KR20180013204A - A composition comprising ethyl acetate fraction of grasshopper for improving liver function as in gradient compounds - Google Patents

Abstract

The present invention relates to a composition comprising ethyl acetate fractions of grasshopper as an active ingredient for improving liver functions. The ethyl acetate fractions of grasshopper of the present invention promote activity of SOD enzyme and Catalase enzyme and directly recover damaged liver tissue functions, thereby being used as a composition for improving liver functions.

Description

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a composition for improving liver function comprising an ethyl acetate fraction of a locust group as an active ingredient,

The present invention relates to a composition for improving liver function, and more specifically to a composition for improving liver function containing a locust group ethyl acetate fraction as an active ingredient.

The liver is the largest organ in the human body. The main function of the liver is the processing of various substances introduced in the body or produced in the body and synthesizing and supplying important substances. There are also several proteins in the blood that play an important role in our body, about 90% of which are made in the liver. In addition, the liver, detoxification and the role of the immune system, such as the various drugs and harmful substances in our body from liver to harmful substances are changed through urine or bile is excreted. In particular, the liver contains immune cells called Cooper cells, which are able to digest bacteria, toxins or foreign substances coming from outside the body and then decompose them and discharge them out of the body. As the liver performs diverse and important functions, various problems occur when the liver function deteriorates. Liver disease is caused by viral hepatitis, alcoholic liver disease caused by alcohol, toxic liver disease caused by drug, autoimmune liver disease caused by abnormality of human immune system, fatty liver accumulating in liver, etc. depending on root cause of disease Respectively.

Ursodeoxycholic acid, immunostimulants, antiviral agents, silymarin, fingugosaccharides, vitamin B group, and methionine are the most commonly used agents for the prevention and treatment of liver diseases, but none of these agents It has yet to provide clinically satisfactory results. Therefore, in recent years, efforts have been made to find ways to prevent liver damage, as well as to prevent liver damage, without the burden on the liver.

As a hepatic function-protecting agent which is extracted from natural plants and most commonly used in clinical practice, there are silymarin preparations extracted from plants, and Japanese Patent Application Laid-Open Nos. 10-2014-0070734 and 10-1317551, -0499877, and 10-0403722, have also been tried in many efforts to develop therapeutic agents for liver diseases from natural products. However, in relation to the improvement of liver function, the number of naturally derived substances used in clinical practice is only a few.

Accordingly, in connection with the improvement of liver function, there has been developed a composition effective for preventing liver damage and prevention of liver damage derived from natural origin and capable of protecting hepatocytes and having no resistance or stability, and a composition for improving liver function It is an urgent situation.

On the other hand, grasshoppers are insects living in the world, which is the middle of the insects belonging to insects, locusts and locusts. It is known that about 20,000 species are distributed around the world in the tropics, and about 200 species are found in Korea . Grasshopper powder (moisture content 5%) has a heat content of 378Kcal per 100g and is composed of 10.7% of fat and 70.4% of protein, and the protein content is remarkably high However, the use of grasshoppers as food materials is not active. Especially, there are no studies related to the improvement of liver function of locusts.

The inventors of the present invention have found that the ethyl acetate fraction of the extract of the locust bean is excellent in improving the liver function, and as a result, the present inventors have completed the present invention .

KR 10-2014-0070734A, (Patent Document 0002) KR 10-10-1317551B, KR 10-0499877B, (patent document 0004) KR 10-0403722B

Accordingly, a main object of the present invention is to provide a composition for improving liver function, which contains an ethyl acetate fraction of an extract of locust bean as an active ingredient.

According to one aspect of the present invention, there is provided a composition for improving liver function comprising an ethyl acetate fraction of an extract of locust bean as an active ingredient.

The present inventors have made extensive efforts to develop a composition effective for improving liver function. As a result, chloroform fractions, ethyl acetate fractions and hexane fractions were obtained from ethanol extracts of grasshoppers and they were administered to animal models of rats. As a result, compared to other fractions, the activity of superoxide dismutase (SOD) And the blood levels of AST and ALT enzymes used as liver damage markers were reduced. As a result, the inventors found that the present invention can be effectively used for improving liver function.

