KR20200028108A - Composition comprising flavonoid derivatives isolated from Sicyos angulatus for preventing or treating liver diseases - Google Patents

Abstract

The present invention relates to a composition for preventing or treating liver diseases containing a flavonoid derivative isolated from Sicyos angulatus as an active component, wherein the flavonoid derivative prevents excessive accumulation of fat in liver cells, and reduces hepatocyte toxicity by alcohol. Therefore, the flavonoid derivative isolated from Sicyos angulatus of the present invention is expected to be usefully used for developing a composition and animal drugs for preventing or treating liver diseases, and health functional food and animal feed additives for alleviating liver diseases.

Description

Composition comprising flavonoid derivatives isolated from Sicyos angulatus for preventing or treating liver diseases}

The present invention relates to a composition for preventing or treating liver disease comprising a flavonoid derivative isolated from Sicyos angulatus as an active ingredient.

In modern society, the obesity population is rapidly increasing due to the convenient living environment due to rapid automation, the increase in food intake, and the reduction in excess nutrition and physical activity due to the increase in eating out. Obesity is widely known to cause chronic diseases such as fatty liver, hypertension, diabetes, and cardiovascular diseases. Recently, 30.7% of Korean adults reported obesity as of 2011, according to national indicators that provide national statistical data. In addition, 1.7 billion people, which account for about 25% of the world's population, are currently overweight (BMI) (25), and 300 million people in the Western region, including 120 million from major countries such as the United States, Europe, and Japan. The above is classified as obese patients (BMI> 30). In the case of children, 1 out of 5 children worldwide is considered obese, and this number is rapidly increasing, making obesity a serious social problem.

Population aging is one of the most important social issues in the world, and with the rapid increase of the aging population, increasing senile diseases and consequently increasing medical expenses are emerging as essential social issues to be solved.

The increase in obesity patients and the increase in aging means that obesity, fatty liver, diabetes, hyperlipidemia, hypertension and cardiovascular disease, which are diseases caused by obesity, increase and the time to suffer from these diseases increases. Therefore, there is an emphasis on the need to develop a therapeutic agent capable of controlling the stage of occurrence of obesity-related diseases and the long-term symptom deterioration process.

Fatty liver (fatty liver) refers to the accumulation of fat in the liver cells due to excessive intake of fat or endogenously increased fat synthesis or excretion in the liver. Fatty liver can be classified into alcoholic fatty liver disease and non-alcoholic fatty liver disease (Yoon, S.K., 2009).

Alcoholic fatty liver disease is the main cause of excessive alcohol intake, which is related to the total intake than the type of alcohol, and even if the same amount is consumed continuously every day, the efficiency of alcohol metabolism decreases and fatty liver occurs more. Alcoholic fatty liver does not stop with fatty liver, but 10 to 35% progress to alcoholic hepatitis, and 8 to 20% progress to alcoholic cirrhosis and eventually lead to liver cancer or end-stage liver failure (Yoon, S.K., 2009).

Non-alcoholic fatty liver disease (NAFLD) is a condition in which excess triglyceride is accumulated in the liver, simple steatosis with only excessive accumulation of fat in the liver, necrosis and inflammation of the liver. It refers to a group of diseases, including non-alcoholic steatohepatitis (NASH) with hyperfibrosis and more advanced cirrhosis (Brunt, EM, 2001). The majority of patients with high prevalence of NAFLD have a simple clinical condition with no significant fibrosis and good clinical progress. However, the severity of the problem is that NASH occurs in about 10-20% of NAFLD patients, and progresses to cirrhosis in about 9-25% of these NASH patients. When cirrhosis develops, the risk of developing liver cancer increases, and after 30 years, 30-40% die from complications of liver disease (Ekstedt, M, et al., 2006; Fassio, E., et al., 2004; Adams, LA, et al., 2005).

Recently, NAFLD is considered as a form of metabolic syndrome in the liver such as obesity, hypertension, type 2 diabetes, and lipid metabolism based on insulin resistance (Sohn, J.H., et al, 2010). Accumulation of triglycerides in the liver is a representative indicator of NAFLD, which appears in the mechanism of NAFLD development, and until now, several mechanisms related to lipid accumulation in the liver are known, but peripheral fatty acids are increased along with increased fatty acid synthesis in the liver. It has been reported that NAFLD plays the largest role in lipid accumulation in the liver (Tilg, H., et al., 2010).

The mechanism of development of NAFLD is particularly closely related to metabolic syndrome, including obesity. The liver plays an important role in carbohydrate metabolism because it is responsible for blood sugar balance through glycogen production and degradation. About 96% of patients with cirrhosis are sugar-resistant, and 30% are clinically diagnosed with diabetes, explaining the importance of mechanisms regulating fat accumulation in the liver. Fatty liver disease is generally associated with insulin resistance, since soluble mediators of immune cells and free fatty acids are usually involved in insulin control systems and NAFLD expression. That is, several inflammatory cytokines, including TNF-α (tumor necrosis factor-α) and IL-6 (interleukin-6) produced by free fatty acids, are an important starting point for the development of a series of liver diseases such as NAFLD, NASH and cirrhosis. . Therefore, the development of NAFLD therapeutics is urgent because NAFLD is rapidly increasing due to the recent spread of metabolic syndrome and population aging.

Prickly pear (Sicyos angulatus L. , Elariaium trifoliatum L.) belongs to the genus Sicyos, and is one of the most diverse genus in the 40 species of Cucurbitaceae. Prickly pear is a plant that grows rapidly by covering the surrounding soil with rapid growth and pushing out other plants. Even in Korea, thornstalk was introduced as a grafted plant for cultivation of cucumbers, etc., but the rapid growth of thornstalk was vigorous enough to dominate Korea's wetland environment and was designated as an environmentally disturbing species designated by the Ministry of Environment. However, before flowering, the young leaves of this plant are listed as edible raw materials in Korean Food Standard Codex (KFSC), so if you are looking for new functionalities based on them, they may be used as raw materials for functional food. However, although it is a plant commonly found in Korea, it is recognized as a target for removal despite its strong vitality and abundant plant resources, and studies related to it are incomplete. Recently, active research of prickly pear has been started, and an attempt is made to separate active substances from prickly pear (Kim, YA, et al., 2017).

Thus, the present inventors were able to complete the present invention by separating the flavonoid derivatives from the spinach extracts in the course of studying the spinach extract, and confirming that the separated flavonoid derivatives are excellent in treating liver diseases.

As a prior art, Korean Registered Patent No. 1802970 describes a composition for the treatment of liver disease including a spinach extract, but the flavonoid derivative isolated from the spinach of the present invention and its liver disease treatment effect are not described. In addition, Korean Registered Patent No. 1849624 describes a biflavonoid derivative and its non-alcoholic fatty liver treatment effect, but is different from the flavonoid derivative of the present invention.

Korean Registered Patent No. 1802970, Composition for the treatment or prevention of liver disease comprising thorn extract as an active ingredient, Nov. 23, 2017. Korean Registered Patent No. 1849624, Composition for preventing, improving or treating metabolic diseases caused by lipid imbalance, including biflavonoid derivatives or salts thereof, registered on April 11, 2018.

Adams, L.A., et al., The natural history of nonalcoholic fatty liver disease: a population-based cohort study, Gastroenterology, 129 (1), 113-121, 2005. Brunt, E.M., Nonalchololic steatohepatitis: definition and pathology, Semin. Liver Dis., 21 (1), 3-16, 2001. Cameron, R.G., et al., Modulation of liver-specific cellular response to ethanol in vitro in hepG2 cells, Toxicol In Vitro., 12 (2), 111-122, 1998. Ekstedt, M., et al., Long-term follow-up patients with NAFLD and elevated liver enzymes, Hepatology, 44 (4), 865-873, 2006. Fassio, E., et al., Natural history of nonalcoholic steatohepatitis: a longitudianl study of repeat liver biopsies, Hepatology, 40 (4), 820-826, 2004. Kim, Y.A., et al., Antioxidant Activity and Anti-inflammatory effects of Sicyos angulatus L. extract, Journal of Oil & Applied Science, 34 (3), 536-544, 2017. Madushani, H.K., et al., Phenolic acid and flavonoid-rich fraction of Sasa quelpaertensis Nakai leaves prevent alcohol induced fatty liver through AMPK activation, J. Ethnopharmacol., 224, 335-348, 2018. Sohn, J.H., et al., Recent update on pathogenesis of nonalcholic fatty liver disease, The Korean Journal of Medicine, 79 (5), 461-474, 2010. Tilg, H., et al., Evolutin of Inflammation in Nonalcoholic Fatty Liver Disease: L The Multiple Parallel Hits Hypothesis, Hepatology, 52 (5), 1836-1846, 2010. Yoon, S.K., Diagnosis and treatment of fatty liver, The Korean Journal of Medicine, 76 (6), 677-679, 2009.

An object of the present invention is to provide a composition for preventing or treating liver disease comprising a flavonoid derivative isolated from prickly pear (Sicyos angulatus ) as an active ingredient.

Another object of the present invention is to provide a novel flavonoid derivative isolated from spiny foil and a method for separating the same.

The present invention is the following camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1), camphor Roll-3-O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 2), camphorol-3-O- (6 -O-α-L-ramnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 3), camphorol-3-O-β-D-glucopyra Nosyl-7-O-α-L-lamnopyranoside (Compound 4), apigenin-7-O-β-D-glucopyranoside (Compound 5), camphorol-3-O-β-D- Glucopyranoside (Compound 6), Camperol-3-O- (6-O-α-L-lamnopyranosyl) -β-D-Glucopyranoside (Compound 7), Camperol-3-O- β-D-Apiofuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 8), quercetin-3-O- (6-OE -Feruloyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 9), quercetin-3-O- (6-OE-coumaroyl) -β-D -Glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 10), Perol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L-lamnopyranoside (Compound 11), camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β- D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7- Prevention of liver disease comprising at least one flavonoid derivative compound selected from the group consisting of O-α-L-lamnopyranosyl- (1-4) -β-D-glucopyranoside (Compound 13), or It relates to a pharmaceutical composition for treatment, a health functional food for improvement, an animal drug for prevention or treatment, and an animal feed additive for improvement.

[Formula 1]

  The flavonoid derivative compound is preferably camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1), Camperol-3-O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 2), camperol-3-O- ( 6-O-α-L-lamnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 3), camphorol-3-O-β-D-gluco Pyranosyl-7-O-α-L-lamnopyranoside (Compound 4), camphorol-3-O-β-D-glucopyranoside (Compound 6), camphorol-3-O- (6- OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L -Ramnopyranoside (Compound 11), Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1- 3) -α-L-lamnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R-guaiacglyceryl) ether 7-O-α-L-lamnopyranosyl -(1-4) -β-D-glucopyranoside (Compound 1 It may be one or more flavonoid derivative compounds selected from the group consisting of 3).

The flavonoid derivative compound may be isolated from the extract of thorns ( Sicyos angulatus ).

The spinach extract is a water, C1 ~ 4 lower alcohol, C1 ~ 4 acetic acid (acetic ester), acetone (acetone), methyl ethyl ketone (methylethylketone) at least one solvent selected from the group consisting of It may be an extracted extract.

The liver disease may be a fatty liver disease.

The fatty liver disease may be non-alcoholic fatty liver disease (NAFLD) or alcoholic fatty liver disease.

