KR20240057057A - Anti-Obesity or Anti-Hyperlipidemic Composition Using Acertannin or Maplexin D - Google Patents

Abstract

The present invention discloses a composition for anti-obesity or anti-hyperlipidemia using asertanin or marplexin D, which inhibits the differentiation of 3T3-L1 mouse preadipocytes into adipocytes and fat accumulation in a concentration-dependent manner.

Description

Composition for anti-obesity or anti-hyperlipidemic use using acertannin or maplexin D {Anti-Obesity or Anti-Hyperlipidemic Composition Using Acertannin or Maplexin D}

The present invention relates to a composition for anti-obesity or anti-hyperlipidemia using acertannin or Maplexin D.

Obesity is an imbalance between energy intake and consumption, resulting in excessive accumulation of body fat and an increase in the number and size of fat cells (Cell.104:531-543 2001). Energy in the body is stored in fat cells in the form of triglyceride. When the stored energy source is depleted, the stored fat is broken down into free fatty acids and glycerol and used as an energy source. However, excessive intake of energy promotes the differentiation of fat cells and increases the amount of stored fat in the body, which is a direct cause of obesity. (Metabolism. 30: 425-427 1981; Biochem J. 1;435(3):723-32 2011).

Obesity not only changes body shape due to fat accumulation in the intestines and abdomen, but also acts as a risk factor that increases the incidence of various diseases (Korean J. Pediatr. 48:126-137. 2005). When visceral fat accumulates excessively, problems arise with sugar metabolism in the body and symptoms such as abnormal hormone secretion and cytokine secretion occur. The increase in triglycerides and LDL-cholesterol and the decrease in HDL-cholesterol due to obesity lead to abnormalities in fat metabolism in the body, decrease insulin receptors present in tissues, and also reduce insulin sensitivity, inhibiting the transport of glucose into cells. This can cause high blood sugar and diabetes. In addition, obesity is known to be closely related to the occurrence of metabolic diseases such as hyperlipidemia, cardiovascular disease, cancer, respiratory disorders, stroke, and osteoarthritis (Med. Int. 22 385-388 1994;. Ann. Intern. Med 103:1024-1029 1985).

Hyperlipidemia refers to a condition in which lipids such as cholesterol, phospholipids, and triglycerides are abnormally increased in the blood. Hyperlipidemia can appear in three forms: (i) hypercholesterolemia, (ii) hypertriglyceridemia, and (ii) a combination of these (hypercholesterolemia and hypertriglyceridemia). Hyperlipidemia itself does not show any unique symptoms, but if there is a lot of fat in the blood, it can stick to the walls of blood vessels, causing arteriosclerosis, and can cause angiocardia, myocardial infarction, cerebrovascular disease, and peripheral vascular occlusion (E. Falk et al. al., Circulation 92, 657-671, 1995).

Adipocytes are created by differentiating from preadipocytes. During adipocyte differentiation, neutral fat accumulation occurs and the expression of adipocyte-specific proteins (aP2, FAS, GLUT4, LPL, leptin) is induced, which includes PPARγ (peroxisome proliferator-activated receptor γ), C/EBP family (CCAAT/enhancer binding proteins; C/EBRα, C/EBRβ and C/EBRδ), ADD1/SREBP1c (adipocyte determination factor 1)/(sterol regulatory element binding protein 1c) It is known that transcription factors or adipocyte signaling substances (adipokines) play a central role (Genes De 2000, 14(11) 1293~1307)(Physiol Rev 1998, 78(3):783~809 ; Annu Rev Biochem 2008, 77:289-312; Genes Dev 1996, 10:1096-1107).