Accordingly, the present invention relates to a pharmaceutical composition for improving liver function comprising an ethyl acetate fraction of an extract of locust bean as an active ingredient.

The kind of grasshopper used in the present invention is not particularly limited, and includes any grasshopper commonly used in the art, preferably a rice grasshopper (Oxya chinensis sinusosa Mistshenko). According to one embodiment of the present invention, the grasshopper of the present invention is a rice-grasshopper.

The extract of the grasshopper used in the composition of the present invention may be obtained by treating the grasshopper-dried powder with an extraction solvent, and various extraction solvents may be used. According to one embodiment of the present invention, the extraction solvent used in the present invention may be a polar solvent. Suitable as the polar solvent are (i) water, (ii) alcohols such as methanol, ethanol, propanol, butanol, n-propanol, iso-propanol, n-butanol, Ethylene glycol), (iii) acetic acid, (iv) dimethyl-formamide (DMFO) or (v) dimethyl sulfoxide (DMSO). According to one embodiment of the present invention, the extraction solvent of the present invention is ethanol.

The term "extract ", as used herein, has the meaning conventionally used in the art as a crude extract.

The term "fraction " of the present invention means a product obtained by a fractionation method for separating a specific component or a specific group from a mixture containing various constituents. In the present invention, the locust bean extract may preferably be obtained by fractionation by various methods such as a solvent fractionation method, an ultrafiltration fractionation method, and a chromatography fractionation method.

For example, fractions obtained by fractionating the locust extract of the present invention with a solvent having similar physical properties as the ethyl acetate solvent, fractions obtained by passing through an ultrafiltration membrane having a constant molecular weight cut-off value, various chromatographies (depending on size, charge, hydrophobicity or affinity Fractions obtained by various purification methods which are additionally carried out, such as those obtained by separation for the purpose of separation, are also defined as fractions and equivalents of the present invention.

The locust fraction used in the present invention can be prepared in powder form by an additional process such as vacuum distillation and freeze drying or spray drying.

As used herein, the term "comprising as active ingredient " is meant to include an amount sufficient to achieve efficacy or activity of the following locust fraction. The locust fraction of the present invention is a fraction isolated from a locust, which is an edible insect, and there is no adverse effect on human body even when administered in an excessive amount. Therefore, the quantitative upper limit of the locust fraction contained in the composition of the present invention can be selected by a person skilled in the art .

According to one embodiment of the present invention, the ethyl acetate fraction of the locust extract of the present invention promotes the activity of superoxide dismutase (SOD) and catalase enzymes. In the present invention, (SOD) activity increased by 44.5% compared to the control group. In addition, the activity of catalase activity in the liver tissue was about 89.9% as compared with the untreated control group in the LPS-only control group. In the experimental group treated with the ethyl acetate fraction of the locust bean ethanol extract of the present invention, 138.9% Respectively.

According to one embodiment of the present invention, the ethyl acetate fraction of the locust extract of the present invention has been shown to reduce the amount of AST and ALT enzyme released when liver cells are damaged. The ethyl acetate fraction of the locust extract of the present invention showed that the serum AST level was reduced to 63.5% of the LPS treatment group and the serum ALT level was reduced to 33.5%. Based on these results, it was shown that the ethyl acetate fraction of the locust extract of the present invention directly regenerates damaged liver tissue function.

The improvement in liver function in this specification means improving the harmful effects of damage caused by liver tissue by factors known to cause liver disease, and these factors include alcoholic liver disease, chronic viral hepatitis (B type and C type ), Chronic biliary duct obstruction, Wilson's disease, hemochromatosis, exposure to drugs and toxins, autoimmune hepatitis, cystic fibrosis, alpha-anti-trypsin deficiency, obesity and schistosomiasis and the like.

The composition of the present invention may be prepared from a pharmaceutical composition.