In addition, the present invention solvents one or more selected from the group consisting of water, C1-4 lower alcohol, C1-4 acetic ester, acetone, and methylethylketone. Preparing a thorny berry extract as; Hexane, ethyl acetate and butanol are sequentially added to the spinach extract to obtain a fraction; And chromatographing each fraction to form camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (1) Compound 1), camperol-3-O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 2), camperol-3 -O- (6-O-α-L-ramnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 3), camphorol-3-O-β -D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 4), apigenin-7-O-β-D-glucopyranoside (Compound 5), camphorol-3-O -β-D-glucopyranoside (Compound 6), camphorol-3-O- (6-O-α-L-ramnopyranosyl) -β-D-glucopyranoside (Compound 7), camphorol -3-O-β-D-apiofuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 8), quercetin-3-O -(6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 9), quercetin-3-O- (6-OE-coumaroyl ) -β-D-glucopyranosyl-7-O- -L-lamnopyranoside (Compound 10), camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranosyl- ( 1-6) -β-D-glucopyranosyl- (1-3) -α-L-lamnopyranoside (Compound 11), camphorol-3-O- (6-OE-feruloyl) -β -D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 12) and trisine 4'-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α-L-lamnopyranosyl- (1-4) -β-D-glucopyranoside (Compound 13) It relates to a method of isolating a flavonoid derivative compound.

The present invention is also a novel compound having the chemical structure of Formula 2, camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-ram Nopyranoside (Compound 1), camphorol-3-O-β-D-apiofuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 8), Quercetin-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 9), camphorol-3 -O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranosyl- (1-6) -β-D-glucopyranosyl- (1- 3) -α-L-lamnopyranoside (Compound 11), camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-gluco Pyranosyl- (1-3) -α-L-ramnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α -L-lamnopyranosyl- (1-4) -β-D-glucopyranoside (Compound 13).

[Formula 2]

Hereinafter, the present invention will be described in detail.

The present invention relates to a method for separating a flavonoid derivative compound and a composition for preventing or treating liver disease comprising the flavonoid derivative compound isolated from an extract of spiny pear (Sicyos angulatus ) as an active ingredient.

The flavonoid derivative compound is camphorol-3-O- (6-OE-feruloyl) of Chemical Formula 1-β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1). , Camphorol-3-O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 2), camphorol-3-O- (6-O-α-L-lamnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 3), camphorol-3-O-β-D- Glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 4), apigenin-7-O-β-D-glucopyranoside (Compound 5), camphorol-3-O-β- D-glucopyranoside (Compound 6), camphorol-3-O- (6-O-α-L-ramnopyranosyl) -β-D-glucopyranoside (Compound 7), camphorol-3- O-β-D-apiofuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 8), quercetin-3-O- (6 -OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 9), quercetin-3-O- (6-OE-coumaroyl) -β -D-glucopyranosyl-7-O-α-L-ram Nopyranoside (Compound 10), camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 11), camphorol-3-O- (6-OE-feruloyl) -β-D-gluco Pyranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R- Guaiacylglyceryl) ether 7-O-α-L-lamnopyranosyl- (1-4) -β-D-glucopyranoside (Compound 13) One or more flavonoid derivative compounds selected from the group consisting of You can. Preferably camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 1), camphorol-3- O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 2), camphorol-3-O- (6-O-α -L-lamnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 3), camphorol-3-O-β-D-glucopyranosyl-7- O-α-L-lamnopyranoside (Compound 4), camperol-3-O-β-D-glucopyranoside (Compound 6), camperol-3-O- (6-OE-feruloyl ) -β-D-glucopyranosyl-7-O-α-L-ramnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 11), Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1-3) -α- L-lamnopyranoside (Compound 12) and trisine 4'-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α-L-lamnopyranosyl- (1-4 ) -β-D-glucopyranoside (Compound 13) It is on the flavonoid derivatives.

The spinach extract is a water, C1 ~ 4 lower alcohol, C1 ~ 4 acetic acid (acetic ester), acetone (acetone), methyl ethyl ketone (methylethylketone) at least one solvent selected from the group consisting of It may be an extracted extract. Methanol, ethanol, propanol, isopropanol, butanol, etc. may be used as the lower alcohol of C1-4, and methyl acetate, ethyl acetate, propyl acetate (propyl acetate) propyl acetate), isopropyl acetate, butyl acetate, isobutyl acetate, and the like can be used. It is preferably ethanol, and more preferably 70% [v / v] ethanol.

The solvent may add a weight of 5 to 100 times the weight of the visible foil. Preferably, 5 to 40 times the weight is added.

In addition, the extract of thorns may be a fraction re-fractionated with an organic solvent. As the organic solvent, C1-4 lower alcohol, ethyl acetate, hexane and acetone may be used, and the C1-4 lower alcohol may be methanol, ethanol, propanol, isopranol, butanol, and the like. Preferably, it may be a fraction fractionated by adding hexane, ethyl acetate and butanol, and more preferably, a fraction fractionated by sequentially adding hexane, ethyl acetate and butanol.

The flavonoid derivatives Compounds 1 to 13 can be separated by using the chromatography of the spinach extract. The chromatography is silica gel column chromatography, reverse phage-18 (RP-18) column chromatography, high-performance liquid chromatography (HPLC), Sepha In dex LH-20 column chromatography, thin layer chromatography (TLC), medium pressure liquid chromatography, ion exchange resin chrmatography, etc. You can choose and use.

Meanwhile, the flavonoid derivative compound of the present invention may be synthesized according to a conventional method in the art, and may be prepared with a pharmaceutically acceptable salt.

In addition, the present invention provides a pharmaceutical composition for preventing or treating liver disease comprising at least one flavonoid derivative compound selected from the group consisting of compounds 1 to 13 of Formula 1 as an active ingredient.

The pharmaceutical composition may include at least one flavonoid derivative compound selected from the group consisting of compounds 1 to 13 of Formula 1 and a pharmaceutically acceptable excipient or carrier.

The pharmaceutical composition may be formulated and used in the form of oral dosage forms, external preparations, suppositories, and sterile injectable solutions, such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., according to a conventional method. Carriers, excipients and diluents that may be included in the pharmaceutical composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, Cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. In the case of formulation, it is prepared using diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents and surfactants. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc. These solid preparations are at least one excipient in at least one flavonoid derivative compound selected from the group consisting of compounds 1-13 of the present invention. For example, it is prepared by mixing starch, calcium carbonate, sucrose or lactose, gelatin, and the like. Also, lubricants such as magnesium stearate and talc are used in addition to simple excipients. Liquid preparations for oral use include suspensions, intravenous solutions, emulsions, syrups, etc. In addition to water and liquid paraffin, which are common diluents, various excipients, such as wetting agents, sweeteners, fragrances, and preservatives, can be included. . Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, and suppositories. Non-aqueous solvents and suspensions may include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate. As a base for suppositories, witepsol, macrogol, tween 61, cacao butter, laurin butter, and glycerogelatin may be used.

The dosage of the pharmaceutical composition will vary depending on the age, gender, weight of the subject to be treated, the specific disease or pathology to be treated, the severity of the disease or pathology, the route of administration, and the judgment of the prescriber. Dosage determination based on these factors is within the level of those skilled in the art, and generally, dosages range from 0.01 to 2000 mg / kg / day. A more preferable dosage is 0.1 to 500 mg / kg / day. Administration may be administered once a day, or may be divided into several times. The above dosage does not limit the scope of the present invention in any way.

The pharmaceutical composition may be administered to various mammals, such as rats, companion animals, livestock, and humans. Any mode of administration can be expected, for example, oral, rectal or intravenous, intramuscular, subcutaneous, intrauterine mucosal or intracranial injection.

The present invention also provides a health functional food for improving liver disease comprising at least one flavonoid derivative compound selected from the group consisting of compounds 1 to 13 of Formula 1 as an active ingredient.

The health functional food may include at least one flavonoid derivative compound selected from the group consisting of compounds 1 to 13 of Formula 1 and a food additive that is food-acceptable.

The health functional food includes tablets, capsules, pills or liquids, and can be added to one or more flavonoid derivative compounds selected from the group consisting of compounds 1 to 13 of Formula 1 of the present invention. Is, for example, various foods, beverages, gums, teas, vitamin complexes, health functional foods, and the like.

In addition, the present invention provides an animal feed additive for improving liver disease comprising at least one flavonoid derivative compound selected from the group consisting of compounds 1 to 13 of Formula 1 as an active ingredient.

In the animal feed additive, at least one flavonoid derivative compound selected from the group consisting of compounds 1 to 13 of Formula 1 is 0.001 to 30% by weight, preferably 0.001 to 10% by weight, most preferably 0.001 to the animal feed. It can be added at ~ 5% by weight.

The liver disease may be fatty liver.

The fatty liver is a disease in which fat synthesis in liver cells is increased due to an increase in fat synthesis or excretion in liver tissue due to excessive intake of fat or endogenous factors, and non-alcoholic fatty liver disease (NAFLD) or It may be alcoholic fatty liver disease. It is preferably a non-alcoholic fatty liver disease.

The non-alcoholic fatty liver disease refers to a state in which triglyceride is excessively accumulated in the liver, and the non-alcoholic fatty liver disease is simple steatosis, non-alcoholic fat accompanying liver necrosis, inflammation and fibrosis. Hepatitis (non-alcoholic steatohepatitis, NASH) and more advanced conditions, cirrhosis of the liver.

The alcoholic fatty liver disease is a liver disease caused by excessive alcohol. When alcohol is excessively consumed, fat synthesis is promoted in the liver and normal energy metabolism is not achieved.

The present invention relates to a composition for preventing or treating liver disease comprising a flavonoid derivative compound isolated from Sicyos angulatus as an active ingredient, wherein the flavonoid derivative compound prevents excessive fat accumulation in liver cells, and is caused by alcohol. It was confirmed to inhibit hepatocellular toxicity.

Through this, the flavonoid derivative compound isolated from the thorns of the present invention will be useful for the development of a composition for preventing or treating liver disease caused by excessive fat accumulation or alcohol, an improved health functional food, and animal feed additive. It is expected.

Figure 1 shows the results confirming the HMBC relationship for confirming the chemical structure of the new compounds, 1, 8, 9, 11 to 13, separated from thorny foil.
Figure 2 shows the results confirming the hepatotoxicity of the compounds 1 to 13 isolated from thorns.
Figure 3 shows the results confirming the change in the content of triglycerides in hepatocytes according to palmitic acid, glucose or insulin treatment for the production of a fat cell model of hepatocytes.
Figure 4 confirms the effect of preventing the accumulation of fat in hepatocytes of compounds 1 to 13 isolated from prickly pear, and 4 (A) compares the effect of preventing the accumulation of fat according to each compound by treating compounds 1 to 13 at the same concentration. As a result, 4 (B) shows the results confirming the effect of preventing fat accumulation according to the treatment concentration of Compounds 1, 3, 4, and 13.
Figure 5 shows the results obtained by confirming the effect of preventing the accumulation of fat in hepatocytes by treatment of compounds 1, 3, 4, 11, 13 among the compounds isolated from thorns.

Hereinafter, a preferred embodiment of the present invention will be described in detail. However, the present invention is not limited to the embodiments described herein and may be embodied in other forms. Rather, the contents introduced here are thorough and complete, and are provided to sufficiently convey the spirit of the present invention to those skilled in the art.

<Example 1. Separation of flavonoid derivative compounds from spiny foil>

10 kg of 70% [v / v] ethanol was added to 1 kg of prickly pear (Sicyos angulatus ) and ultrasonic extraction at room temperature was repeated three times to obtain an extract. The obtained extract was concentrated under reduced pressure to obtain 115 g of 70% [v / v] ethanol extract.

After adding 3 liters of distilled water and suspending the obtained 70% [v / v] ethanol extract, mixing with 3 liters of n-hexane and shaking for 5 minutes, the process of separating the n-hexane layer was repeated twice. Thereafter, 2 liters of ethyl acetate was mixed with the residue (water layer) from which the n-hexane layer had been removed, followed by shaking for 5 minutes and separating the ethyl acetate layer twice, and 2 l of butanol to the residue (water layer) from which the ethyl acetate layer was removed. After mixing and shaking for 5 minutes, the process of separating the butanol layer was repeated twice. The n-hexane layer, ethyl acetate layer, and butanol layer separated from each other were concentrated under reduced pressure to obtain n-hexane fraction, ethyl acetate fraction, and butanol fraction.

