Japan), and then freeze-dried (Ilshin Lab, Dongducheon, Korea) to produce a powder. The powder was stored at -20 ℃ and used as a sample in the experiment.

[0056] 3. 3T3-L1 cell differentiation and treatment with TNF-α and extracts

[0057] The 3T3-L1 cells used in the experiment were obtained from the American Type Culture Collection (Manassas, VA, USA). Cells were cultured in Dulbecco's modified Eagle's medium (DMEM) (Invitrogen, Waltham, MA, USA) with 10% bovine serum (BS) (Invitrogen, Waltham, MA, USA) and 1% antibiotic-antimycotic solution (Invitrogen, Waltham, MA, USA) added, in a 5% CO2, 37°C incubator. For cell differentiation, 3T3-L1 cells were seeded and cultured until 100% confluence, after which the DMEM medium was replaced with one containing 10% fetal bovine serum (FBS) (Welgene, Gyeongsan, Korea). After 2 days, differentiation was induced by replacing the DMEM medium containing 10% FBS with 2 mM dexamethasone, 0.5 mM 3-isobutyl-methylxanthine, and 1 μg / mL insulin. After 2 days, the medium was replaced with DMEM containing 10% FBS containing 1 μg / mL insulin and cultured for 2 days. The medium was then replaced with DMEM containing 10% FBS and cultured for an additional 2 days. Fully differentiated 3T3-L1 cells were treated with fermented Cudrania tricuspidata twig extract at various concentrations for 4 hours, followed by treatment with 10 ng / mL TNF-α for 24 hours and analysis.

[0059] 4. Glucose Absorption Analysis

[0060] 3T3-L1 cells (1×10⁴ cells / well) were seeded into each well of a 96-well plate, and after differentiation induction was complete, they were cultured for 4 hours in KRPH buffer (121 mM NaCl, 4.9 mM KCl, 1.2 mM MgSO₄, 0.33 mM CaCl₂, 12 mM HEPES, pH 7.4). After treating with the extract for 6 hours, 10 ng / mL TNF-α was added for 15 minutes, followed by 10 nM insulin for 30 minutes. After removing the medium, 900 μL of KRPH and 10 μg / mL insulin were added for 1 hour, followed by 10 mM 2-deoxyglucose (2-DG) and incubation for 30 minutes. Glucose uptake was measured using a glucose uptake assay kit (Abcam, Cambridge, UK).

[0062] 5. Enzyme Immunoassay (ELISA)

[0063] 3T3-L1 cells (1×10⁴ cells / well) were seeded into each well of a 96-well plate, and fully differentiated cells were treated with the extract for 6 hours, followed by treatment with 10 ng / mL TNF-α for 24 hours. Adiponectin and inflammatory cytokines contained in the medium obtained after treatment were measured using enzyme-linked immunosorbent assay kits (Enzo Life Sciences, Farmingdale, NY, USA).

[0065] 6. Western blot assay

[0066] After treatment, the cells were washed with PBS, and proteins were extracted by treating with lysis buffer (RIPA buffer, 50 mM Tris-HCl, pH 8.0, 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS) for 20 minutes at 4°C. Proteins were quantified using a BCA protein assay kit (Thermofisher, Waltham, MA, USA), and equal amounts of protein were electrophoresed on 10% SDS-PAGE. After electrophoresis, proteins were transferred to a PVDF membrane (Roche Diagnostics GmbH, Mannheim, Germany) and blocked with blocking buffer (Bio-Rad, Hercules, CA, USA) for 10 minutes at room temperature. The primary antibody was diluted to 1:1,000–1:4,000 and incubated at 4°C for 12 hours, after which the membrane was washed three times for 15 minutes each with TBS-T (Tris-buffered saline with Tween-20) buffer. The secondary antibody was diluted to 1:1,000–1:2,500 and incubated at room temperature for 2 hours, after which the membrane was washed three times for 15 minutes each with TBS-T buffer. The membrane was treated with an enhanced chemiluminescent substrate (Thermofisher, Waltham, MA, USA), and the protein bands generated on the membrane were identified using an image reader (Microchemi 4.2, DNR, Neve Yamin, ISR) to analyze the level of protein expression.