According to one embodiment of the invention, the composition of the present invention comprises (a) a pharmaceutically effective amount of the ethyl acetate fraction of the locust extract of the inventive locust as described above; And (b) a pharmaceutically acceptable carrier. As used herein, the term "pharmaceutically effective amount" means an amount sufficient to achieve efficacy or activity of the ethyl acetate fraction of the locust bean ethanol extract described above.

When the composition of the present invention is prepared from a pharmaceutical composition, the pharmaceutical composition of the present invention includes a pharmaceutically acceptable carrier. The pharmaceutically acceptable carriers to be contained in the pharmaceutical composition of the present invention are those conventionally used in the present invention and include lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia rubber, calcium phosphate, alginate, gelatin, But are not limited to, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrups, methylcellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. It is not. The pharmaceutical composition of the present invention may further contain a lubricant, a wetting agent, a sweetening agent, a flavoring agent, an emulsifying agent, a suspending agent, a preservative, etc., in addition to the above components. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington ' s Pharmaceutical Sciences (19th ed., 1995).

The pharmaceutical composition of the present invention may be administered orally or parenterally, and may be administered orally in accordance with an embodiment of the present invention.

The appropriate dosage of the pharmaceutical composition of the present invention may vary depending on such factors as formulation method, administration method, age, body weight, sex, pathological condition, food, administration time, route of administration, excretion rate, . The dose of the ethyl acetate fraction of the locust bean ethanol extract, which is a physiologically active substance contained in the pharmaceutical composition of the present invention, is in the range of 0.01-100 mg / kg on an adult basis.

The pharmaceutical composition of the present invention may be formulated into a unit dosage form by using a pharmaceutically acceptable carrier and / or excipient according to a method which can be easily carried out by those having ordinary skill in the art to which the present invention belongs. Or by intrusion into a multi-dose container. The formulations may be in the form of solutions, suspensions, syrups or emulsions in oils or aqueous media, or in the form of excipients, powders, powders, granules, tablets or capsules, and may additionally contain dispersing or stabilizing agents.

In addition, the composition of the present invention can be provided as a food composition.

When the composition of the present invention is prepared as a food composition, it contains not only the soybean fermentation product of the present invention as an active ingredient but also a component that is ordinarily added at the time of food production, for example, protein, carbohydrate, fat, And flavoring agents. Examples of the above-mentioned carbohydrates are monosaccharides such as glucose, fructose, and the like; Disaccharides such as maltose, sucrose, oligosaccharides and the like; And polysaccharides such as dextrin, cyclodextrin and the like, and sugar alcohols such as xylitol, sorbitol and erythritol. Natural flavors such as tau martin and stevia extract (e.g., rebaudioside A and glycyrrhizin), and synthetic flavors (saccharin, aspartame, etc.) can be used.

For example, when the food composition of the present invention is prepared as a drink, it may further contain citric acid, liquid fructose, sugar, glucose, acetic acid, malic acid, juice, mulberry extract, jujube extract, licorice extract, .

In addition, the composition of the present invention may be provided as a functional food composition.

Functional foods means foods that are recognized as pharmacological effects through individual accreditation by the Korea Food and Drug Administration.

The functional food of the present invention is a food prepared by adding the ethyl acetate fraction of the locust extract of the locust which is an active ingredient of the present invention to food materials such as beverage, tea, spice, gum, confectionery, or by encapsulation, This means that it takes a certain effect on health when consumed, but unlike general medicine, there is an advantage that there is no side effect that may occur when a drug is taken for a long time by using food as a raw material.

In the functional food of the present invention, the ethyl acetate fraction of the extract of the locust bean of the present invention may be added in an amount not detrimental to the original taste of the food, When the food is in the form of powder, granules, tablets, capsules or drinks, it may be added usually in the range of 0.001-100 wt%, 0.01-100 wt% or 0.1-100 wt% with respect to the target food.