A portion of the ethyl acetate fraction (12.2 g) was subjected to silica gel column chromatography (silica gel) under a concentration gradient elution condition of hexane: ethyl acetate: methanol (10: 1: 0 → 0: 0: 1 [v / v / v]). After column chromatography) (column size: 7 × 50 cm, particle size: 63 to 200 μm), five fractions were obtained based on a thin-layer chromatography profile (SA.1 to SA). .5).

The SA.4 fraction was subjected to a reverse phase 18 (RP-18) column chromatography under a gradient gradient elution condition of methanol: distilled water (4: 6 → 1: 0 [v / v]) (column size: 3.5 × 30 Cm, particle size: 40 ~ 63㎛) to obtain 8 small fractions (SA.4.1 ~ SA.4.8).

Silica gel column chromatography (column size: 4 × 40 cm, particle size: 63 to 200) of the small fraction SA.4.2 in a concentration gradient elution condition of ethyl acetate: methanol (9: 1 → 0: 1 [v / v]) Μm) to obtain 6 small fractions (SA.4.2.1 to SA.4.2.6).

The SA.4.2.2 small fraction (120 mg) was subjected to high-performance liquid chromatography (HPLC) (Optima-pack ODS-H80 column 20 × 150 mm; 10 μm particle size; 2 mL / min flow rate; UV detection: 254 nm; mobile phase: MeCN in H ₂ O containing 0.1% HCO ₂ H (0-45 min: 18% MeCN) to perform compound 3 (7 mg, retention time ( t _R ) = 33.4 min), compound 4 (3 mg, t _R = 37 min), compound 5 (2 mg, t _R = 43.4 min), compound 6 (1.2 mg, t _R = 38 min), compound 8 (2.2 mg, t _R = 31.5 min) , Compound 11 (2.3 mg, t _R = 32.3 min), Compound 12 (2.2 mg, t _R = 36.3 min) and Compound 13 (1.7 mg, t _R = 38.3 min) were isolated.

In addition, the SA.4.2.3 small fraction (220 mg) was methanol: distilled water (7: 3 [v / v]) isocratic conditions, Sephadex LH-20 (sephadex LH-20) (column size: 2 × 30 Cm) Column chromatography was performed to obtain 4 fractions (SA.4.2.3.1 to SA.4.2.3.4).

The SA.4.2.3.2 fraction (150 mg) was HPLC (Optima-pack ODS-H80 column 20 × 150 mm; 10 μm particle size; 2 ml / min flow rate; UV detection: 254 nm; mobile phase: MeCN in H ₂ O containing 0.1% HCO ₂ H (0-45 min: 20% MeCN) (0-60 min)) to perform compound 1 (5 mg, t _R = 55 min), compound 2 (5 mg, t _R = 37 min ), Compound 9 (1.9 mg, t _R = 50 min) and Compound 10 (1.7 mg, t _R = 48 min) were isolated.

The SA.4.2.6 small fraction (80 mg) was HPLC (Optima-pack ODS-H80 column 20 × 150 mm; 10 μm particle size; 2 ml / min flow rate; UV detection: 254 nm; mobile phase: MeCN in H ₂ O containing 0.1% HCO ₂ H (0-45 min: 16% MeCN) (0-35 min)) was performed to isolate compound 7 (1 mg, t _R = 32.5 min).

<Example 2. Confirmation of physicochemical structure of flavonoid derivative compound>

Example 2-1. Camperol-3-O- (6-O-E-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1).

Kaempferol-3-O- (6-O-E-feruroyl) -β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside;

Yellow powder;

[ a ] _D -92.2 (c 0.5, methanol);

UV (methanol) λ _max (log ε) 201 (4.19), 265 (3.75), 328 (3.41) nm;

IR (KBr) ν _max 3299, 1656, 1588, 1291, 1181, 1085 cm ^-1 ;

HRESIMS m / z 769. 2006 [M-H] ^- (calcd for C ₃₇ H ₃₇ O ₁₈ , 769.1985);

¹ H NMR (DMSO- d ₆ , 500 MHz): δ _H 8.02 (2H, d, J = 8.0 Hz, H-2 ', 6'), 7.35 (1H, d, J = 15.5 Hz, H-7 '''), 7.20 (1H, d, J = 1.2 Hz, H-2 '''), 6.95 (1H, dd, J = 8.0, 1.2 Hz, H-6'''), 6.86 (2H, d, J = 8.0 Hz, H-3 ', 5'), 6.78 (1H, d, J = 8.0 Hz, H-5 '''), 6.73 (1H, d, J = 2.0 Hz, H-8), 6.33 (1H, d, J = 2.0 Hz, H-6), 6.25 (1H, d, J = 15.5 Hz, H-8 '''), 5.47 (1H, br s, H-1''''), 5.45 (1H, d, J = 7.5 Hz, H-1``), 4.24 (1H, dd, J = 11.5, 2.0 Hz, H-6''a), 4.12 (1H, m, H-6 '' b), 3.84 (1H, br s, H-2 ''''), 3.81 (3H, s, OMe-9 '''), 3.62 (1H, dd, J = 9.0, 2.6 Hz, H-3''''), 3.40 (1H, m, H-4''''), 3.30 (1H, m, H-5''), 3.28 (1H, m, H-3''), 3.25 (1H, m, H-2``), 3.24 (1H, m, H-5 ''''), 3.23 (1H, m, H-4 ''), 1.09 (3H, d, J = 6.0 Hz, H- 6 '''');

¹³ C NMR (DMSO- d ₆ , 125 MHz): see Table 1.

Compound 1 was isolated as a yellow powder, and the [α] _D value measured at 20 ° C. was -92.2 ( c 0.5, methanol), and the molecular formula was C ₃₇ H ₃₈ O ₁₈ determined from HRESIMS 769.2006 (calcd. 769.1985). . Compound 1 exhibited strong IR absorption at ν _max OH (3299 cm ^-1 ), C = O (1656 cm ^-1 ), suggesting the presence of α, β-unsaturated ketone groups and at λ _max 328 nm. It showed a strong UV absorption band, suggesting the presence of ferulic acid. As a result of ¹ H NMR analysis of Compound 1, the anomer proton was observed at 5.45 (d, J = 7.5 Hz), and 5.47 (br s), confirming that the compound was attached with glucose and rhamnose residues. . ¹ H NMR and ¹³ C NMR analysis results (see Table 1), A2B2 type aromatic rings (δ _H 8.02, 2H, d, J = 8.0 Hz; δ _C 130.9), (δ _H 6.86, 2H, d, J = 8.0 Hz; δ _C 115.1), trans double bond (δ _H 7.35, 1H, d, J = 15.5 Hz; δ _C 144.9), (δ _H 6.25, 1H, d, J = 15.5 Hz; δ _C 113.9), ABX Types of aromatic rings (δ _H 7.20, 1H, d, J = 1.2 Hz; δ _C 110.9), (δ _H 6.95, 1H, dd, J = 8.0, 1.2 Hz; δ _C 123.2), (δ _H 6.78, 1H , d, J = 8.0 Hz; δ _C 115.4), two carbonyl carbons (δ _C 177.6, 166.3) and two meta-coupling protons (δ _H 6.73, 1H, d, J = 2.0 Hz), (δ _H 6.33, 1H, d, J = 2.0 Hz) confirmed that compound 1 is a camphorol glycoside compound in which ferulic acid is polymerized. From the HSQC and HMBC experiments of Compound 1 (see FIG. 1), β-D-glucopyranosyl capture at the C-3 position from δ _H 5.45 (1H, d, J = 7.5 Hz) It was confirmed from the HMBC linkage to C-3 (δ _C 133.4) and the α-L-rhamnopyranosyl capture at the C-7 position was found to be C-7 from δ _H 5.47 (1H, br s). (δ _C 161.5). Also β-D- glucopyranosyl (β-D-glucopyranosyl) H -6 '(δ C' (δ H 4.24, 1H, dd, J = 11.5, 2.0 Hz) of the ferulic acid from the C-9 '''of the 166.3). Through this, it was confirmed that the novel compound 1 is Kaempferol-3-O- (6-OE-feruroyl) -β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside.

Example 2-2. Camperol-3-O- (6-O-E-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 2).

Kaempferol-3-O- (6-O-E-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside;

Yellow powder;

ESIMS m / z 739.1 [M-H] ^- ;

¹ H NMR (DMSO- d ₆ , 500 MHz): δ _H 8.02 (2H, d, J = 8.0 Hz, H-2 ', 6'), 7.38 (2H, d, J = 8.0 Hz, H-2 '',6''), 7.32 (1H, d, J = 15.5 Hz, H-7'''), 6.95 (1H, dd, J = 8.0, 1.2 Hz, H-6 '''), 6.85 (2H , d, J = 8.0 Hz, H-3 ', 5'), 6.77 (2H, d, J = 8.0 Hz, H-3 ''',5'''), 6.32 (1H, br s, H- 6), 6.12 (1H, d, J = 15.5 Hz, H-8 '''), 5.48 (1H, br s, H-1''''), 5.45 (1H, d, J = 7.5 Hz, H -1``), 4.24 (1H, d, J = 11.6 Hz, H-6''a), 4.10 (1H, dd, J = 11.6, 6.0 Hz, H-6''b), 3.84 (1H, br s, H-2`` ''), 3.62 (1H, dd, J = 9.0, 2.6 Hz, H-3 ''''), 3.40 (1H, m, H-4 ''''), 3.32 (1H, m, H-5``), 3.29 (1H, m, H-3 ''), 3.25 (1H, m, H-2 ''), 3.22 (1H, m, H-5 ''''), 3.19 (1H, m, H-4``), 1.10 (3H, d, J = 6.0 Hz, H-6'''');

¹³ C NMR (methanol- d ₄ , 125 MHz): δ _C 155.9 (C-2), 133.3 (C-3), 177.6 (C-4), 160.7 (C-5), 99.3 (C-6), 161.5 (C-7), 94.6 (C-8), 156.9 (C-9), 105.5 (C-10), 120.6 (C-1 '), 130.2 (C-2'), 115.1 (C-3 ' ), 160.2 (C-4 '), 115.1 (C-5'), 130.2 (C-6 '), 100.9 (C-1''), 74.3 (C-2''), 76.2 (C-3''), 70.0 (C-4''), 74.1 (C-5''), 62.8 (C-6''), 124.9 (C-1'''), 130.9 (C-2 '''), 115.7 (C-3 '''), 159.9 (C-4'''), 115.7 (C-5 '''), 130.9 (C-6'''), 144.6 (C-7 '''), 113.6 (C-8 '''), 166.2 (C-9'''), 98.3 (C-1 ''''), 69.8 (C-2 ''''), 70.2 (C-3 ''''), 71.6 (C-4''''), 69.7 (C-5''''), 17.9 (C-6'''').

Example 2-3. Camperol-3-O- (6-O-α-L-ramnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 3).