3T3-L1, a preadipocyte derived from a mouse embryo, is a pre-adipocyte that has been widely used as an in vitro model in drug screening for obesity, hyperlipidemia, etc. and in studying the mechanisms of lipid metabolism disorders (Biotechnology and Bioprocess Engineering 20(1):157-167, 2015; Prev Nutr Food Sci (17):1~7, 2012: J. Antibiot. 58(10): 634-39, 2005), morphological changes in cells, hormones They differentiate into adipocytes through a complex process involving sensitivity, gene and protein expression, etc. Early differentiation involves PPARγ (peroxisome proliferator-activated receptor γ) and the C/EBP family (CCAAT/enhancer binding proteins), C/EBRβ and C/EBRδ. Late differentiation involves C/EBRα and PPARγ (peroxisome proliferator-activated receptor γ). is known to play a central role (Genes De 2000, 14(11) 1293~1307)(Physiol Rev 1998, 78(3):783~809; Annu Rev Biochem 2008, 77:289~312; Genes Dev 1996, 10:1096-1107). During differentiation of these adipocytes, accumulation of neutral fat occurs and expression of adipocyte-specific proteins (aP2, FAS, GLUT4, LPL, leptin) is induced.

The present invention discloses the anti-obesity and anti-hyperlipidemic activities of asertanin or marplexin D.

The purpose of the present invention is to provide an anti-obesity composition using asertanin or marplexin D.

Another object of the present invention is to provide a composition for antihyperlipidemia using acertanin or marplexin D.

Another object or specific object of the present invention will be presented below.

In the present invention, as confirmed in the examples and experimental examples below, acertanin and marplexin D, isolated and identified from Siberian lanceolate, induce differentiation into adipocytes and fat accumulation of 3T3-L1, a mouse preadipocyte, at a concentration level. It is completed by confirming that it is suppressed dependently.

Considering the experimental results described above, the present invention in one aspect relates to (i) asertanin, its isomer, its prodrug, its solvate or its hydrate or (ii) marplexin D, its isomer, its prodrug. It can be viewed as an anti-obesity composition containing a drug, its solvate, or its hydrate as its active ingredient, and in another aspect, (i) asertanin, its isomer, its prodrug, its solvate, or its hydrate or ( ii) It can be viewed as an anti-hyperlipidemia composition containing Marplexin D, its isomer, its prodrug, its solvate, or its hydrate as its active ingredient.

The structural formulas of acertanin and maplexin D can be found in Chemical Formulas 1 and 2 below, respectively. Both are types of polyphenols contained in Acer ginnala , etc., and acertanin causes apoptosis in human dermal papilla cells. It is reported to have the effect of controlling apoptosis, and Marplexin D is reported to have high antioxidant activity (Kor. J. Pharmacogn. 49(1):7-14, 2018; Evid Based Complement Alternat Med. 2017 ;2017:6945912).

<Formula 1>

<Formula 2>

As used herein, isomers include essentially pure diastereomers, including optical isomers, as well as conformation isomers (i.e. isomers that differ only in the angle of one or more chemical bonds), and position isomers. (In particular, tautomers) or geometric isomers (eg, cis-trans isomers).

Also, in this specification, “prodrug” refers to a drug whose physical and chemical properties are adjusted by chemically changing a drug. It does not itself show physiological activity, but is affected by chemical or enzymatic action in the body after administration. It refers to a drug that can be converted into the original drug and exert medicinal effects.

Also, in this specification, “hydrate” refers to a compound to which water is bound, and includes embedded compounds in which there is no chemical bond between water and the compound.

Also, in this specification, “solvate” refers to a compound formed between a solute molecule or ion and a solvent molecule or ion.

In addition, in this specification, “active ingredient” refers to an ingredient that exhibits the desired activity alone or can exhibit activity in combination with a carrier that is not active on its own.

Also, in this specification, “anti-obesity” means reducing body fat, suppressing body fat accumulation, and/or reducing body weight. Therefore, it includes the prevention of obesity and the treatment of obesity, and further includes reducing body weight/body fat for beauty or health purposes (aka diet purposes) even though body weight is not classified as obesity or overweight.

In addition, in this specification, the term "obesity" refers to a state in which adipose tissue is abnormally increased, whether it is obesity due to genetic factors or obesity due to environmental factors, and according to the classification of body mass index (BMI), the term "obesity" refers to a state in which fat tissue is abnormally increased. This includes obesity (if BMI is 30.0 or more), obese (if BMI is 25 to 30), and overweight (if BMI is 23 to 25).