The features and advantages of the present invention are summarized as follows:

(a) The present invention provides a pharmaceutical composition for improving liver function comprising an ethyl acetate fraction of an extract of locust bean as an active ingredient.

(b) The ethyl acetate fraction of the locust extract of the present invention promotes the activity of superoxide dismutase (SOD) and catalase enzymes and directly functions to repair the damaged liver tissue function and is used as a composition for improving liver function There is a possible advantage.

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood that the scope of the present invention is not limited by these examples in accordance with the gist of the present invention, and it is to be understood by those skilled in the art that the present invention is not limited thereto It will be obvious.

Example

Throughout this specification, "% "used to denote the concentration of a particular substance, unless stated otherwise, is defined as the percent solids / solids (wt / wt) The liquid / liquid is (vol / vol)%.

< Example  1> grasshopper Fraction  Produce

The rice grasshoppers cultivated for edible purposes were individually harvested from the farms and excised for 3 days by fasting. After sterilization by UV irradiation, 500 g of powdered grasshopper powder was prepared. The prepared rice seeds powder was immersed in 1.5 L of 70% ethanol, and then subjected to low-temperature ultrasonic extraction three times and concentrated under reduced pressure at 0.7 atm to obtain about 250 g of an ethanol extract. The obtained ethanol extract was dissolved in 250 ml of a 20% nucleic acid solution, and the solution was put in a separatory funnel composed of 250 ml of hexane, 250 ml of chloroform, 250 ml of ethyl acetate, and 250 ml of butanol. The ethyl acetate layer was separated and purified in the separatory funnel to give approximately 0.6 g of ethyl acetate fraction.

<Example 2> Verification of liver function improvement effect

2-1 Laboratory animals

Seven-week-old female Sprague-Dawley rats weighing 170 ~ 180g were purchased from Nara ViteTech (Korea, Kangnam, Korea) and fed for 1 week in a laboratory environment (temperature 22-24 ℃, humidity 55-66% , And 12-hour light / dark cycle).

2-2 Experimental group

The experimental group was divided into 4 groups of 3 per group by the nude method so as to have an average weight similar to that of the control group. After 3 weeks of administration of the LPS-treated group and the fraction of Example 1 in the normal control group (no treatment group) (Experimental group). Lipopolysaccharide (LPS), known to cause liver damage similar to hepatitis, constitutes the cell wall of Gram-negative bacteria and causes a variety of responses such as local inflammation, antibody production, and hematemesis.

Specifically, the fraction of Example 1 was administered to Sprague-Dawley rats at 7 weeks of age for 3 weeks per 3 kg of body weight (kg). LPS was dissolved in physiological saline solution at the end of the 3-week period, After intraperitoneal injection, 4 hours after induction of liver injury, anesthetized with ether and sacrificed, blood was collected from the posterior vena cava and serum was separated from the supernatant using a centrifuge. Liver was quenched with liquid nitrogen after weighing and stored at -70 ° C until experiment.

2-3 Measurement of SOD (superoxide dismutase) enzyme activity

SOD enzyme activity was measured using Cyman's SOD assay kit. The experimental procedure was as follows. First, dilute 20 μl of the radial buffer into wells, add 10 μl of Standard and Sample to each well, and add 20 μl of xanthine oxidase. After incubating for 20 minutes in a shaker, the absorbance at 440-460 nm was measured.

Superoxide ions are produced by various biochemical reactions during aerobic metabolism, and the hydroxyl radicals generated from them destroy the tissues and macromolecules and cause them to lose function. The cytoplasmic Cu / Zn SOD, including MnSOD in mitochondria, plays an important role in rapidly converting O ₂ ^- disproportionation to hydrogen peroxide and defending oxygen free radicals. SOD is an enzyme that converts active oxygen (O ^2- ) to less reactive H ₂ O ₂ and is one of the in vivo detoxification systems.

As shown in Table 1 below, the SOD activity of the liver tissue was 60.2% as compared with the untreated group in the LPS treatment group because the SOD activity was decreased as the liver inflammation was induced by the administration of LPS .