Kaempferol-3-O- (6-O-α-L-rhamnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside;

Yellow powder;

HRESIMS m / z 739.1 [M-H] ^- ;

¹ H NMR (DMSO- d ₆ , 500 MHz): δ _H 8.01 (2H, d, J = 8.7 Hz, H-2 ', 6'), 6.88 (2H, d, J = 8.7 Hz, H-3 ' , 5 '), 6.77 (1H, br s, H-8), 6.44 (1H, br s, H-6), 5.54 (1H, s, H-1``''), 5.35 (1H, d, J = 7.4 Hz, H-1``), 4.38 (1H, br s, H-1 '''), 3.84 (1H, br s, H-2''''), 3.67 (1H, d, J = 10.6 Hz, H-6``a), 3.64 (1H, dd, J = 9.6, 2.4, H-3 ''''), 3.48 (1H, m, H-4 ''), 3.47 (1H, m, H-3`` '), 3.46 (1H, m, H-2''), 3.45 (1H, m, H-5''''), 3.44 (1H, m, H-5''' ), 3.40 (1H, m, H-2`` '), 3.34 (1H, m, H-3''), 3.22 (1H, m, H-6''b), 3.20 (1H, m, H) -4``), 3.19 (1H, m, H-4 ''''), 3.11 (1H, m, H-4 '''), 3.10 (1H, m, H-5''), 1.11 ( 3H, d, J = 6.0 Hz, H-6 ''''), 0.97 (3H, d, J = 6.0 Hz, H-6 ''');

¹³ C NMR (DMSO- d ₆ , 125 MHz): δ _C 156.0 (C-2), 133.5 (C-3), 177.6 (C-4), 160.8 (C-5), 99.3 (C-6), 161.6 (C-7), 94.6 (C-8), 157.2 (C-9), 105.6 (C-10), 120.6 (C-1 '), 130.9 (C-2'), 115.1 (C-3 ' ), 160.2 (C-4 '), 115.1 (C-5'), 130.9 (C-6 '), 101.2 (C-1''), 74.1 (C-2''), 76.3 (C-3''), 70.3 (C-4''), 75.8 (C-5''), 66.7 (C-6''), 100.7 (C-1'''), 70.2 (C-2 '''), 70.5 (C-3 '''), 71.8 (C-4'''), 68.2 (C-5 '''), 17.7 (C-6'''), 98.3 (C-1 '''') , 69.8 (C-2`` ''), 70.0 (C-3 ''''), 71.6 (C-4 ''''), 69.8 (C-5 ''''), 17.9 (C-6 '''').

Example 2-4. Camperol-3-O-β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 4).

Kaempferol-3-O-β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside;

Yellow powder;

ESIMS m / z 593.1 [M-H] ^- ;

¹ H NMR (DMSO- d ₆ , 400 MHz): δ _H 8.07 (2H, d, J = 8.8 Hz, H-2 ', 6'), 6.88 (2H, d, J = 8.8 Hz, H-3 ' , 5 '), 6.82 (1H, br s, H-8), 6.44 (1H, br s, H-6), 5.54 (1H, s, H-1'''), 5.47 (1H, d, J = 7.3 Hz, H-1``), 3.83 (1H, br s, H-2 '''), 3.64 (1H, dd, J = 9.3, 2.4 Hz, H-3'''), 3.53 (1H , d, J = 11.6 Hz, H-6``a), 3.48 (1H, m, H-5 ''), 3.45 (1H, m, H-5 '''), 3.34 (1H, m, H -3``), 3.32 (1H, m, H-6''b), 3.27 (1H, m, H-2 ''), 3.46 (1H, m, H-3 ''), 3.19 (1H, m, H-4 ''''), 1.11 (3H, d, J = 6.0 Hz, H-6 '''');

¹³ C NMR (DMSO- d ₆ , 100 MHz): δ _C 156.0 (C-2), 133.4 (C-3), 177.5 (C-4), 160.9 (C-5), 99.4 (C-6), 161.5 (C-7), 94.9 (C-8), 156.6 (C-9), 105.6 (C-10), 120.6 (C-1 '), 130.9 (C-2'), 115.1 (C-3 ' ), 160.2 (C-4 '), 115.1 (C-5'), 130.9 (C-6 '), 100.6 (C-1''), 74.1 (C-2''), 76.4 (C-3''), 70.2 (C-4''), 74.5 (C-5''), 60.8 (C-6''), 98.3 (C-1'''), 69.9 (C-2 '''), 70.5 (C-3 '''), 71.5 (C-4'''), 69.8 (C-5 '''), 17.9 (C-6''').

Example 2-5. Apigenin-7-O-β-D-glucopyranoside (Compound 5).

Apigenin-7-O-β-D-glucopyranoside;

Yellow powder;

ESIMS m / z 433.1 [M + H] ⁺ ;

¹ H NMR (DMSO- d ₆ , 600 MHz): δ _H 7.95 (2H, d, J = 8.0 Hz, H-2 ', 6'), 6.94 (2H, d, J = 8.0 Hz, H-3 ' , 5 '), 6.87 (1H, s, H-3), 6.82 (1H, br s, H-8), 6.44 (1H, br s, H-6), 5.06 (1H, d, J = 7.3 Hz , H-1 ''), 3.70 (1H, d, J = 11.3 Hz, H-6''a), 3.48 (1H, dd, J = 11.0, 5.6 Hz, H-6''b), 3.46 ( 1H, m, H-5 ''), 3.31 (1H, m, H-3 ''), 3.28 (1H, m, H-2 ''), 3.19 (1H, m, H-4 '');

¹³ C NMR (DMSO- d ₆ , 150 MHz): δ _C 164.2 (C-2), 103.0 (C-3), 182.0 (C-4), 161.4 (C-5), 99.5 (C-6), 162.9 (C-7), 94.8 (C-8), 156.9 (C-9), 105.3 (C-10), 120.9 (C-1 '), 128.6 (C-2'), 116.0 (C-3 ' ), 161.1 (C-4 '), 116.0 (C-5'), 128.6 (C-6 '), 99.8 (C-1''), 73.0 (C-2''), 76.4 (C-3''), 69.5 (C-4''), 77.1 (C-5''), 60.5 (C-6'').

Example 2-6. Camperol-3-O-β-D-glucopyranoside (Compound 6).

Kaempferol-3-O-β-D-glucopyranoside;

Yellow powder;

ESIMS m / z 447.1 [M-H] ^- ;

¹ H NMR (DMSO- d ₆ , 600 MHz): δ _H 8.10 (2H, d, J = 8.6 Hz, H-2 ', 6'), 6.90 (2H, d, J = 8.6 Hz, H-3 ' , 5 '), 6.90 (1H, d, J = 1.6 Hz, H-8), 6.20 (1H, d, J = 1.6 Hz, H-6), 5.40 (1H, d, J = 7.6 Hz, H- 1 ''), 3.56 (1H, d, J = 11.0 Hz, H-6''a), 3.34 (1H, m, H-3 ''), 3.23 (1H, m, H-4 ''), 3.22 (1H, m, H-6''b), 3.15 (1H, m, H-5 ''), 3.11 (1H, m, H-2 '');

¹³ C NMR (DMSO- d ₆ , 150 MHz): δ _C 156.0 (C-2), 133.4 (C-3), 177.5 (C-4), 160.9 (C-5), 99.4 (C-6), 161.5 (C-7), 94.9 (C-8), 156.6 (C-9), 105.6 (C-10), 120.6 (C-1 '), 130.9 (C-2'), 115.1 (C-3 ' ), 160.2 (C-4 '), 115.1 (C-5'), 130.9 (C-6 '), 100.6 (C-1''), 74.1 (C-2''), 76.4 (C-3''), 70.2 (C-4''), 74.5 (C-5''), 60.8 (C-6''), 98.3 (C-1'''), 69.9 (C-2 '''), 70.5 (C-3 '''), 71.5 (C-4'''), 69.8 (C-5 '''), 17.9 (C-6''').

Example 2-7. Camperol-3-O- (6-O-α-L-ramnopyranosyl) -β-D-glucopyranoside (Compound 7).

Kaempferol-3-O- (6-O-α-L-rhamnopyranosyl) -β-D-glucopyranoside;

Yellow powder;

ESIMS m / z 594.1 [M-H] ^- ;

¹ H NMR (DMSO- d ₆ , 600 MHz): δ _H 7.97 (2H, d, J = 8.8 Hz, H-2 ', 6'), 6.87 (2H, d, J = 8.8 Hz, H-3 ' , 5 '), 6.34 (1H, br s, H-8), 6.14 (1H, br s, H-6), 5.28 (1H, d, J = 7.7 Hz, H-1``), 4.37 (1H , s, H-1 '''), 3.64 (1H, d, J = 11.0 Hz, H-6''a), 3.40 (1H, br s, H-2'''), 3.24 (1H, m , H-3`` '), 3.48 (1H, m, H-5''), 3.45 (1H, m, H-5'''), 3.34 (1H, m, H-3 ''), 3.32 (1H, m, H-6``b), 3.27 (1H, m, H-2 ''), 3.46 (1H, m, H-3 ''), 3.19 (1H, m, H-4 ''''), 0.98 (3H, d, J = 6.0 Hz, H-6 '''');

¹³ C NMR (DMSO- d ₆ , 100 MHz): δ _C 156.9 (C-2), 133.2 (C-3), 177.4 (C-4), 161.2 (C-5), 98.8 (C-6), 164.2 (C-7), 93.8 (C-8), 159.9 (C-9), 103.9 (C-10), 120.9 (C-1 '), 130.9 (C-2'), 115.1 (C-3 ' ), 159.9 (C-4 '), 115.1 (C-5'), 130.9 (C-6 '), 101.3 (C-1''), 74.2 (C-2''), 76.3 (C-3''), 69.9 (C-4''), 75.7 (C-5''), 66.9 (C-6''), 100.8 (C-1'''), 70.3 (C-2 '''), 70.6 (C-3 '''), 71.8 (C-4'''), 68.2 (C-5 '''), 17.7 (C-6''').

Example 2-8. Camperol-3-O-β-D-apiopuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-lamnopyranoside (Compound 8).

Kaempferol-3-O-β-D-apiofuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-rhamnopyranoside;

Red sword;

[ a ] _D- 60.3 (c 0.5, methanol);

UV (methanol) λ _max (log ε) 201 (4.40), 271 (3.80), 358 (3.30) nm;

IR (KBr) ν _max 3345, 1657, 1603, 1348, 1175, 1077 cm ^-1 ;

HRESIMS m / z 725.1961 [M-H] ^- (calcd for C ₃₂ H ₃₇ O ₁₉ , 725.1935);

¹ H NMR (DMSO- d ₆ , 800 MHz): δ _H 7.98 (2H, d, J = 8.0 Hz, H-2 ', 6'), 6.91 (2H, d, J = 8.0 Hz, H-3 ' , 5 '), 6.86 (1H, d, J = 2.0 Hz, H-8), 6.44 (1H, d, J = 2.0 Hz, H-6), 5.74 (1H, d, J = 2.1 Hz, H- 1 ''''), 5.59 (1H, br s, H-1 '''), 4.48 (1H, d, J = 7.7 Hz, H-1'''), 4.19 (1H, d, J = 2.4 Hz, H-2``), 4.10 (1H, br s, H-2 '''), 3.70 (1H, dd, J = 8.7, 3.2 Hz, H-3'''), 3.68 (1H, dd , J = 11.7, 1.9 Hz, H-6 '''' a), 3.60 (1H, d, J = 9.3 Hz, H-4''a), 3.52 (1H, m, H-4 ''') , 3.51 (1H, m, H-5`` '), 3.49 (1H, d, J = 9.3 Hz, H-4''b), 3.43 (1H, dd, J = 11.7, 5.7 Hz, H-6 '''' b), 3.31 (1H, d, J = 11.0 Hz, H-5''a), 3.30 (1H, d, J = 11.0 Hz, H-5''b), 3.18 (1H, m , H-3 ''''), 3.16 (1H, m, H-5 ''''), 3.09 (1H, m, H-2 ''''), 3.09 (1H, m, H-4 ''''), 1.14 (3H, d, J = 6.0 Hz, H-6''');

¹³ C NMR (DMSO- d ₆ , 200 MHz): see Table 1.