[0068] 7. Production of a hyperlipidemia model

[0069] In this experiment, 50 male ICR white rats aged 8 weeks were purchased and used. For one week prior to the start of the experimental diet, the rats were acclimatized on a solid compound feed. Afterward, based on body weight, they were randomly assigned to five groups of 10 rats each and reared for 50 days in an optimal environment (temperature 20±2°C, 12-hour light-dark cycle). The experimental diets were divided into a Normal Control group, a Hyperlipidemia Treatment Negative Control group, a Hyperlipidemia Treatment + Sample 1 experimental group, a Hyperlipidemia Treatment + Sample 2 experimental group, and a Hyperlipidemia Treatment Positive Control group. The dietary compositions for the Normal Control group, the control group, and the experimental group to be used in this experiment were formulated, and all food and drinking water were provided via a free-form method. Body weight and food intake were measured at regular intervals.

[0071] 8. Analysis of changes in adipose and liver tissues

[0072] After blood collection, tissues such as liver and fat were extracted from experimental animals, washed with PBS, and stored at -70°C until use. After the diet was completed, fat was extracted, weighed, and analyzed. For the morphology and analysis of fatty liver, a portion of the middle lobe of the liver was excised, washed with physiological saline, and fixed using 3.75% paraformaldehyde. After fixation, the tissues were washed with physiological saline and dehydrated using ethanol of various concentrations. Subsequently, blocks were prepared using Tissue-Tek (Sakura prohosp) and sectioned to a thickness of 15–30 μm using a micrometer (HM 325, Thermo Scientific, Florida, FL, USA). To observe fat in the adipose tissue and liver tissue, the tissues were stained with Oil Red O for 10 minutes and observed under a 100x magnification optical microscope (Olympus CX-31, Japan).

[0074] 9. Biochemical analysis in serum

[0075] To perform biochemical analysis in the serum, blood was collected from the heart after 24 hours of fasting and anesthesia with ethyl ether. The collected blood was left at room temperature for 20 minutes, and then centrifuged at 8,000 rpm for 20 minutes to separate the serum. The separated serum was stored at -70℃ until use. For biochemical analysis, total cholesterol, triglycerides (TG), and high-density lipoprotein cholesterol (HDL-C) were measured using a Vitros 250 chemistry system (Ortho Clinical Diagnostics) to analyze the effect of reducing body fat. Low-density lipoprotein cholesterol (LDL-C), very low-density lipoprotein cholesterol (VLDL-C), glucose, and protein were analyzed using the Friedewald equation concentration calculation method.

[0077] 10. Analysis of changes in arteriosclerosis index, arteriosclerosis correlation coefficient, cardiac risk index, and coronary artery index

[0078] Based on the results obtained from biochemical analysis, changes in four cardiovascular-related risk indices (Atherogenic index (AI), Atherogenic coefficient (AC), Cardiac risk ratio (CRR), and Coronary artery index (CAI)) were analyzed.

[0080] 11. Insulin Resistance Analysis

[0081] The homeostasis model of insulin resistance (HOMA-IR) was calculated using measured blood glucose and insulin concentrations.

[0083] Example 1. Analysis of the effect of fermented Cudrania tricuspidata twigs on improving insulin resistance

[0084] 1-1. Regulation of glucose absorption by fermented Cudrania tricuspidata twigs

[0085] To confirm the mitigating effect of fermented Cudrania tricuspidata twigs on TNF-α-induced insulin resistance, changes in glucose absorption were examined using 2-deoxyglucose (2-DG). Compared to the control group, glucose absorption decreased to 49% in the TNF-α-alone treatment group (untreated group with fermented Cudrania tricuspidata twigs), but it was confirmed that glucose absorption increased to 57%, 65%, and 81% in the groups treated with fermented Cudrania tricuspidata twigs at concentrations of 100, 200, and 300 μg / mL, respectively. Fig. 1 ). Fig. 1 This shows the effect of fermented Cudrania tricuspidata branches on glucose uptake in TNF-α-treated 3T3-L1 adipocytes. All differentiated 3T3-L1 adipocytes were treated with 10 ng / mL TNF-α alone or pretreated with fermented Cudrania tricuspidata branches for 24 hours.

[0087] 1-2. Analysis of the Effects of Fermented Cudrania Tilia on Adiponectin Production

[0088] Adiponectin and TNF-α act in opposition to each other in adipose tissue and play antagonistic roles in the regulation of insulin resistance. Adiponectin exhibits anti-inflammatory effects and increases insulin sensitivity, whereas TNF-α is known to reduce the production of adiponectin in adipocytes and increase insulin resistance. As a result of confirming the inhibitory effect of fermented Cudrania tricuspidata twig extract on the reduction of adiponectin induced by TNF-α, it was confirmed that compared to the control group, the TNF-α-alone treatment group (group untreated with fermented Cudrania tricuspidata twig extract) showed a decrease to 10.95 ng / ml, whereas the groups treated at concentrations of 100, 200, and 300 μg / mL showed an increase in adiponectin production to 13.91, 20.96, and 28.20 ng / ml, respectively. Fig. 2 ). Fig. 2 This shows the effect of fermented Cudrania tricuspidata branches on adiponectin levels in TNF-α-treated 3T3-L1 adipocytes. All differentiated 3T3-L1 adipocytes were treated with 10 ng / mL TNF-α alone or pretreated with fermented Cudrania tricuspidata branches for 24 hours. Adiponectin levels were determined by ELISA.