In addition, in this specification, “antihyperlipidemia” refers to lowering the concentration of neutral fat in the blood, lowering the concentration of cholesterol in the blood, or lowering both the concentration of neutral fat in the blood and the concentration of cholesterol in the blood.

The anti-obesity composition or anti-hyperlipidemia composition of the present invention (hereinafter "composition of the present invention") contains the active ingredients in an arbitrary amount (effective amount) depending on the use, formulation, purpose of formulation, etc., as long as it can exhibit anti-obesity activity and anti-hyperlipidemia activity. ), and a typical effective amount will be determined within the range of 0.001% by weight to 20.0% by weight based on the total weight of the composition. Here, “effective amount” means that when the composition of the present invention is administered to mammals, preferably humans, to which it is applied during the administration period recommended by medical experts, etc., the intended functional and pharmacological effects, such as anti-obesity effect and anti-hyperlipidemia effect, are achieved. It refers to the amount of active ingredient contained in the composition of the present invention that can be expressed. Such effective amounts can be determined experimentally within the scope of the ordinary ability of those skilled in the art.

In addition to the active ingredients, the composition of the present invention has already been proven safe in the art for enhancing and reinforcing the anti-obesity effect or anti-hyperlipidemia effect, or for improving the convenience of taking or ingesting through the addition of similar activities such as blood pressure regulating activity. It may further include any compounds or natural extracts that have been validated and known to have the corresponding activity.

These compounds or extracts include compounds, extracts, and pharmaceuticals listed in compendial documents such as the Pharmacopoeia of each country (in Korea, the “Korean Pharmacopoeia”) and the Code of Health Functional Foods of each country (in Korea, it is the “Standards and Specifications for Health Functional Foods” notified by the Ministry of Food and Drug Safety). The laws of each country that govern the manufacture and sale of compounds or extracts and health functional foods that have received product approval in accordance with the laws of each country (in Korea, this is the Pharmaceutical Affairs Act) (in Korea, it is the “Health Functional Foods Act”) 」) includes compounds or extracts whose functionality has been recognized.

For example, according to Korea's Health Functional Foods Act, Garcinia cambogia bark extract, conjugated linolenic acid (free fatty acid), conjugated linolenic acid (triglyceride), green tea extract, chitosan, and Lactobacillus gasseri are recognized as functional ingredients for 'body fat reduction'. BNR17 ( Lactobacillus gasseri BNR17), L-carnitine tartrate, green mate extract, green coffee bean extract, perilla leaf extract, soybean germ extract, etc. complex, ginseng leaf alcohol extract powder, lactoferrin (milk purified protein), lemon balm extract mixed powder , mate thermal water extract, seaweed and other complex extracts (xanthigen), fermented vinegar pomegranate complex, pu-erh tea extract, soybean peptide complex, vegetable oil diglyceride, wild mango seed extract, medium chain fatty acid (MCFA)-containing oil, Complex extracts such as coleus forskohlli extract, chitooligosaccharide, finger root extract powder, hibiscus, etc., powder containing GABA derived from L-glutamic acid recognized as a 'blood pressure regulation' functional ingredient, katsuobushi oligopeptide, natto bacteria culture powder, Soybean) peptide complex, salmon peptide, olive leaf extract, sardine peptide, casein hydrolyzate, coenzyme Q10, grape seed enzymatically decomposed extract powder, Haitai oligopeptide, etc., DHA concentrated oil and globin singer recognized as functional ingredients for 'improving blood neutral fat' Decomposition products, indigestible maltodextrin, bamboo leaf extract, vegetable oil diglyceride, sardine refined fish oil, refined squid oil, etc., L-arabinose, nopal extract, cinnamon extract powder, guava leaf extract, Indigestible maltodextrin, freeze-dried silkworm powder, hemp spirit extract, banaba leaf extract, leaf extract, etc., fermentation-generated amino acid complex recognized as a functional ingredient for 'improving fatigue', Hovenia vulgaris fruit extract, rhodiola extract, etc., with 'anti-stress' functionality These compounds or extracts include L-theanine, asiaganda extract, milk protein hydrolyzate, and ginseng leaf extract, which are recognized as ingredients.