In the experimental group treated with the ethyl acetate fraction of the present invention, the SOD activity was elevated again and the liver function was restored to normal, which was 44.5% higher than that of the LPS treatment group. Therefore, it is considered that the ethyl acetate fraction of the locust extract of the present invention increases the activity of the SOD enzyme to inhibit the production of active oxygen, thereby restoring the damaged liver tissue function.

[Table 1]

2-4 Serum With AST  ALT measurement

2-3 SOD Enzyme Activities Serum was separated from the experimental rats after the incubation and stored at -70 ° C, and serum levels of ATS and ATL were measured. The levels of AST (sGOT) and ALT (sGPT) were measured using the IDTox ^TM Aspartate Transaminase (AST) Color Endopoint Assay Kit and the IDTox ^TM Alanine Transaminase (ALT) Color Endopoint Assay kit (ID LABS, Respectively.

AST and ALT enzymes are known to be released when cells become damaged and secreted into the blood, and the amount of these enzymes present in plasma is frequently used as a measure of tissue damage, especially liver damage.

As shown in Table 2, serum ALT levels were increased by about 220% in the LPS-treated group compared to the normal group. These elevated levels were again lowered in the experimental group treated with the ethyl acetate fraction of the extract of LPS and the locust extract of the present invention. The experimental group was increased by 40% compared to the normal group, but the level of the treated fraction of the present invention was 63.5% , The serum ALT level decreased by 33.5%.

In addition, serum AST levels were about 2.5 times higher in the LPS-treated group than in the normal group, and the elevated levels were again lowered in the experimental group treated with the ethyl acetate fraction of the locust extract of the present invention. 52.6%, but the serum AST level was reduced by 23.9% by the fraction treatment of the present invention to 76.1% of the LPS treatment group.

[Table 2]

From the above results, the ethyl acetate fraction of the locust bean ethanol extract of the present invention was shown to directly restore the damaged liver tissue function.

2-5. Measurement of activity of catalase (CAT) in liver tissue

The activity of catalase in liver tissue was measured using a catalase assay kit (cyman, NO 707002). 30 μl of methanol, 100 μl of assay buffer and 20 μl of hydrogen peroxide were dispensed into 20 μl of liver tissue, treated with foil, and reacted at room temperature for 20 minutes. After that, 30 μL of potassium hydroxide and 30 μL of catalasepurpald were mixed and subjected to foil treatment for 10 minutes at room temperature. After addition of 10 μl of catalase potassium periodate, the reaction was allowed to proceed at room temperature for 5 minutes, and the absorbance was measured at 540 nm.

Catalase is the enzyme responsible for the reduction of hydrogen peroxide to water and forms the basis of an antioxidant enzyme system in the body together with SOD. In general, it is known that catalase activity is increased when the activity of SOD is increased due to the generation of active oxygen by various metabolism by stress. Catalase is mainly distributed in intracellular peroxidesomes and mainly acts when H ₂ O ₂ concentration is high.

As shown in Table 3, catalase activity in the liver was about 89.9% in the LPS-treated group and 138.9% in the experimental group.

From the above results, it was found that the ethyl acetate fraction of the extract of the locust bean of the present invention effectively restored the damage of the liver tissue by protecting the tissues of the living body from the peroxides generated during in vivo metabolism by removing active oxygen .

[Table 3]

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the same is by way of illustration and example only and is not to be construed as limiting the scope of the present invention. Accordingly, the actual scope of the present invention will be defined by the appended claims and their equivalents.

Claims (4)

A pharmaceutical composition for improving liver function comprising an ethyl acetate fraction of an extract of locust bean as an effective ingredient. The pharmaceutical composition according to claim 1, wherein the fraction promotes the activity of a superoxide dismutase (SOD) enzyme and a catalase enzyme. A food composition for improving liver function comprising a locust ethyl acetate fraction as an active ingredient. The food composition according to claim 3, wherein the fraction promotes the activity of a superoxide dismutase (SOD) enzyme and a catalase enzyme.