[0090] 1-3. Analysis of the Effects of Fermented Cudrania Tilia Twigs on the Production of Inflammatory Cytokines

[0091] IL-6 induces inflammation in the liver and adipose tissue, regulates the expression of GLUT4 and IRS-1, and increases insulin resistance, while MCP-1 induces macrophage infiltration into adipose tissue, maintains an inflammatory state, and induces insulin resistance by suppressing the expression of genes such as LPL, GLUT4, and PPARγ. To confirm the inhibitory effect of fermented Cudrania tricuspidata twig extract on TNF-α-induced inflammation, the production levels of MCP-1 and IL-6 were measured. As a result, IL-6 production increased due to TNF-α, with levels of 45.91 pg / mL in the control group and 158.02 ng / mL in the group treated with TNF-α alone; however, in the groups treated with fermented Cudrania tricuspidata twig extract, IL-6 production was found to be 139.51, 110.66, and 83.97 pg / mL, respectively, which was lower than that of the group treated with TNF-α alone. Fig. 3 A). In addition, regarding MCP-1, the level was 139.97 pg / mL in the control group and 139.97 pg / mL in the group treated with TNF-α alone, indicating that MCP-1 production was increased by TNF-α ( Fig. 3 B). In the group treated with fermented Cudrania tricuspidata twigs, the production of MCP-1 was 113.97, 92.98, and 77.87 pg / mL, which was found to be reduced compared to the group treated with TNF-α alone. Fig. 3 This shows the effect of fermented Cudrania tricuspidata branches on inflammatory cytokine levels in TNF-α-treated 3T3-L1 adipocytes. All differentiated 3T3-L1 adipocytes were treated with 10 ng / mL TNF-α alone or pretreated with fermented Cudrania tricuspidata branches for 24 hours. Cytokine levels were determined by ELISA.

[0093] 1-4. Analysis of the Effects of Fermented Cudrania Tilia on Insulin Resistance-Related Gene Expression

[0094] TNF-α is known to induce insulin resistance in adipocytes by inhibiting the activity of PPARγ. PPARγ is a factor that plays a crucial role in adipocyte differentiation, maintenance of blood glucose homeostasis, and lipid metabolism, inducing insulin sensitivity enhancement, a reduction in inflammation-related factors, and increased adiponectin secretion. To investigate the effect of TNF-α on PPARγ expression related to insulin resistance in 3T3-L1 cells, PPARγ expression was examined. The results showed a sharp decrease in expression in the group treated with TNF-α alone compared to the control group. Conversely, an increase in PPARγ protein expression was observed in the group treated with fermented Cudrania tricuspidata twig extract. Fig. 4 This shows the effect of fermented Cudrania tricuspidata branches on PPARγ expression in TNF-α-treated 3T3-L1 adipocytes. All differentiated 3T3-L1 adipocytes were treated with 10 ng / mL TNF-α alone or pretreated with fermented Cudrania tricuspidata branches for 24 hours. Protein expression was determined by Western blot.

[0095] GLUT4 is involved in insulin sensitivity and insulin-induced glucose uptake, and mediates the influx of intracellular glucose induced by insulin stimulation. TNF-α is known to inhibit the expression of GLUT4; a decrease in GLUT4 leads to reduced insulin sensitivity, increased insulin resistance, and a reduction in intracellular glucose influx. To determine whether fermented Cudrania tricuspidata twig extract improves TNF-α-induced insulin resistance, changes in the expression of GLUT4, an insulin signaling factor, were examined. The results showed that compared to the control group, GLUT4 expression decreased sharply in the group treated with TNF-α alone. Conversely, an increase in GLUT4 protein expression was observed in the group treated with the fermented Cudrania tricuspidata twig extract. Fig. 5This shows the effect of Cudrania tricuspidata fermentation on GLUT expression in TNF-α-treated 3T3-L1 adipocytes. All differentiated 3T3-L1 adipocytes were treated with 10 ng / mL TNF-α alone or pretreated with fermented Cudrania tricuspidata branches for 24 hours. Protein expression was determined by Western blot.