The composition of the present invention can be considered a food composition in a specific aspect.

The food composition of the present invention can be manufactured in any form, for example, beverages such as tea, juice, carbonated beverages, and electrolyte drinks, processed oils such as milk and yogurt, gums, rice cakes, Korean snacks, bread, snacks, noodles, etc. It can be manufactured into health functional food preparations such as foods, tablets, capsules, pills, granules, liquids, powders, flakes, pastes, syrups, gels, jellies, bars, etc.

In addition, the food composition of the present invention can have any product classification in terms of legal and functional classification as long as it complies with the enforcement laws and regulations at the time of manufacture and distribution. For example, it is a health functional food according to the Korean 「Act on Health Functional Foods」, or confectionery, beans, tea, and beverages according to each food type according to the food code of the Korean 「Food Sanitation Act」 (Ministry of Food and Drug Safety Notification 「Food Standards and Specifications」) , special purpose food, etc.

The food composition of the present invention may contain food additives in addition to the active ingredients. Food additives can generally be understood as substances that are added to, mixed with, or infiltrated into food when manufacturing, processing, or preserving food. Since they are consumed daily and for a long period of time with food, their safety must be guaranteed. In the food additive code of each country that regulates the manufacturing and distribution of food (in Korea, it is the Food Sanitation Act), food additives with guaranteed safety are limited in terms of ingredients or functions. In the Korea Food Additive Code (Ministry of Food and Drug Safety Notification “Food Additive Standards and Specifications”), food additives are classified into chemical synthetics, natural additives, and mixed preparations in terms of composition. These food additives are classified into sweeteners and flavors in terms of function. It is classified into preservatives, emulsifiers, acidulants, thickeners, etc.

Sweeteners are used to impart an appropriate sweetness to foods, and both natural and synthetic ones can be used in the food composition of the present invention. Preferably, a natural sweetener is used, and natural sweeteners include sugar sweeteners such as corn syrup solids, honey, sucrose, fructose, lactose, and maltose.

Flavoring agents are used to improve taste or aroma, and both natural and synthetic ones can be used. Preferably, natural products are used. When using natural products, they can serve the purpose of enhancing nutrition in addition to flavor. Natural flavoring agents may be obtained from apples, lemons, tangerines, grapes, strawberries, peaches, etc., or may be obtained from green tea leaves, coriander leaves, bamboo leaves, cinnamon, chrysanthemum leaves, jasmine, etc. You can also use things obtained from ginseng (red ginseng), bamboo shoots, aloe vera, and ginkgo nuts. Natural flavoring agents may be liquid concentrates or solid extracts. In some cases, synthetic flavoring agents may be used, and as synthetic flavoring agents, esters, alcohols, aldehydes, terpenes, etc. may be used.

Preservatives include calcium sorbate, sodium sorbate, potassium sorbate, calcium benzoate, sodium benzoate, potassium benzoate, EDTA (ethylenediaminetetraacetic acid), etc., and emulsifiers include acacia gum, carboxymethyl cellulose, xanthan gum, and pectin. etc. may be used, and as acidulants, acidic acid, malic acid, fumaric acid, adipic acid, phosphoric acid, gluconic acid, tartaric acid, ascorbic acid, acetic acid, phosphoric acid, etc. may be used. In addition to improving taste, acidulants may be added to ensure that the food composition has an appropriate acidity for the purpose of suppressing the growth of microorganisms. As a thickening agent, a suspending agent, settling agent, gel forming agent, bulking agent, etc. may be used.

In addition to the food additives described above, the food composition of the present invention may contain bioactive substances or minerals known in the art and whose safety is guaranteed as food additives for the purpose of supplementing and reinforcing functionality and nutrition.