Compound 8 was isolated as a reddish gum, and the [α] _D value measured at 20 ° C. was -60.3 ( c 0.5, methanol), and the molecular formula was C ₃₂ H ₃₈ O ₁₉ with HRESIMS 725.1961 (calcd. 725.1935). As a result of ¹ H NMR analysis of compound 8, the anomeric proton was observed at 5.74 (d, J = 2.1 Hz), 5.59 (br s), and 4.48 (d, J = 7.7 Hz), and apiofuranos, rhamnose, and glucose residue It was confirmed that the compound is attached. ¹ H NMR and ¹³ C NMR analysis results (see Table 1), A2B2 type aromatic rings (δ _H 7.98, 2H, d, J = 8.0 Hz; δ _C 130.7), (δ _H 6.91, 2H, d, J = 8.0 Hz; δ _C 115.5), one carbonyl carbon (δ _C 177.0) and two meta-coupled protons (δ _H 6.86, 1H, d, J = 2.0 Hz), (δ _H 6.48, 1H, d, J = 2.0 Hz), it was confirmed that compound 8 is a camphorol glycoside compound. From the HSQC and HMBC experiments of compound 8 (see FIG. 1), β-D-apiofuranosyl capture at the C-3 position was δ _H 5.74 (1H, d, J = 2.1 Hz). From the HMBC connection to C-3 (δ _C 133.7) and the α-L-rhamnopyranosyl capture at the C-7 position was found to be C- from δ _H 5.59 (1H, br s). 7 (δ _C 161.4). Also, β-D-glucopyranosyl (β-D) from α-L-rhamnopyranosyl H-3 ′ '' (δ _H 3.70, 1H, dd, J = 8.7, 3.2 Hz) -glucopyranosyl) to C-1 '''' (δ _C 104.6) was confirmed HMBC linkage. Through this, it was confirmed that the new compound 8 is Kaempferol-3-O-β-D-apiofuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-rhamnopyranoside.

Example 2-9. Quercetin-3-O- (6-O-E-feruloyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 9).

Quercetin-3-O- (6-O-E-feruroyl) -β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside;

Yellow powder;

[ a ] _D -115.0 (c 0.5, methanol);

UV (methanol) λ _max (log ε) 201 (4.12), 265 (3.71), 326 (3.40) nm;

IR (KBr) ν _max 3304, 1656, 1598, 1446, 1286, 1181, 1085 cm ^-1 ;

HRESIMS m / z 785.1964 [M-H] ^- (calcd for C ₃₇ H ₃₇ O ₁₉ , 785.1935);

¹ H NMR (DMSO- d ₆ , 800 MHz): δ _H 7.58 (1H, d, J = 2.1 Hz, H-2 '), 7.56 (1H, dd, J = 8.4, 2.1 Hz, H-6') , 7.34 (1H, d, J = 15.8 Hz, H-7 '''), 7.20 (1H, d, J = 1.8 Hz, H-2'''), 6.94 (1H, dd, J = 8.2, 1.8 Hz, H-6`` '), 6.93 (1H, d, J = 8.4 Hz, H-5'), 6.77 (1H, d, J = 8.1 Hz, H-5 '''), 6.69 (1H, d, J = 2.0 Hz, H-8), 6.30 (1H, d, J = 2.1 Hz, H-6), 6.24 (1H, d, J = 15.8 Hz, H-8 '''), 5.49 (1H , d, J = 7.6 Hz, H-1``), 5.46 (1H, br s, H-1 ''''), 4.24 (1H, d, J = 10.1 Hz, H-6''a), 4.13 (1H, dd, J = 11.9, 6.2 Hz, H-6''b), 3.84 (1H, br s, H-2 ''''), 3.81 (3H, s, OMe-9 ''') , 3.61 (1H, dd, J = 8.2, 3.2 Hz, H-3 ''''), 3.39 (1H, m, H-2 ''), 3.30 (1H, m, H-5 ''), 3.28 (1H, m, H-4 ''''), 3.27 (1H, m, H-3 ''), 3.23 (1H, m, H-5 ''''), 1.09 (3H, d, J = 6.2 Hz, H-6 '''');

¹³ C NMR (DMSO- d ₆ , 200 MHz): see Table 1.

Compound 9 was isolated as a yellow powder, and the [α] _D value measured at 20 ° C. was -115.0 ( c 0.5, methanol), and the molecular formula was determined from HRESIMS 785.1964 (calcd. 785.1935) with C ₃₇ H ₃₈ O ₁₉ . . Compound 9 has a structure in which a hydroxyl group is added to the non-glycoside B ring of compound 1, and the molecular weight is increased to 16. Analysis results of ¹ H NMR and ¹³ C NMR of compound 9 (see Table 1), ABX type aromatic ring of B ring (δ _H 7.58, 1H, d, J = 2.1 Hz; δ _C 116.3), (δ _H 6.93, 1H, d, J = 8.4 Hz; δ _C 115.2), (δ _H 7.56, 1H, dd, J = 8.4, 2.1 Hz; δ _C 121.7). Coupling constants of anomer protons confirmed the connection to the β and α positions of the glucopyranosyl (J = 7.6 Hz) and rhamsnopyranosyl (br s) groups, respectively. From the HSQC and HMBC experiment results of compound 9 (see Fig. 1), the linking order of sugars was confirmed, and thus new compound 9 was Quercetin-3-O- (6-OE-feruroyl) -β-D-glucopyranosyl-7-O- It was confirmed that α-L-rhamnopyranoside.

Example 2-10. Quercetin-3-O- (6-O-E-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 10).

Quercetin-3-O- (6-O-E-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside;

Yellow powder;

ESIMS m / z 755.1 [M-H] ^- ;

¹ H NMR (DMSO- d ₆ , 600 MHz): δ _H 7.59 (1H, d, J = 2.0 Hz, H-2 '), 7.56 (1H, dd, J = 8.4, 2.1 Hz, H-6') , 7.38 (2H, d, J = 8.6 Hz, H-2 ''',6'''), 6.83 (1H, d, J = 8.4 Hz, H-5 '), 6.77 (2H, d, J = 8.5 Hz, H-3`` ', 5'''), 6.69 (1H, d, J = 2.0 Hz, H-8), 6.30 (1H, d, J = 2.0 Hz, H-6), 5.49 ( 1H, d, J = 7.6 Hz, H-1``), 5.48 (1H, br s, H-1 ''''), 4.23 (1H, d, J = 10.3 Hz, H-6''a) , 4.11 (1H, dd, J = 11.0, 6.2 Hz, H-6``b), 3.84 (1H, br s, H-2 ''''), 3.61 (1H, m, H-3 ''''), 3.40 (1H, m, H-2``), 3.38 (1H, m, H-4''), 3.30 (1H, m, H-5''), 3.28 (1H, m, H- 3 ''), 3.28 (1H, m, H-4 ''''), 3.28 (1H, m, H-3 ''), 3.22 (1H, m, H-5 ''''), 1.10 ( 3H, d, J = 6.1 Hz, H-6 '''');

¹³ C NMR (DMSO- d ₆ , 150 MHz): δ _C 156.7 (C-2), 133.3 (C-3), 177.5 (C-4), 160.7 (C-5), 99.2 (C-6), 161.5 (C-7), 94.3 (C-8), 155.8 (C-9), 105.4 (C-10), 120.8 (C-1 '), 116.2 (C-2'), 144.5 (C-3 ' ), 148.7 (C-4 '), 115.1 (C-5'), 121.6 (C-6 '), 100.7 (C-1''), 74.3 (C-2''), 76.2 (C-3''), 70.0 (C-4''), 73.9 (C-5''), 62.8 (C-6''), 124.9 (C-1'''), 130.1 (C-2 '''), 115.6 (C-3 '''), 159.7 (C-4'''), 115.6 (C-5 '''), 130.1 (C-6'''), 144.8 (C-7 '''), 113.6 (C-8 '''), 166.2 (C-9'''), 98.3 (C-1 ''''), 70.0 (C-2 ''''), 70.2 (C-3 ''''), 71.5 (C-4''''), 69.8 (C-5''''), 17.8 (C-6'''').

Example 2-11. Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranosyl- (1-6) -β-D-glucopyranosyl -(1-3) -α-L-ramnopyranoside (Compound 11).

Kaempferol-3-O- (6-OE-feruroyl) -β-D-glucopyranosyl-7-O-α-L-rhamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α- L-rhamnopyranoside;

Yellow sword;

[ a ] _D -78.5 (c 0.5, methanol);

UV (methanol) λ _max (log ε) 201 (3.76), 265 (3.50), 326 (3.40) nm;

IR (KBr) ν _max 3304, 1656, 1598, 1446, 1286, 1181, 1085 cm ^-1 ;

HRESIMS m / z 1077.3108 [M-H] ^- (calcd for C ₄₉ H ₅₇ O ₂₇ , 1077.3093);

¹ H NMR (DMSO- d ₆ , 800 MHz): δ _H 8.14 (2H, d, J = 8.8 Hz, H-2 ', 6'), 7.49 (1H, d, J = 15.8 Hz, H-7 '''), 7.10 (1H, d, J = 1.8 Hz, H-2 '''), 6.92 (1H, dd, J = 8.1, 1.8 Hz, H-6'''), 6.87 (2H, d, J = 8.0 Hz, H-3 ', 5'), 6.77 (1H, d, J = 8.1 Hz, H-5 '''), 6.59 (1H, d, J = 2.0 Hz, H-8), 6.32 (1H, d, J = 2.0 Hz, H-6), 6.39 (1H, d, J = 15.8 Hz, H-8 '''), 5.44 (1H, d, J = 1.2 Hz, H-1''''), 5.33 (1H, d, J = 7.6 Hz, H-1 ''), 4.48 (1H, d, J = 7.7 Hz, H-1 '''''), 4.41 (1H, dd, J = 11.8, 1.8 Hz, H-6``a), 4.38 (1H, d, J = 1.2 Hz, H-1 ''''''), 4.23 (1H, dd, J = 11.9, 7.0 Hz, H -6``b), 4.08 (1H, br s, H-2 ''''), 3.72 (1H, dd, J = 9.0, 2.5 Hz, H-3 ''''), 3.71 (1H, d , J = 10.0 Hz, H-6 '''''a), 3.68 (3H, s, OMe-9'''), 3.51 (1H, m, H-4 ''''), 3.50 (1H, m, H-5``), 3.45 (1H, br s, H-5 ''''), 3.41 (1H, m, H-3 ''''''), 3.26 (1H, m, H- 2 ''), 3.26 (1H, br s, H-4 '''''), 3.25 (1H, m, H-6''''' b), 3.25 (1H, br s, H-2 ''''''), 3.23 (1H, m, H-3''), 3.17 (1H, m, H-5'''''), 3.15 (1H, m, H-4 '') 3.14 ( 1H, m, H-2`` '''), 3.09 (1H, m, H -5`` ''''), 3.06 (1H, m, H-4 ''''''), 1.10 (3H, d, J = 6.0 Hz, H-6 ''''), 0.97 (3H , d, J = 6.0 Hz, H-6 '''''');

¹³ C NMR (DMSO- d ₆ , 200 MHz): see Table 1.

Compound 11 was isolated as a yellow gum, and the [α] _D value measured at 20 ° C. was -78.5 ( c 0.5, methanol), and the molecular formula was determined from HRESIMS 1077.3108 (calcd. 1077.3093) with C ₄₉ H ₅₈ O ₂₇ . . Compound 11 is a structure in which β-D-glucopyranosyl and α-L-lamnopyranosyl are added to Compound 1. Β-D-glucopyranosyl is connected to H-3 '''' of α-L-lamnopyranosyl linked to C-7 of A-ring, H-6 '' of β-D-glucopyranosyl The α-L-lamnopyranosyl capture is additionally linked to '''fromH-3''''(δ _H 3.72) to C-1''''' (δ _C 105.0), H-6 ', respectively. It was confirmed by HMBC connection from '''' (δ _H 3.71) to C-1 '''''' (δ _C 100.6). The coupling constant of the anomer proton of compound 11 is linked to the β and α positions of the glucopyranosyl ( J = 7.6 Hz) ( J = 7.7 Hz) and rhamnopyranosyl ( J = 1.2 Hz) groups, respectively. Was confirmed. Therefore, the new compound 11 is Kaempferol-3-O- (6-OE-feruroyl) -β-D-glucopyranosyl-7-O-α-L-rhampyranosyl- (1-6) -β-D-glucopyranosyl- (1- It was confirmed that 3) -α-L-rhamnopyranoside.

Example 2-12. Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-lamnopyrano Seed (Compound 12).