[0097] Example 2. Analysis of the effects of improving obesity and triglycerides using an experimental animal model

[0098] 2-1. Analysis of Body Weight and Dietary Intake in Hyperlipidemia Models

[0099] To analyze the hyperlipidemia-inhibiting efficacy of fermented Cudrania tricuspidata twig extract (CTSB extract), a hyperlipidemia model was induced by treatment with tyloxapol (300 mg / kg), and fermented Cudrania tricuspidata twig extract was administered orally at daily concentrations of 500 mg / kg / day and 1000 mg / kg / day. For comparative analysis, administration of the lipid-lowering agent fenofibrate (200 mg / kg / day) was used as a positive control. The results regarding average body weight over 50 days show average body weight at 5-day intervals. In the control group fed only a regular diet (regular diet group), the average body weight was confirmed to have increased from 34.8 ± 0.8 g (day 0) to 46.1 ± 1.4 g (day 50); in the negative control group fed only a regular diet after hyperlipidemia model treatment, it was confirmed to have increased from 36.3 ± 1.5 g (day 0) to 43.6 ± 1.3 g (day 50); in the positive control group administered fenofibrate after hyperlipidemia model treatment (Model+fenofibrate group), it increased from 36.5 ± 0.7 g (day 0) to 45.1 ± 1.2 g (day 50); and in Experimental Group 1 (Model+CTSB500 group), administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after hyperlipidemia model treatment, it increased from 36.1 ± 1.0 g (day 0) to 43.1 ± 0.4 g (day 50); and in the fermented Experimental group 2 (Model+CTSB1000 group), administered a high concentration (1000 mg / kg / day) of Cudrania tricuspidata twig extract, was confirmed to have weights ranging from 35.5 ± 0.8 g (day 0) to 42.8 ± 1.5 g (day 50). Fig. 6 ). Fig. 6 This shows the effects of fermented CTSB extract and fenofibrate on the body weight of mice with hyperlipidemia and obesity. Sample treatment started on day 3 and ended on day 50.

[0100] The average weight gain rate was confirmed to be 131.7 ± 6.8% in the control group fed only a regular diet and 121.3 ± 7.5% in the model group fed only a regular diet after hyperlipidemia model treatment, when the pre-treatment weight of all diet groups was set to 100%. Additionally, the control group administered fenofibrate after hyperlipidemia model treatment showed 125.2 ± 8.1%, the group receiving low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after hyperlipidemia model treatment (Model+CTSB500 group) showed 120.3 ± 4.6%, and the group receiving high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after hyperlipidemia model treatment (Model+CTSB1000 group) showed 120.6 ± 5.9%. Fig. 7 ). Fig. 7 This shows the effects of fermented CTSB extract and fenofibrate on the weight gain rate of mice with hyperlipidemia and obesity.

[0101] Average diet results at 5-day intervals Fig. 8 As shown in [figure], the average daily dietary intake results were confirmed to be 81-105 g in the control group fed only a regular diet, 82-119 g in the model group fed only a regular diet after hyperlipidemia model treatment, 79-115 g in the positive control group administered fenofibrate after hyperlipidemia model treatment, 84-119 g in Experimental Group 1 (Model+CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after hyperlipidemia model treatment, and 86-125 g in Experimental Group 2 (Model+CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after hyperlipidemia model treatment. Fig. 8 ). Fig. 8 This shows the effects of fermented CTSB extract and fenofibrate on the diet of mice with hyperlipidemia and obesity.

[0102] The dietary efficiency values ​​derived from this were confirmed to be 0.031 in the normal diet group, 0.020 in the negative control group, 0.024 in the positive control group (Model+fenofibrate group), 0.020 in experimental group 1 (Model+CTSB500 group), and 0.021 in experimental group 2 (Model+CTSB1000 group). Fig. 9 ). Fig. 9 This shows the effects of fermented CTSB extract and fenofibrate on the dietary efficiency of mice with hyperlipidemia and obesity. As such, no significant changes in body weight or food intake were observed with the intake of fermented Cudrania tricuspidata twig extract.

[0104] 2-2. Analysis of Liver, Kidney, and Spleen Tissue Weights in a Hyperlipidemia Model

[0105] After the 50-day diet was completed, tissues from each group were extracted, processed, and analyzed. Fig. 10 This shows the effects of fermented CTSB extract and fenofibrate on liver tissue weight, kidney weight, and spleen weight in mice with hyperlipidemia and obesity.