Such physiologically active substances include catechins contained in green tea, vitamins such as vitamin B1, vitamin C, vitamin E, and vitamin B12, tocopherol, dibenzoylthiamine, etc., and minerals include calcium preparations such as calcium citrate and magnesium stearate. Magnesium preparations such as iron citrate, iron preparations such as chromium chloride, potassium iodine, selenium, germanium, vanadium, zinc, etc. are included.

The food composition of the present invention may contain the above-described food additives in an appropriate amount to achieve the purpose of addition depending on the product type.

Regarding other food additives that can be included in the food composition of the present invention, each country's food code or food additive code can be referred to.

The composition of the present invention may be considered a pharmaceutical composition in other specific embodiments.

The pharmaceutical composition of the present invention contains a pharmaceutically acceptable carrier in addition to the active ingredient and can be prepared into an oral formulation or a parenteral formulation depending on the route of administration by a conventional method known in the art. Here, the route of administration may be any suitable route, including topical route, oral route, intravenous route, intramuscular route, and direct absorption through mucosal tissue, and two or more routes may be used in combination. An example of a combination of two or more routes is when two or more types of drugs according to the administration route are combined, for example, when one drug is administered firstly through an intravenous route and the other drug is administered secondarily through a local route.

Pharmaceutically acceptable carriers are well known in the art depending on the route of administration or dosage form, and for specific details, reference can be made to the pharmacopoeia of each country, including the Korean Pharmacopoeia.

When the pharmaceutical composition of the present invention is prepared as an oral dosage form, it can be prepared as powder, granules, tablets, pills, sugar-coated tablets, capsules, solutions, gels, syrups, suspensions, and wafers along with a suitable carrier according to methods known in the art. It can be manufactured in a dosage form such as: Examples of suitable carriers include sugars such as lactose, glucose, sucrose, dextrose, sorbitol, mannitol, and xylitol, starches such as corn starch, potato starch, and wheat starch, cellulose, methylcellulose, ethylcellulose, sodium carboxymethylcellulose, Cellulose such as hydroxypropylmethylcellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, magnesium stearate, mineral oil, malt, gelatin, talc, polyol, vegetable oil, ethanol, grease. Serol, etc. can be mentioned. When formulating, appropriate binders, lubricants, disintegrants, colorants, diluents, etc. can be included as needed. Suitable binders include starch, magnesium aluminum silicate, starch ferrite, gelatin, methylcellulose, sodium carboxymethylcellulose, polyvinylpyrrolidone, glucose, corn sweetener, sodium alginate, polyethylene glycol, and wax, and as a lubricant, oleic acid. Examples include sodium, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride, silica, talcum, stearic acid, its magnesium and calcium salts, and polydethylene glycol. Disintegrants include starch and methyl cellulose. , agar, bentonite, xanthan gum, starch, alginic acid or its sodium salt, etc. Additionally, diluents include lactose, dextrose, sucrose, mannitol, sorbitol, cellulose, glycine, etc.

When the pharmaceutical composition of the present invention is prepared as a parenteral formulation, it can be formulated in the form of injections, transdermal administration, nasal inhalation, and suppositories along with a suitable carrier according to methods known in the art. When formulated as an injection, an aqueous isotonic solution or suspension can be used as a suitable carrier. Specifically, an isotonic solution such as PBS (phosphate buffered saline) containing triethanolamine, sterile water for injection, or 5% dextrose can be used. . When formulated for transdermal administration, it can be formulated in the form of ointments, creams, lotions, gels, external solutions, paste preparations, linear preparations, and aerol preparations. In the case of nasal inhalation, it can be formulated in the form of an aerosol spray using suitable propellants such as dichlorofluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, and carbon dioxide. When formulated as a suppository, the carrier is Wethepsol ( witepsol), Tween 61, polyethylene glycols, cocoa fat, laurel paper, polyoxyethylene sorbitan fatty acid esters, polyoxyethylene stearates, sorbitan fatty acid esters, etc. can be used.