Kaempferol-3-O- (6-O-E-feruroyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-rhamnopyranoside;

Yellow powder;

[ a ] _D -64.9 (c 0.5, methanol);

UV (methanol) λ _max (log ε) 311 (4.10) nm;

IR (KBr) ν _max 342, 2910, 1702, 1645, 1643, 1142, 606 cm ^-1 ;

HRESIMS m / z 931.2523 [M-H] ^- (calcd for C ₄₃ H ₄₇ O ₂₃ , 931.2514);

¹ H NMR (DMSO- d ₆ , 800 MHz): δ _H 8.09 (2H, d, J = 8.9 Hz, H-2 ', 6'), 7.35 (1H, d, J = 15.8 Hz, H-7 '''), 7.18 (1H, d, J = 1.8 Hz, H-2 '''), 6.95 (1H, dd, J = 8.2, 1.8 Hz, H-6'''), 6.86 (2H, d, J = 8.9 Hz, H-3 ', 5'), 6.77 (1H, d, J = 8.1 Hz, H-5 '''), 6.76 (1H, d, J = 2.0 Hz, H-8), 6.32 (1H, d, J = 2.0 Hz, H-6), 6.24 (1H, d, J = 15.8 Hz, H-8 '''), 5.48 (1H, br s, H-1''''), 5.41 (1H, d, J = 7.6 Hz, H-1 ''), 4.48 (1H, d, J = 7.6 Hz, H-1 '''''), 4.18 (1H, dd, J = 11.2, 3.7 Hz, H-6``a), 4.11 (1H, br s, H-2 ''''), 4.07 (1H, dd, J = 11.3, 7.5 Hz, H-6''b), 3.79 (3H , s, OMe-9 '''), 3.75 (1H, dd, J = 9.1, 3.1 Hz, H-3''''), 3.70 (1H, d, J = 11.0 Hz, H-6''''' a), 3.52 (1H, m, H-4 ''''), 3.51 (1H, m, H-5 ''''), 3.45 (1H, m, H-6 '''''b ), 3.30 (1H, br s, H-5``), 3.28 (1H, m, H-3 ''), 3.26 (1H, m, H-2 ''), 3.23 (1H, br s, H -4``), 3.18 (1H, br s, H-3 '''''), 3.16 (1H, m, H-5'''''), 3.09 (1H, m, H-2 '''''), 3.09 (1H, m, H-4'''''), 1.11 (3H, d, J = 6.1 Hz, H-6 '''');

¹³ C NMR (DMSO- d ₆ , 200 MHz): see Table 1.

Compound 12 was isolated as a yellow powder, and the [α] _D value measured at 20 ° C. was -64.9 ( c 0.5, methanol). Compound 12 is a structure in which one of rhamnopyranosyl of compound 11 is removed, the molecular weight of HRESIMS is reduced by 146, and the molecular formula is determined to be C ₄₃ H ₄₈ O ₂₃ . Coupling constants of anomer protons confirmed the connection of glucopyranosyl ( J = 7.6 Hz) and rhamnopyranosyl (br s) groups to β and α positions, respectively. From the HSQC and HMBC experiment results of compound 12 (see FIG. 1), the linking order of sugars was confirmed, and thus, new compound 12 was Kaempferol-3-O- (6-OE-feruroyl) -β-D-glucopyranosyl-7-O- It was confirmed that β-D-glucopyranosyl- (1-3) -α-L-rhamnopyranoside.

Example 2-13. Trisine 4′-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α-L-ramnopyranosyl- (1-4) -β-D-glucopyranoside ( Compound 13).

Tricin 4′-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α-L-rhamnopyranosyl- (1-4) -β-D-glucopyranoside;

Red colored amorphous powder;

[ a ] _D -39.8 (c 0.5, methanol);

IR (KBr) ν _max 3410, 1658, 1613, 1495, 1459, 1352, 1256, 1133 cm ^-1 ;

HRESIMS m / z 833.2513 [M-H] ^- (calcd for C ₃₉ H ₄₅ O ₂₀ , 833.2510);

¹ H NMR (DMSO- d ₆ , 800 MHz): δ _H 7.32 (2H, s, H-2 ', 6'), 6.93 (1H, br s, H-2 ''), 6.90 (1H, d, J = 2.0 Hz, H-8), 6.39 (1H, d, J = 2.0 Hz, H-6), 6.75 (1H, dd, J = 8.0, 1.6 Hz, H-6 ''), 6.69 (1H, d, J = 8.1 Hz, H-5``), 5.21 (1H, d, J = 7.7 Hz, H-1 '''), 5.12 (1H, br s, H-1''''), 4.79 (1H, br s, H-7``), 4.37 (1H, m, H-8 ''), 3.87 (6H, s, OMe-3 ', 5'), 3.74 (3H, s, OMe-4 ''), 3.70 (1H, m, H-9''a), 3.49 (1H, m, H-9''b), 3.69 (1H, br s, H-2 ''''), 3.77 ( 1H, br s, H-5`` ''), 3.51 (1H, m, H-4 '''), 3.48 (1H, m, H-2'''), 3.48 (1H, m, H- 3 '''), 3.47 (1H, m, H-6''' a), 3.46 (1H, m, H-6 '''b), 3.34 (1H, m, H-3'''') , 3.21 (1H, m, H-4`` ''), 3.18 (1H, m, H-5 '''), 1.18 (3H, d, J = 6.0 Hz, H-6'''');

¹³ C NMR (DMSO- d ₆ , 200 MHz): see Table 1.

Compound 13 was isolated as a red amorphous powder, and the [α] _D value measured at 20 ° C. was -39.8 ( c 0.5, methanol), and the molecular formula was determined to be C ₃₉ H ₄₆ O ₂₀ calculated from HRESIMS measurements. As a result of ¹ H NMR analysis of Compound 13, the anomer proton was confirmed at 5.21 (1H, d, J = 7.7 Hz) and 5.12 (1H, br s) to confirm that the compound was attached with glucose and rhamnose residues. ¹ H NMR, ¹³ C NMR analysis results (see Table 1), A2 type aromatic rings (δ _H 7.32, 2H, s, δ _C 104.3) and ABX type aromatic rings (δ _H 6.93, br s; δ _C 104.3 ), (δ _H 6.75, 1H, dd, J = 8.0, 1.6 Hz; δ _C 119.7), (δ _H 6.69, 1H, d, J = 8.1 Hz; δ _C 114.7), three methoxy groups (δ _H 3.87 , 6H, s) (δ _H 3.74, 3H, s) and two methoxy groups (δ _H 3.87, 6H, s) are present in A2 type aromatic rings C-3 ', 5' (δ _C 152.9) Ham was confirmed by HMBC connection. In addition, by confirming the HMBC linkage from H-8 (δ _H 4.37, 1H, m) of the guiaacylglycerol capture to C-4 '(δ _C 139.7) of the A2 type aromatic ring, compound 13 polymerizes guiaacylglycerol. It was confirmed that it was a trisine glycoside compound. From the HSQC and HMBC experiments of compound 13 (see FIG. 1), the β-D-glucopyranosyl capture at the C-7 position was shown by C-7 (δ _C 162.8) from δ _H 5.21 (1H, d, J = 7.7 Hz). ) Was confirmed from the HMBC linkage to) and the α-L-lamnopyranosyl capture of the C-4 '''position of glucose from δ _H 5.12 (1H, br s) to C-4''' (δ _C 76.4) Confirmed from HMBC connection. In addition, the relative arrangement was confirmed through the ROESY connection of two oxymesain protons (δ _H 4.37, 4.79) of the guaiacylglycerol capture, and the absolute arrangement of the protons was compared to the experimental values and calculated values of circular polarization dichroism β position. Decided. The time dependent density functional theory (TDDFT) function was used, and the Boltzmann distribution of the lowest five heteromorphic forms with the lowest energy was 99%. Through this, it was confirmed that the novel compound 13 is Tricin 4'-O- (7''R, 8''R-guaiacylglyceryl) ether 7-O-α-L-rhamnopyranosyl- (1-4) -β-D-glucopyranoside .

No. Compound 1a Compound 8b Compound 9b Compound 11b Compound 12b No. Compound 13b 2 156.9, C 157.3, C 156.8, C 155.8, C 155.8, C 2 163.9, C 3 133.4, C 133.7, C 133.4, C 133.6, C 133.5, C 3 105.1, CH 4 177.6, C 177.0, C 177.5, C 177.5, C 177.6, C 4 182.2, C 5 160.8, C 160.9, C 160.8, C 160.7, C 160.7, C 5 161.0, C 6 99.2, CH 99.5, CH 99.2, CH 99.1, CH 99.2, CH 6 99.6, CH 7 161.5, C 161.4, C 161.5, C 161.1, C 161.2, C 7 162.8, C 8 94.6, CH 94.8, CH 94.4, CH 94.6, CH 94.8, CH 8 95.0, CH 9 155.9, C 155.9, C 155.8, C 157.3, C 156.9, C 9 157.0, C 10 105.5, C 105.7, C 105.5, C 105.5, C 105.5, C 10 105.6, C One' 120.8, C 120.5, C 120.8, C 120.8, C 120.6, C One' 124.9, C 2' 130.9, CH 130.7, CH 116.3, CH 131.0, CH 131.1, CH 2' 104.3, CH 3 ' 115.1, CH 115.5, CH 144.5, C 115.0, CH 115.0, CH 3 ' 152.9, C 4' 160.2, C 160.1, C 148.8, C 160.0, C 160.1, C 4' 139.7, C 5 ' 115.1, CH 115.5, CH 115.2, CH 115.0, CH 115.0, CH 5 ' 152.9, C 6 ' 130.9, CH 130.7, CH 121.7, CH 131.0, CH 131.1, CH 6 ' 104.3, CH One'' 101.0, CH 109.2, CH 100.8, CH 101.5, CH 101.6, CH OMe 56.3, CH ₃ 2'' 74.3, CH 77.1, CH 74.4, CH 75.6, CH 74.0, CH One'' 133.2, C 3 '' 76.2, CH 79.4, C 76.3, CH 76.0, CH 76.2, CH 2'' 110.9, CH 4'' 70.1, CH 75.1, CH ₂ 70.0, CH 70.2, CH 69.8, CH 3 '' 146.9, C 5 '' 74.1, CH 62.7, CH ₂ 74.0, CH 74.1, CH 74.1, CH 4'' 145.4, C 6 '' 62.8, CH ₂ 62.9, CH ₂ 63.4, CH ₂ 62.9, CH ₂ 5 '' 114.7, CH One''' 125.5, C 98.2, CH 125.5, C 125.3, C 125.5, C 6 '' 119.7, CH 2''' 110.9, CH 68.9, CH 110.9, CH 110.5, CH 110.9, CH 7 '' 72.1, CH 3 '' ' 147.8, C 80.7, CH 147.8, C 147.6, C 147.8, C 8'' 86.5, CH 4''' 149.3, C 70.5, CH 149.3, C 149.2, C 149.3, C 9 '' 60.3, CH ₂ 5 '' ' 115.4, CH 69.6, CH 115.4, CH 115.1, CH 115.4, CH OMe 55.5, CH ₃ 6 '' ' 123.2, CH 17.8, CH ₃ 123.1, CH 123.0, CH 123.1, CH One''' 98.2, CH 7 '' ' 144.9, CH 144.9, CH 145.2, CH 145.0, CH 2''' 77.3, CH 8''' 113.9, CH 113.9, CH 113.9, CH 113.9, CH 3 '' ' 77.2, CH 9 '' ' 166.3, C 166.3, C 166.6, C 166.2, C 4''' 76.4, CH OMe 55.6, CH ₃ 55.6, CH ₃ 55.4, CH ₃ 55.6, CH ₃ 5 '' ' 69.8, CH One'''' 98.3, CH 104.6, CH 98.4, CH 97.9, CH 98.1, CH 6 '' ' 60.5, CH ₂ 2'''' 69.8, CH 73.9, CH 69.8, CH 69.2, CH 68.2, CH One'''' 100.5, CH 3 '' '' 70.2, CH 76.2, CH 70.2, CH 81.2, CH 80.7, CH 2'''' 70.4, CH 4'''' 71.6, CH 69.9, CH 71.6, CH 70.1, CH 70.5, CH 3 '' '' 70.5, CH 5 '' '' 69.8, CH 76.7, CH 69.8, CH 69.6, CH 68.9, CH 4'''' 71.8, CH 6 '' 17.8, CH ₃ 60.9, CH ₂ 17.8, CH ₃ 17.8, CH ₃ 17.8, CH ₃ 5 '' '' 68.4, CH One''''' 105.0, CH 104.6, CH 6 '' 18.0, CH ₃ 2''''' 73.8, CH 72.9, CH 3 '' '' ' 76.4, CH 76.2, CH 4''''' 70.5, CH 69.5, CH 5 '' '' ' 74.2, CH 76.7, CH 6 '' '' ' 66.7, CH ₂ 60.9, CH ₂ a: Recorded in DMSO-d ₆ at 125 MHz.
b: Recorded in DMSO-d ₆ at 200 MHz.