[0106] The weight of liver tissue was confirmed to be 2.3 ± 0.1 g in the control group fed only a normal diet, 2.5 ± 0.2 g in the negative control group fed only a normal diet after hyperlipidemia model treatment, 2.3 ± 0.2 g in the positive control group (Model + fenofibrate group) administered fenofibrate after hyperlipidemia model treatment, 2.4 ± 0.2 g in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after hyperlipidemia model treatment, and 2.2 ± 0.1 g in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after hyperlipidemia model treatment. Fig. 10 of A).

[0107] The weight of kidney tissue was confirmed to be 0.88 ± 0.05 g in the control group fed only a normal diet, 0.82 ± 0.03 g in the negative control group fed only a normal diet after hyperlipidemia model treatment, 0.82 ± 0.03 g in the control group administered fenofibrate after hyperlipidemia model treatment (Model + fenofibrate group), 0.81 ± 0.06 g in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after hyperlipidemia model treatment, and 0.83 ± 0.04 g in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after hyperlipidemia model treatment. Fig. 10 of B).

[0108] The weight of spleen tissue was confirmed to be 0.12 ± 0.01 g in the control group fed only a normal diet, 0.11 ± 0.01 g in the negative control group fed only a normal diet after hyperlipidemia model treatment, 0.12 ± 0.02 g in the positive control group (Model + fenofibrate group) administered fenofibrate after hyperlipidemia model treatment, 0.12 ± 0.02 g in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after hyperlipidemia model treatment, and 0.12 ± 0.01 g in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after hyperlipidemia model treatment. Fig. 10 of C).

[0109] Through these results, it was observed that in the hyperlipidemia obesity model, liver weight increased slightly, while kidney and spleen weights decreased slightly. In the hyperlipidemia obesity model, no significant changes were observed in liver and kidney weights in the experimental group that consumed fermented Cudrania tricuspidata twig extract, and while spleen weight increased on average, no significant change was observed.

[0111] 2-3. Analysis of Hepatic Fat and Epididymal Fat in a Hyperlipidemia Model

[0112] Fig. 11 This study shows the effects of fermented CTSB extract and fenofibrate on inhibiting epididymal fat and hepatic fat accumulation in mice with hyperlipidemia and obesity. After the diet was completed for 50 days, liver tissue and epididymal fat were extracted from each group to analyze the degree of hepatic fat and body fat accumulation.

[0113] In the tissue weight analysis of epididymal adipose tissue, the values ​​were confirmed to be 1.2 ± 0.1 g in the control group fed only a regular diet, 1.0 ± 0.1 g in the negative control group fed only a regular diet after treatment with the hyperlipidemia obesity model, 1.0 ± 0.2 g in the positive control group (Model + fenofibrate group) administered fenofibrate after treatment with the hyperlipidemia model, 1.0 ± 0.2 g in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model, and 0.9 ± 0.1 g in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model. Fig. 11 of A).

[0114] As a result of observing and analyzing the density of lipids and fats in the liver tissues of each group by staining them with Oil Red 0, when the fat density of the control group fed only a regular diet was set to 100%, the values ​​were confirmed to be 161 ± 6% for the negative control group consuming only a regular diet after treatment with the hyperlipidemia obesity model, 135 ± 4% for the positive control group administered fenofibrate after treatment with the hyperlipidemia model, 131 ± 5% for Experimental Group 1 (Model+CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model, and 129 ± 3% for Experimental Group 2 (Model+CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model. Fig. 11 of B).

[0115] Body fat accumulation in the epididymis tissue was actually reduced in the hyperlipidemia model group compared to the control group fed only a regular diet. In the hyperlipidemia model, the weight of epididymis fat was lower on average in all groups that consumed the lipid-lowering agent fenofibrate and fermented Cudrania tricuspidata twig extract compared to the control group fed only a regular diet. Similar levels were observed at low concentrations (500 mg / kg / day) of fenofibrate and fermented Cudrania tricuspidata twig extract, while the greatest average reduction was observed at high concentrations (1000 mg / kg / day).

[0116] Fat in liver tissue increased by 61% in the hyperlipidemia model group compared to the control group fed only a normal diet; however, a significant decrease was observed in all groups administered the lipid-lowering agent fenofibrate and fermented Cudrania tricuspidata twig extract. Specifically, the rate of increase decreased by 135% in the group administered fenofibrate, 131% in the group administered low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract, and 129% in the group administered high concentration (1000 mg / kg / day). The group administered high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract showed the highest inhibition rate of fat accumulation.