Regarding the specific formulation of pharmaceutical compositions, it is known in the art, and references can be made to, for example, Remington's Pharmaceutical Sciences (19th ed., 1995). The above documents are considered a part of this specification.

The preferred dosage of the pharmaceutical composition of the present invention is in the range of 0.001 mg/kg to 10 g/kg per day, preferably 0.001 mg/kg to 1 g, depending on the patient's condition, weight, gender, age, patient's severity, and administration route. It may be in the /kg range. Administration can be done once a day or divided into several times. These dosages should not be construed as limiting the scope of the invention in any respect.

In another aspect, the present invention relates to a method for isolating acertanin or marplexin D from a thorn vine extract.

The method of the present invention includes the steps of (a) preparing a solid extract by immersing and extracting the Bluethorn Vine plant in an extraction solvent mixed with water and ethanol, and then removing the extraction solvent; (b) preparing the solid extract in distilled water; After suspending in , sequentially fractionating with dichlormethane, ethyl acetate, and n-butanol, and then concentrating under reduced pressure to obtain an ethyl acetate fraction, and (c) performing chromatography on the ethyl acetate fraction to produce acertanin or It includes the step of isolating marplexin D.

In the method of the present invention, the extraction solvent mixed with water and ethanol in step (a) is preferably 70% ethanol.

In addition, in the method of the present invention, step (c) is performed using a mixed solvent of methanol and water (20%, 40%, 60%, 80%) in a column filled with (c-1) Diaion HP-20 resin. , and 100% methanol) as a mobile phase, performing chromatography and dividing into five fractions; and (c-2) performing chromatography on the 40% methanol fraction among the five fractions to separate acertanin or maplexin D. Here, Diaion HP-20 resin is a high porous type synthetic adsorbent made of a copolymer of styrene and DVB (Divinyl benzene), and has a large number of pores distributed. It is a resin that is used for adsorption, separation, and purification of various biologically active substances such as vitamins, antibiotics, steroids, and enzymes because it has a high specific surface area and excellent adsorption ability.

As described above, according to the present invention, a composition for anti-obesity or anti-hyperlipidemia using asertanin or marplexin D can be provided. The composition for anti-obesity or anti-hyperlipidemia of the present invention can be commercialized into drugs, functional foods, etc.

Figure 1 is a schematic diagram of the separation process of acertanin and marplexin D from a thorn vine extract.
Figures 2 and 3 show ¹ H NMR, ¹³ H NMR, ¹ H- ¹ H COZY, and HMBC spectral data of acertanine and marplexin D, respectively.
Figure 4 shows the results of evaluating the degree of fat accumulation in preadipocytes.

Hereinafter, the present invention will be described with reference to examples. However, the scope of the present invention is not limited to these examples.

<Example> Isolation, identification, and anti-obesity and anti-hyperlipidemia activity tests of acertanin and marplexin D

<Example 1> Isolation and identification of acertanin and marplexin D from Siberian thorn vine extract

Acertannin and maplexin D were isolated from Smilax sieboldii extract. First, the green thorn vine outpost?? 404 g of the 70% ethanol extract of the dried product was suspended in 10 L of distilled water, sequentially fractionated with equal amounts of dichloromethane, ethyl acetate, and saturated n-butanol, and then concentrated under reduced pressure to obtain each fraction. For 25 g of the ethyl acetate fraction, 5 sub-fractions (G99- 12-1 to G99-12-5) were obtained, and for subfraction No. 2 (G99-12-2), the mobile phase was acetonitrile-water gradient elution (0 min; 5% acetonitrile, 110 min; 30% aceto). Reverse phase column (RP-18) medium pressure chromatography was performed using nitrile to obtain 12 small fractions (G99-20-1 to G99-20-12). For sub-fraction No. 7 (G99-20-7), acetonitrile-water as the mobile phase was eluted using a reversed-phase column (0 min; 15% acetonitrile, 50 min; 27% acetonitrile, flow rate: 10 mL/min). RP-18) Preparative HPLC was performed to separate and purify the active compounds <Formula 1> and <Formula 2>. A schematic diagram of the separation process of the above compound is shown in [Figure 1]. As can be seen in [FIG. 2] and [FIG. 3] below, the two isolated compounds are each of the <Formula 1> from ¹ H NMR, ¹³ H NMR, ¹ H- ¹ H COZY, and HMBC spectrum data. Asertanin (2,6-di- O -galloyl-1,5-anhydro-D-glucitol), Marplexin D (2,4-di- O -galloyl-1,5- of <Formula 2>) each was identified as anhydro-D-glucitol). The molecular formula, molecular weight, CASRN (CAS Registry Number), etc. of each chemical can be confirmed in [Figure 2] and [Figure 3] (Code No. and Remark in Figures 2 and 3 are reference numbers for each compound of the inventor. ).