<Example 3. Cytotoxicity evaluation for hepatocytes>

MTT (3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide using hepG2 cells, a hepatocyte cell line, to confirm the cytotoxicity of the flavonoid derivative compound isolated from prickly pear in Example 1 ) Assay was performed.

HepG2 cells use DMEM (Dulbecco's modified Eagle's medium) medium containing 10% fetal bovine serum (FBS), 1% PS (penicillin + streptomycin) and L-glutamine (2 ml based on 500 ml medium). And cultured in a cell incubator at 37 ° C and 5% CO ₂ conditions. The cultured HepG2 cells were placed in a 96-well plate to be 5 × 10 ³ cells per well, and the HepG2 cells were cultured to fill about 75% of the plate area. Compounds 1 to 13 isolated in Example 1 were each treated in wells to have a concentration of 100 uM and reacted for 24 hours. At this time, HepG2 cells, which were not treated with anything, were used as normal controls. After 24 hours, 2 μl / ml MTT solution was treated with 20 μl for each well, and after incubation for 4 hours in a cell incubator at 37 ° C. and 5% CO ₂ conditions, the culture solution was removed and DMSO (dimethylsulfoxide) was added to each well. After treating 100 µl each, the cell viability was analyzed by measuring the absorbance at 550 nm, and the results are shown in FIG. 2. At this time, the cell viability of the normal control was based on 100%, and the cell viability by treatment with each compound was calculated.

As shown in Figure 2, it was confirmed that the compounds 1 to 13 isolated from thorns are not cytotoxic to hepatocytes and do not affect hepatocyte growth.

<Example 4. Confirmation of the effect of preventing fat accumulation in hepatocytes>

Example 4-1. Fat accumulation-inducing cell model preparation

Fatty liver is a disease caused by fat accumulation in liver cells due to increased fat synthesis or discharge in liver tissue due to excessive consumption of fat or endogenous factors, and can be divided into non-alcoholic fatty liver disease and alcoholic fatty liver disease. Alcoholic fatty liver disease is caused by excessive alcohol intake, which reduces the efficiency of alcohol metabolism, resulting in increased fatty liver. Non-alcoholic fatty liver disease occurs even without a history of alcohol intake, and excessive accumulation of triglycerides in the liver.

Therefore, as fat accumulation in alcoholic fatty liver disease and non-alcoholic fatty liver disease is reported, a cell model that induces fat accumulation in the hepatocyte cell line is made, and treatment of liver disease caused by fat accumulation such as fatty liver of compounds 1 to 13 of the present invention The effect was confirmed.

In order to prepare a cell model that induces excessive accumulation of fat in the hepatocyte cell line, neutralization in hepatocytes is performed by treating carbohydrates (glucose), fatty acids (palmitic acid) or insulin by simulating obesity and insulin resistance, respectively or simultaneously. The accumulation of fat (triglyceride) was confirmed. In more detail, hepG2 cells, a hepatocyte cell line, were cultured in a 12-well plate and treated with 0.1% bovine serum albumin (BSA), 0.2mM palmitic acid, high concentration of glucose (30mM) or high concentration of insulin (50nM) for 24 hours. Fat accumulation with insulin tolerance was induced. Thereafter, the cultured cells were washed with cold phosphate buffered saline (PBS) and treated with 1% triton X-100 to obtain an analytical sample. The obtained analysis sample was analyzed for the amount of triglyceride in the sample using the triglyceride quantitative kit (Fluorometric Serum Triglyceride Quantification, Cell Biolabs, USA) and the experimental method provided by the manufacturer, and the results are shown in FIG. 3. .

As shown in FIG. 3, it was confirmed that the accumulation of triglycerides in HepG2 cells treated with glucose and palmitic acid increased compared to HepG2 cells treated with glucose alone. In addition, it was confirmed that when the insulin was additionally treated with glucose and palmitic acid, the accumulation of triglyceride in HepG2 cells increased more than that with the treatment with glucose and palmitic acid.

Thus, in order to confirm the effect of preventing the accumulation of fat of the compounds 1 to 13 of the present invention, a method of simultaneously treating glucose, palmitic acid, and insulin in HepG2 cells was used.

Example 4-2. Confirmation of the effect of reducing the accumulation of fat in hepatocytes by flavonoid derivative compounds

HepG2 cells, a hepatocyte cell line, were cultured in 12-well plates, and compounds 1 to 13 isolated in Example 1 were respectively treated at a concentration of 40 uM, and 0.1% BSA, 0.2 mM palmitic acid, 50 nM insulin, and 30 mM glucose were treated for 24 hours. Did. Thereafter, the cultured cells were washed with cold PBS and treated with 1% Triton X-100 to obtain an analytical sample. The obtained analysis sample was analyzed for the amount of triglyceride in HepG2 cells treated with each compound using the triglyceride quantitation kit and the experimental method provided by the same manufacturer as the triglyceride analysis method of Example 4-1, The results are shown in Fig. 4 (A).

As shown in Figure 4 (A), compared to the triglyceride content of hepatocytes treated with palmitic acid, glucose and insulin, the triglyceride content in hepatocytes treated with compounds 1 to 13 isolated in Example 1 is reduced. Confirmed. In particular, it was confirmed that the decrease in triglyceride content in hepatocytes treated with compounds 1-4, 6, 11-13 was large.

In addition, the triglyceride reduction effect according to the compound concentration was confirmed by using some compounds (Compounds 1, 3, 4, 13) in the same manner as the method for confirming the effect of preventing the triglyceride accumulation, and the results are shown in FIG. 4 (B). It is shown in. At this time, the compound was treated so that the concentration was 10, 20, and 40 uM, and the treatment with 80 uM of fenofibrate (fenofibrate, Sigma, USA) used as a therapeutic agent for nonalcoholic fatty liver disease was used as a positive control.

As shown in FIG. 4 (B), when the compounds 1, 3, 4, and 13 were treated by concentration, it was confirmed that the content of triglycerides in the hepatocytes was decreased depending on the concentration of the compound treatment.

Example 4-3. Check the effect of preventing fat accumulation in hepatocytes through triglyceride staining

Among the compounds isolated in Example 1, compounds 1, 3, 4, 11 and 13 were treated to confirm the change in triglycerides in hepatocytes using Oil Red O staining.

In the same manner as in Example 4-2, HepG2 cells were treated with each compound and palmitic acid, glucose, and insulin, and instead of the compound, fenofibrate was treated with the same concentration as the compound, and used as a positive control. Thereafter, the cultured cells were treated with 5% formalin for 30 minutes to fix the cells, and then treated with Oil Red O reagent (Sigman, USA) for 15 minutes to stain triglycerides. Thereafter, the cells were washed with a 60% isopropanol solution and washed three times with distilled water. For staining of the cell nucleus, a hematoxylin solution (Sigma, USA) was further treated and reacted at room temperature for 2 minutes. The stained red oil droplets were identified using a fluorescence microscope (Olympus ix70 Fluorescence Microscope, Olympus Corporation, JPN), and the results are shown in FIG. 5.

As shown in FIG. 5, in the case of hepatocytes treated with palmitic acid, glucose, and insulin, a portion stained in red by an oil red OH reagent was remarkably large, whereas fenofibrate used as a positive control and separation in Example 1 In the hepatocytes treated with one compound 1, 3, 4, 11 and 13, it was confirmed that the portion stained with red was reduced.

In addition, although not shown in the specification of the present invention, in Example 1 above using the alcohol-induced fatty liver-induced animal cell model described in Cameron, RG, et al., 1998. and Madushani, HK, et al, 2018. As a result of confirming the effect of inhibiting hepatotoxicity by alcohol by treating the separated compounds 1 to 13, it was confirmed that the decrease in the survival rate of hepatocytes by alcohol was suppressed in all of the hepatocytes to which the compounds 1 to 13 were administered.

Through the above results, compounds 1 to 13, which are flavonoid derivatives isolated from thorns of the present invention, prevent excessive accumulation of fat in liver tissue and suppress the toxicity of liver cells by alcohol, thereby treating alcoholic and non-alcoholic fatty liver. It was found that there is.

Hereinafter, a formulation example of the composition will be described, but the present invention will be described in detail rather than to limit it.

<Formulation Example 1. Pharmaceutical preparation>

Formulation Example 1-1. Preparation of tablets

20 g of compound 4 of the present invention was mixed with 175.9 g of lactose, 180 g of potato starch and 32 g of colloidal silicic acid, respectively. After adding 10% gelatin solution to this mixture, it was ground and passed through a 14 mesh sieve. This mixture was dried, and a mixture obtained by adding 160 g of potato starch, 50 g of talc and 5 g of magnesium stearate was purified.

Formulation Example 1-2. Preparation of injection liquid

100 mg of the compound 4 of the present invention, 0.6 g of sodium chloride and 0.1 g of ascorbic acid were dissolved in distilled water to make 100 ml. The solution was placed in a bottle and sterilized by heating at 20 ° C for 30 minutes.

<Formulation Example 2. Preparation of health functional food>

Formulation Example 2-1. Production of health functional food

20 g of the compound 4 of the present invention, the appropriate amount of the vitamin mixture, 70 μg of vitamin A acetate, 1.0 mg of vitamin E, 0.13 mg of vitamin B1, 0.15 mg of vitamin B2, 0.5 mg of vitamin B6, 0.2 μg of vitamin B12, 10 mg of vitamin C, 10 μg of biotin , Nicotinic acid amide 1.7mg, folic acid 50µg, calcium pantothenate 0.5mg, mineral mixture amount, ferrous sulfate 1.75mg, zinc oxide 0.82mg, magnesium carbonate 25.3mg, potassium phosphate 15mg, dicalcium phosphate 55mg, Potassium citrate 90mg, calcium carbonate 100mg, magnesium chloride 24.8mg was prepared as a granule, but can be prepared by modifying it into various formulations depending on the application. In addition, the composition ratio of the above-mentioned vitamin and mineral mixture may be arbitrarily modified, and the above ingredients may be mixed and prepared according to a conventional health functional food manufacturing method.

Formulation Example 2-2. Production of health functional beverages

A beverage was prepared by mixing 1 g of the compound 4 of the present invention, 0.1 g of citric acid, 100 g of fructooligosaccharide, and 900 g of purified water, followed by stirring, heating, filtering, sterilizing, and refrigerating according to a conventional beverage preparation method.