[0118] 2-4. Analysis of Blood Glucose, Insulin, and Insulin Resistance in a Hyperlipidemia Model

[0119] Fig. 12 This shows the effects of fermented CTSB extract and fenofibrate on serum glucose levels, serum insulin levels, and insulin resistance (HOMA-IR) in mice with hyperlipidemia and obesity.

[0120] After the 50-day diet was completed, serum glucose concentrations were measured as follows: 161 ± 14 mg / dL in the control group fed only a regular diet; 159 ± 9 mg / dL in the negative control group fed only a regular diet after treatment with the hyperlipidemia obesity model; 153 ± 11 mg / dL in the positive control group (Model + fenofibrate group) administered fenofibrate after treatment with the hyperlipidemia model; 176 ± 7 mg / dL in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model; and 172 ± 6 mg / dL in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model. Fig. 12A). When compared to a negative control group that consumed only a normal diet after treatment with a hyperlipidemia obesity model, blood glucose levels were maintained without significant change with the intake of the lipid-lowering agent fenofibrate and fermented Cudrania tricuspidata branch extract, but blood glucose levels increased slightly on average with the intake of the fermented Cudrania tricuspidata branch extract and decreased slightly on average with the intake of the lipid-lowering agent.

[0121] Serum insulin concentrations were measured as 0.041 ± 0.009 ng / mL in the control group consuming only a regular diet, 0.038 ± 0.006 ng / mL in the negative control group consuming only a regular diet after treatment with the hyperlipidemia obesity model, 0.035 ± 0.005 ng / mL in the positive control group (Model + fenofibrate group) administered fenofibrate after treatment with the hyperlipidemia model, 0.049 ± 0.011 ng / mL in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model, and 0.040 ± 0.004 ng / mL in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model. Fig. 12 B). When compared to a negative control group that consumed only a normal diet after treatment with a hyperlipidemia model, the intake of the lipid-lowering agent fenofibrate resulted in a slight average decrease in insulin, whereas the intake of fermented Cudrania tricuspidata twig extract increased insulin concentration on average, but no significant change was observed.

[0122] Serum insulin resistance levels were calculated using blood glucose and insulin concentrations of each group and were confirmed to be 0.018 ± 0.001 in the control group consuming only a regular diet, 0.015 ± 0.001 in the negative control group consuming only a regular diet after treatment with the hyperlipidemia obesity model, 0.013 ± 0.001 in the positive control group (Model + fenofibrate group) administered fenofibrate after treatment with the hyperlipidemia model, 0.022 ± 0.002 in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model, and 0.017 ± 0.001 in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model. Fig. 12 C). When compared to a negative control group that consumed only a normal diet after treatment with a hyperlipidemia obesity model, insulin resistance decreased slightly on average with the intake of the lipid-lowering agent fenofibrate, but the intake of fermented Cudrania tricuspidata twig extract did not show a significant change.

[0124] 2-5. Analysis of Total Protein, Total Cholesterol, Triglycerides, HDL-Cholesterol, LDL-Cholesterol, and VLDL in a Hyperlipidemia Model

[0125] Fig. 13 This shows the effects of fermented CTSB extract and fenofibrate on total protein, total cholesterol, triglycerides, HDL-cholesterol, LDL-cholesterol, and VLDL-cholesterol in mice with hyperlipidemia and obesity.

[0126] After the 50-day diet was completed, serum total protein concentrations were measured as 5.3 ± 0.4 mg / dL in the control group fed only a regular diet, 5.3 ± 0.5 mg / dL in the negative control group fed only a regular diet after treatment with the hyperlipidemia obesity model, 5.2 ± 0.4 mg / dL in the positive control group (Model + fenofibrate group) administered fenofibrate after treatment with the hyperlipidemia model, 5.3 ± 0.3 mg / dL in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model, and 5.3 ± 0.4 mg / dL in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model. Fig. 13 A). When compared to a negative control group that consumed only a normal diet after treatment with a hyperlipidemia obesity model, total protein was maintained without significant change with the intake of the lipid-lowering agent fenofibrate and fermented Cudrania tricuspidata branch extract.