< Example 2> Anti-obesity, anti-hyperlipidemia activity test

3T3-L1 (Zenbio SP-L1-F), a mouse preadipocyte, was cultured in 3T3-L1 Preadipocyte Medium (Zenbio PM-1-L1) at 37°C and 5% CO2. 3T3-L1 preadipocytes were distributed in a 24 well plate at a cell count of 5×10 ⁴ cells/well and maintained for 2 more days when 100% confluency was reached. Preadipocyte differentiation was induced for 2 days with 3T3-L1 Differentiation Medium (Zenbio DM-2-L1) containing MDI (Isobutylmethylxanthine, dexamethasone, insulin), and cultured for 48 hours, every 2 days for 6 days. The culture medium was replaced with 3T3-L1 Adipocyte Medium (Zenbio AM-1-L1). During the induction of adipocyte differentiation, samples were treated with each culture medium at different concentrations, and the degree of adipocyte differentiation was observed on day 8, when differentiation was completed. The degree of adipocyte differentiation was first confirmed under a microscope through Oil Red O staining, and the degree of adipocyte staining was measured by measuring the amount of differentiated fat in the cells at 510 nm absorbance.

The results are shown in [Figure 4]. Both of the above two compounds inhibited neutral fat accumulation in preadipocytes in a concentration-dependent manner.

Claims (6)

For anti-obesity use containing (i) asertanin, its isomer, its prodrug, its solvate, or its hydrate, or (ii) Maplexin D, its isomer, its prodrug, its solvate, or its hydrate as an active ingredient. Pharmaceutical composition.
For anti-obesity use containing (i) asertanin, its isomer, its prodrug, its solvate, or its hydrate, or (ii) Maplexin D, its isomer, its prodrug, its solvate, or its hydrate as an active ingredient. Food composition.
An antihyperlipidemic pharmaceutical containing i) asertanin, its isomer, its prodrug, its solvate, or its hydrate, or (ii) Maplexin D, its isomer, its prodrug, its solvate, or its hydrate as an active ingredient. enemy composition.
For antihyperlipidemia containing (i) asertanin, its isomer, its prodrug, its solvate, or its hydrate or (ii) Maplexin D, its isomer, its prodrug, its solvate, or its hydrate as an active ingredient. Food composition.
(a) preparing a solid extract by immersing and extracting the Bluethorn Vine plant in an extraction solvent mixed with water and ethanol, and then removing the extraction solvent;
(b) suspending the solid extract in distilled water, sequentially fractionating it with dichloromethane, ethyl acetate, and normal butanol, and then concentrating under reduced pressure to obtain an ethyl acetate fraction, and
(c) A method of isolating acertanin or maplexin D from a Sertanin extract, comprising the step of performing chromatography on the ethyl acetate fraction to separate asertanin or maplexin D.
According to clause 5,
The extraction solvent mixed with water and ethanol in step (a) is 70% ethanol,
In step (c), a mixed solvent of methanol and water (20%, 40%, 60%, 80%, and 100% methanol) is used in a column filled with (c-1) Diaion HP-20 resin. Using the mobile phase, performing chromatography and dividing into five fractions; and (c-2) performing chromatography on a 40% methanol fraction among the five fractions to separate acertanin or maplexin D.