Claims (24)

Camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1) of Formula 1, Camphorol-3 -O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 2), camphorol-3-O- (6-O- α-L-Lamnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 3), camphorol-3-O-β-D-glucopyranosyl-7 -O-α-L-lamnopyranoside (Compound 4), apigenin-7-O-β-D-glucopyranoside (Compound 5), camphorol-3-O-β-D-glucopyrano Seed (Compound 6), Camperol-3-O- (6-O-α-L-ramnopyranosyl) -β-D-glucopyranoside (Compound 7), Camperol-3-O-β-D -Apiofuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 8), quercetin-3-O- (6-OE-ferulo 1) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 9), quercetin-3-O- (6-OE-coumaroyl) -β-D-glucopyra Nosyl-7-O-α-L-lamnopyranoside (Compound 10), camphorol-3-O- (6-OE -Feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L- Ramnopyranoside (Compound 11), Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1-3 ) -α-L-lamnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α-L-lamnopyranosyl- (1-4) -β-D-glucopyranoside (Compound 13) A pharmaceutical composition for preventing or treating liver disease comprising at least one flavonoid derivative compound selected from the group consisting of active ingredients.
[Formula 1]

According to claim 1,
The flavonoid derivative compound is camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1), camphorol- 3-O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 2), camphorol-3-O- (6-O -α-L-ramnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 3), camphorol-3-O-β-D-glucopyranosyl- 7-O-α-L-lamnopyranoside (Compound 4), camperol-3-O-β-D-glucopyranoside (Compound 6), camperol-3-O- (6-OE-ferrule Royl) -β-D-glucopyranosyl-7-O-α-L-rhamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L-rhamnopi Lanoside (Compound 11), Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1-3)- α-L-lamnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α-L-lamnopyranosyl- (1 -4) -β-D-glucopyranoside (Compound 13) Standing select one liver disease, characterized in that at least one flavonoid derivative compounds preventing or pharmaceutical composition. According to claim 1,
The flavonoid derivative compound is a pharmaceutical composition for preventing or treating liver disease, characterized in that the compound isolated from the extract of thorns ( Sicyos angulatus ). According to claim 3,
The spinach extract is a water, C1 ~ 4 lower alcohol, C1 ~ 4 acetic acid (acetic ester), acetone (acetone), methyl ethyl ketone (methylethylketone) at least one solvent selected from the group consisting of A pharmaceutical composition for the prevention or treatment of liver disease, characterized in that the extracted extract. The method according to any one of claims 1 to 4,
The liver disease is a fatty liver (fatty liver) pharmaceutical composition for preventing or treating liver disease, characterized in that the disease. The method of claim 5,
The fatty liver disease is a non-alcoholic fatty liver disease (non-alcoholic fatty liver disease, NAFLD) or alcoholic fatty liver disease (alcoholic fatty liver disease), pharmaceutical composition for preventing or treating liver disease. Camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1) of Formula 1, Camphorol-3 -O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 2), camphorol-3-O- (6-O- α-L-Lamnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 3), camphorol-3-O-β-D-glucopyranosyl-7 -O-α-L-lamnopyranoside (Compound 4), apigenin-7-O-β-D-glucopyranoside (Compound 5), camphorol-3-O-β-D-glucopyrano Seed (Compound 6), Camperol-3-O- (6-O-α-L-ramnopyranosyl) -β-D-glucopyranoside (Compound 7), Camperol-3-O-β-D -Apiofuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 8), quercetin-3-O- (6-OE-ferulo 1) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 9), quercetin-3-O- (6-OE-coumaroyl) -β-D-glucopyra Nosyl-7-O-α-L-lamnopyranoside (Compound 10), camphorol-3-O- (6-OE -Feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L- Ramnopyranoside (Compound 11), Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1-3 ) -α-L-lamnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α-L-lamnopyranosyl- (1-4) -β-D-glucopyranoside (Compound 13) Health functional food for improving liver disease comprising at least one flavonoid derivative compound selected from the group consisting of.
[Formula 1]

The method of claim 7,
The flavonoid derivative compound is camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1), camphorol- 3-O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 2), camphorol-3-O- (6-O -α-L-ramnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 3), camphorol-3-O-β-D-glucopyranosyl- 7-O-α-L-lamnopyranoside (Compound 4), camperol-3-O-β-D-glucopyranoside (Compound 6), camperol-3-O- (6-OE-ferrule Royl) -β-D-glucopyranosyl-7-O-α-L-rhamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L-rhamnopi Lanoside (Compound 11), Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1-3)- α-L-lamnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α-L-lamnopyranosyl- (1 -4) -β-D-glucopyranoside (Compound 13) Standing health functional food for improving liver diseases, characterized in that one or more flavonoid derivatives are selected. The method of claim 7,
The flavonoid derivative compound is a health functional food for improving liver disease, characterized in that the compound isolated from the extract of thorns ( Sicyos angulatus ). The method of claim 9,
The spinach extract is a water, C1 ~ 4 lower alcohol, C1 ~ 4 acetic acid (acetic ester), acetone (acetone), methyl ethyl ketone (methylethylketone) at least one solvent selected from the group consisting of Health functional food for improving liver disease, characterized in that the extracted extract. The method according to any one of claims 7 to 10,
The liver disease is a health functional food for improving liver disease, characterized in that the fatty liver (fatty liver) disease. The method of claim 11,
The fatty liver disease is a non-alcoholic fatty liver disease (non-alcoholic fatty liver disease, NAFLD) or alcoholic fatty liver disease (alcoholic fatty liver disease) health functional food for improving liver disease. Camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1) of Formula 1, Camphorol-3 -O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 2), camphorol-3-O- (6-O- α-L-Lamnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 3), camphorol-3-O-β-D-glucopyranosyl-7 -O-α-L-lamnopyranoside (Compound 4), apigenin-7-O-β-D-glucopyranoside (Compound 5), camphorol-3-O-β-D-glucopyrano Seed (Compound 6), Camperol-3-O- (6-O-α-L-ramnopyranosyl) -β-D-glucopyranoside (Compound 7), Camperol-3-O-β-D -Apiofuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 8), quercetin-3-O- (6-OE-ferulo 1) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 9), quercetin-3-O- (6-OE-coumaroyl) -β-D-glucopyra Nosyl-7-O-α-L-lamnopyranoside (Compound 10), camphorol-3-O- (6-OE -Feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L- Ramnopyranoside (Compound 11), Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1-3 ) -α-L-lamnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α-L-lamnopyranosyl- (1-4) -β-D-glucopyranoside (Compound 13) An animal drug for the prevention or treatment of liver disease comprising at least one flavonoid derivative compound selected from the group consisting of.
[Formula 1]

The method of claim 13,
The flavonoid derivative compound is camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1), camphorol- 3-O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 2), camphorol-3-O- (6-O -α-L-ramnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 3), camphorol-3-O-β-D-glucopyranosyl- 7-O-α-L-lamnopyranoside (Compound 4), camperol-3-O-β-D-glucopyranoside (Compound 6), camperol-3-O- (6-OE-ferrule Royl) -β-D-glucopyranosyl-7-O-α-L-rhamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L-rhamnopi Lanoside (Compound 11), Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1-3)- α-L-lamnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α-L-lamnopyranosyl- (1 -4) -β-D-glucopyranoside (Compound 13) Standing select one liver disease, characterized in that at least one flavonoid derivatives are the prevention and animal drugs for the treatment. The method of claim 13,
The flavonoid derivative compound is an animal drug for the prevention or treatment of liver disease, characterized in that the compound separated from the extract of thorns. The method of claim 15,
The spinach extract is a water, C1 ~ 4 lower alcohol, C1 ~ 4 acetic acid (acetic ester), acetone (acetone), methyl ethyl ketone (methylethylketone) at least one solvent selected from the group consisting of Animal drug for the prevention or treatment of liver disease, characterized in that the extracted extract. The method according to any one of claims 13 to 16,
The liver disease is an animal drug for preventing or treating liver disease, characterized in that it is a fatty liver disease. Camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1) of Formula 1, Camphorol-3 -O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 2), camphorol-3-O- (6-O- α-L-Lamnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 3), camphorol-3-O-β-D-glucopyranosyl-7 -O-α-L-lamnopyranoside (Compound 4), apigenin-7-O-β-D-glucopyranoside (Compound 5), camphorol-3-O-β-D-glucopyrano Seed (Compound 6), Camperol-3-O- (6-O-α-L-ramnopyranosyl) -β-D-glucopyranoside (Compound 7), Camperol-3-O-β-D -Apiofuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 8), quercetin-3-O- (6-OE-ferulo 1) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 9), quercetin-3-O- (6-OE-coumaroyl) -β-D-glucopyra Nosyl-7-O-α-L-lamnopyranoside (Compound 10), camphorol-3-O- (6-OE -Feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L- Ramnopyranoside (Compound 11), Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1-3 ) -α-L-lamnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α-L-lamnopyranosyl- (1-4) -β-D-glucopyranoside (Compound 13) An animal feed additive for improving liver disease comprising at least one flavonoid derivative compound selected from the group consisting of active ingredients.
[Formula 1]

The method of claim 18,
The flavonoid derivative compound is camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1), camphorol- 3-O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 2), camphorol-3-O- (6-O -α-L-ramnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 3), camphorol-3-O-β-D-glucopyranosyl- 7-O-α-L-lamnopyranoside (Compound 4), camperol-3-O-β-D-glucopyranoside (Compound 6), camperol-3-O- (6-OE-ferrule Royl) -β-D-glucopyranosyl-7-O-α-L-rhamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L-rhamnopi Lanoside (Compound 11), Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1-3)- α-L-lamnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R-guaiacylglyceryl) ether 7-O-α-L-lamnopyranosyl- (1 -4) -β-D-glucopyranoside (Compound 13) Standing selecting one animal feed additive for the improvement of the liver disease, characterized in that at least one flavonoid derivative compound. The method of claim 18,
The flavonoid derivative compound is an animal feed additive for improving liver disease, characterized in that the compound is separated from the thorn extract. The method of claim 20,
The spinach extract is a water, C1 ~ 4 lower alcohol, C1 ~ 4 acetic acid (acetic ester), acetone (acetone), methyl ethyl ketone (methylethylketone) at least one solvent selected from the group consisting of Animal feed additive for improving liver disease, characterized in that the extracted extract. The method according to any one of claims 18 to 21,
The liver disease is an animal feed additive for improving liver disease, characterized in that the fatty liver (fatty liver) disease. Extract of thorns by using thorns as a solvent, at least one selected from the group consisting of water, lower alcohols of C1-4, acetic esters of C1-4, acetone, and methylethylketone Preparing a;
Hexane, ethyl acetate and butanol are sequentially added to the spinach extract to obtain a fraction; And
Chromatography each of the fractions to form camphorol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (compound) 1), camperol-3-O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 2), camperol-3- O- (6-O-α-L-lamnopyranosyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 3), camphorol-3-O-β- D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 4), apigenin-7-O-β-D-glucopyranoside (Compound 5), camphorol-3-O- β-D-glucopyranoside (Compound 6), camphorol-3-O- (6-O-α-L-lamnopyranosyl) -β-D-glucopyranoside (Compound 7), camphorol- 3-O-β-D-apiofuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 8), quercetin-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 9), quercetin-3-O- (6-OE-coumaroyl) -β-D-glucopyranosyl-7-O-α-L-ram Pyranoside (Compound 10), Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 11), camphorol-3-O- (6-OE-feruloyl) -β-D-gluco Pyranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-ramnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R- Guiaacylglyceryl) ether 7-O-α-L-ramnopyranosyl- (1-4) -β-D-glucopyranoside (Compound 13) selected from the group consisting of at least one flavonoid derivative compound How to separate. Novel compound camphorol-3-O- (6-OE-ferruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranoside (Compound 1) ), Camphorol-3-O-β-D-apiofuranosyl-7-O-β-D-glucopyranosyl- (1-3) -α-L-lamnopyranoside (Compound 8), quercetin -3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-ramnopyranoside (Compound 9), camphorol-3-O- (6- OE-feruloyl) -β-D-glucopyranosyl-7-O-α-L-lamnopyranosyl- (1-6) -β-D-glucopyranosyl- (1-3) -α-L -Ramnopyranoside (Compound 11), Camperol-3-O- (6-OE-feruloyl) -β-D-glucopyranosyl-7-O-β-D-glucopyranosyl- (1- 3) -α-L-lamnopyranoside (Compound 12) and trisine 4′-O- (7 ″ R, 8 ″ R-guaiacglyceryl) ether 7-O-α-L-lamnopyranosyl -(1-4) -β-D-glucopyranoside (Compound 13).
[Formula 2]

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