[0127] Serum total cholesterol concentrations were measured as 104 ± 7 mg / dL in the control group fed only a regular diet, 245 ± 11 mg / dL in the negative control group fed only a regular diet after treatment with the hyperlipidemia obesity model, 119 ± 6 mg / dL in the positive control group (Model + fenofibrate group) administered fenofibrate after treatment with the hyperlipidemia model, 195 ± 7 mg / dL in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model, and 180 ± 9 mg / dL in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model. Fig. 13B). When compared to a negative control group that consumed only a normal diet after treatment with a hyperlipidemia-obesity model, the intake of the lipid-lowering agent fenofibrate and fermented Cudrania tricuspidata branch extract showed a significant change in total cholesterol, with the greatest decrease observed in the group that consumed the lipid-lowering agent fenofibrate. The group that consumed the fermented Cudrania tricuspidata branch extract showed a concentration-dependent decrease.

[0128] Serum triacylglycerol concentrations were measured as 93 ± 4 mg / dL in the control group fed only a normal diet, 518 ± 16 mg / dL in the negative control group fed only a normal diet after treatment with the hyperlipidemia obesity model, 152 ± 7 mg / dL in the positive control group (Model + fenofibrate group) administered fenofibrate after treatment with the hyperlipidemia model, 295 ± 11 mg / dL in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model, and 266 ± 13 mg / dL in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model. Fig. 13 C). When compared to a control group that consumed only a normal diet after treatment with a hyperlipidemia obesity model, triacylglycerol levels showed significant changes with the intake of the lipid-lowering agent fenofibrate and fermented Cudrania tricuspidata branch extract. The group that consumed the lipid-lowering agent fenofibrate showed the greatest decrease, while the group that consumed the fermented Cudrania tricuspidata branch extract showed a concentration-dependent decrease.

[0129] Serum high-density lipoprotein cholesterol (HDL-cholesterol) concentrations were measured as 110 ± 4 mg / dL in the control group fed only a normal diet, 147 ± 3 mg / dL in the negative control group fed only a normal diet after treatment with the hyperlipidemia obesity model, 136 ± 7 mg / dL in the positive control group (Model + fenofibrate group) administered fenofibrate after treatment with the hyperlipidemia model, 145 ± 4 mg / dL in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model, and 148 ± 2 mg / dL in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model. Fig. 13 D). When compared to a negative control group that consumed only a normal diet after treatment with a hyperlipidemia obesity model, high-density lipoprotein cholesterol (HDL-cholesterol) showed significant changes with the intake of the lipid-lowering agent fenofibrate and fermented Cudrania tricuspidata branch extract, with the largest increase observed in the group that consumed the fermented Cudrania tricuspidata branch extract and a concentration-dependent increase.

[0130] Serum low-density lipoprotein cholesterol (LDL-cholesterol) concentrations were measured as follows: 12.6 ± 3.8 mg / dL in the control group fed only a regular diet; 201.6 ± 11.2 mg / dL in the negative control group fed only a regular diet after treatment with the hyperlipidemia obesity model; 13.4 ± 0.4 mg / dL in the positive control group administered fenofibrate after treatment with the hyperlipidemia model (Model + fenofibrate group); 109 ± 5.2 mg / dL in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model; and 85.2 ± 9.6 mg / dL in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model. Fig. 13 E). When compared to a control group that consumed only a normal diet after treatment with a hyperlipidemia obesity model, the intake of the lipid-lowering agent fenofibrate and fermented Cudrania tricuspidata branch extract showed a significant change in low-density lipoprotein cholesterol (LDL-cholesterol), with the greatest decrease observed in the group that consumed the lipid-lowering agent fenofibrate and a concentration-dependent decrease observed in the group that consumed the fermented Cudrania tricuspidata branch extract.

[0131] Serum very low-density lipoprotein cholesterol (VLDL-cholesterol) concentrations were measured as follows: 18.6 ± 0.8 mg / dL in the control group fed only a regular diet; 103.6 ± 3.2 mg / dL in the negative control group fed only a regular diet after treatment with the hyperlipidemia obesity model; 30.4 ± 1.4 mg / dL in the positive control group (Model + fenofibrate group) administered fenofibrate after treatment with the hyperlipidemia model; 59 ± 2.2 mg / dL in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model; and 53.2 ± 2.6 mg / dL in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model. Fig. 13 F). When compared to a negative control group that consumed only a normal diet after treatment with a hyperlipidemia obesity model, the intake of the lipid-lowering agent fenofibrate and fermented Cudrania tricuspidata branch extract showed a significant change in very low-density lipoprotein cholesterol (VLDL-cholesterol), with the greatest decrease observed in the group that consumed the lipid-lowering agent fenofibrate and a concentration-dependent decrease observed in the group that consumed the fermented Cudrania tricuspidata branch extract.

[0133] 2-6. Analysis of Atherosclerosis Index, Atherosclerosis Correlation Coefficient, Cardiac Risk Index, and Coronary Artery Index in a Hyperlipidemia Model

[0134] After the diet was completed for 50 days, serum from each group was isolated and biochemically analyzed to derive total cholesterol, triglycerides, HDL (high density lipoprotein), LDL (low density lipoprotein), and VLDL (very low density lipoprotein), and an analysis of four cardiovascular risk indicators using these was performed. Fig. 14This shows the effects of fermented CTSB extract and fenofibrate on the arteriosclerosis index (AI), arteriosclerosis coefficient (AC), cardiac risk ratio (CRR), and coronary artery index (CAI) in mice with hyperlipidemia and obesity.

[0135] The Atherogenic Index (AI) values ​​for each group were confirmed to be 0.85 ± 0.03 in the control group fed only a regular diet, 3.52 ± 0.09 in the negative control group fed only a regular diet after treatment with the hyperlipidemia obesity model, 1.12 ± 0.03 in the positive control group administered fenofibrate after treatment with the hyperlipidemia model (Model + fenofibrate group), 2.03 ± 0.09 in Experimental Group 1 (Model + CTSB500 group) administered a low concentration (500 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model, and 1.80 ± 0.07 in Experimental Group 2 (Model + CTSB1000 group) administered a high concentration (1000 mg / kg / day) of fermented Cudrania tricuspidata twig extract after treatment with the hyperlipidemia model. Fig. 14 A). When compared to a negative control group that consumed only a normal diet after treatment with a hyperlipidemia obesity model, the intake of the lipid-lowering agent fenofibrate significantly reduced the arteriosclerosis index, and the group that consumed fermented Cudrania tricuspidata branch extract showed a lower rate of reduction compared to the lipid-lowering agent, but showed a concentration-dependent reduction.

[0136] The atherogenic coefficient (AC) values ​​for each group were confirmed to be -0.06 ± -0.005 in the control group, 0.91 ± 0.04 in the negative control group, -0.16 ± 0.01 in the positive control group, 0.46 ± 0.03 in experimental group 1 (Model + CTSB500 group), and 0.30 ± 0.009 in experimental group 2 (Model + CTSB1000 group). Fig. 14B). When compared to a negative control group that consumed only a normal diet after treatment with a hyperlipidemia obesity model, the atherosclerosis correlation coefficient was significantly reduced by the intake of the lipid-lowering agent fenofibrate, and the concentration-dependent and effective atherosclerosis correlation coefficient reduction effect of fermented Cudrania tricuspidata branch extract was confirmed.

[0137] The cardiac risk ratio (CRR) values ​​for each group were confirmed to be 0.95 ± 0.06 in the control group, 1.67 ± 0.09 in the negative control group, 0.88 ± 0.07 in the positive control group, 1.34 ± 0.05 in experimental group 1 (Model + CTSB500 group), and 1.22 ± 0.04 in experimental group 2 (Model + CTSB1000 group). Fig. 14 C). When compared to a negative control group that consumed only a normal diet after treatment with a hyperlipidemia obesity model, the intake of the lipid-lowering agent fenofibrate significantly reduced the cardiac risk index, and the concentration-dependent and effective effect of the fermented Cudrania tricuspidata branch extract in reducing the cardiac risk index was confirmed.

[0138] The coronary artery index (CAI) values ​​for each group were confirmed to be 0.11 ± 0.01 in the normal diet control group, 1.37 ± 0.05 in the negative control group, 0.10 ± 0.01 in the positive control group, 0.75 ± 0.02 in experimental group 1 (Model + CTSB500 group), and 0.58 ± 0.02 in experimental group 2 (Model + CTSB1000 group). Fig. 14 D). When compared to a negative control group that consumed only a normal diet after treatment with a hyperlipidemia obesity model, the intake of the lipid-lowering agent fenofibrate significantly reduced the coronary artery index, and the concentration-dependent and effective coronary artery index reduction effect of the fermented Cudrania tricuspidata branch extract was confirmed.

Claims

Claim 1 A mixture of Cudrania tricuspidata branches and Cudrania tricuspidata fruit puree with Reishi mushrooms ( G. lucidum A health functional food for hyperlipidemia containing a 70% ethanol extract of a fermented product fermented with a strain of KACC 42231) as an active ingredient. Claim 2 delete Claim 3 delete Claim 4 delete Claim 5 delete Claim 6 delete Claim 7 delete Claim 8 delete Claim 9 delete Claim 10 delete

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