KR20110054880A - Chitosan oligosaccharide comprising of deep sea water for detoxicating alcohol or eliminating alcoholic - Google Patents

Abstract

PURPOSE: A chitosan oligosaccharide containing deep seawater is provided to enhance alcohol dehydrogenase activity, aldehyde dehydrogenase activity, and MEOS activity. CONSTITUTION: A composition for alcohol detoxification and hangover relief contains chitooligosaccharides containing deep seawater as an active ingredient. A method for evaluating the chitosan oligosaccharides comprises: a step of mixing chiosan oligosaccharide and deep seawater; a step of applying to ICR mouse by oral administration of the chitosan mixture; a step of collecting blood from the mouse; and a step of measuring ADH and ALDH enzymes and MEOS activities.

Description

Chitosan Oligosaccharide comprising of deep sea water for detoxicating alcohol or eliminating alcoholic}

The present invention relates to chitosan oligosaccharides containing deep sea water for alcohol detoxification and hangover resolution.

Today, the consumption of alcohol is increasing day by day, chronic or high intake of alcohol causes hangovers and alcoholic liver disease. Alcohol is absorbed by the stomach 25% in 30 minutes in vivo and transported to the small intestine for at least 90%. The alcohol absorbed in the small intestine is mainly metabolized in the liver, and the part not absorbed is excreted in the lungs or urine and sweat. Ingested alcohol is converted to acetaldehyde by alcohol dehydrogenase (ADH), and then oxidized by aldehyde dehydrogenase (ALDH) to acetic acid, which is partially Exposed to ₂ In the case of excessive or chronic alcohol intake, more than 1/2 to 2/3 is metabolized by the microsomal ethanol oxidizing system (MEOS). Alcohol is known to have various effects on liver metabolism depending on the amount of intake. Acetaldehyde produced during oxidation, rather than alcohol itself, causes damage to liver cells. Acetaldehyde, which is produced as an alcohol breakdown product only when excess alcohol is consumed in the body, is transmitted to the brain and converted into many harmful compounds, which can cause symptoms such as increased pulse, sweating, flushing, nausea and vomiting. The metabolic rate of alcohol in the liver can be regulated by several factors that influence the activity of alcohol dehydrogenase, aldehyde dehydrogenase, and MEOS.

Chitin and chitosan have various physiological functions, so they are applied in various industries such as medicine, food, materials, etc., but because they are high molecular materials and water-insoluble, most of them cannot be absorbed in the human body when they are ingested. come. Therefore, physiological activity research is actively conducted by separating water-soluble chitosan oligosaccharides that are easier to absorb in the body than chitin and chitosan, and is expected to be developed as a new functional material in various fields such as food additives, diagnostic reagents, infection inhibitors, and anticancer agents. Deep sea water is a cold water with a depth of more than 200 m that does not reach sunlight. It is rich in nutrients essential for the growth of marine plants, and its application is gradually increasing.

However, there have been no reports of chitosan oligosaccharides or deep seawater for alcohol detoxification and hangover in connection with alcoholic liver disease or disclosed in patent literature.

An object of the present invention is to provide an effect on the alcohol degradation and metabolism of deep sea water-containing chitosan oligosaccharides.

The above object of the present invention comprises the steps of diluting chitosan oligosaccharides (3 kDa <MW <5 kDa) and deep sea water (hardness 4,000); Orally administering the deep seawater-containing chitosan oligosaccharides to an ICR mouse; Quantifying and evaluating blood alcohol using frozen serum after collecting blood from the mice; Extracting the liver from the mouse to measure mitochondria fraction, cytosol and microsome to measure the activity of ADH, ALDH enzyme and MEOS; Achieved through the step of quantifying the protein.

The deep seawater-containing chitosan oligosaccharide of the present invention has not only increased alcohol dehydrogenase activity and aldehyde dehydrogenase activity, but also significantly increased MEOS activity and significantly reduced blood alcohol concentration in clinical animals. There is.

Experimental material

Lyophilized sample chitosan oligosaccharide (3 kDa <MW <5 kDa) used in this experiment was provided from Chitolife Co., Ltd. (Seoul, Korea) and used in the experiment. The deep sea water was taken from 1,100 m of the sea floor in the East Sea and desalted and concentrated with a reverse osmosis system.The deep sea water adjusted to a hardness of 4,000 by adding desalted deep sea water (hardness 0) and minerals was treated with waterbis (Seoul, Dilution was used for the experiment. The mineral content of the desalted deep sea water (hardness 0) and deep sea water with hardness 4,000 is shown in [Table 1].

Animal breeding and diet

Animals used in the present invention was used to purchase a 4 weeks old male ICR mouse of 23 ~ 24g body weight from Orient (Korea). The experiment was carried out after adapting to the environment for one week while supplying sufficient solid feed and water. During the experiment period, the room temperature was 20-22 ℃, the humidity was 40-50%, and the contrast was changed to a 12-hour cycle. The experimental group was the control group [alcohol (25% EtOH) administration group], alcohol + seawater depth (hardness 1,500) administration group, alcohol + chitosan oligosaccharide (2.5mg / day) administration group, alcohol + seawater depth (hardness 1,500) + chitosan oligosaccharide (2.5mg / day) The administration group was divided into 4 groups of 7 animals each. The samples were orally administered and 25% alcohol was orally administered 30 minutes later. Feed intake was measured at regular time each day and body weight was measured at weekly intervals while observing the condition of the animals for a total of three weeks.

Blood, liver odor and enzyme source isolation

After fasting the experimental animals for 24 hours, blood was collected by cardiac sampling, and blood was centrifuged at 1,000 ° C. for 15 minutes at 4 ° C., and serum was separated therefrom. The separated serum was used for blood alcohol concentration analysis. After collecting the blood, the experimental animals were extracted with liver, injected with 0.15 M KCl into the portal vein, perfusion of the liver, extracted and washed with 0.15 M KCl, and 0.25M sucrose was added at a rate of 9 ml per 1 g of pure liver tissue. It was crushed using. The homogenate was centrifuged at 600 × g for 10 minutes to remove debris, the supernatant was again centrifuged at 10,000 × g for 20 minutes, and then 0.2 ml of 0.25M sucrose solution containing 0.1M CaCl ₂ in pellet. (1/5) was added to obtain the mitochondria fraction to be used as an enzyme source for ALDH activity measurement. The supernatant was ultracentrifuged at 105,000 × g for 1 hour to separate the microsomal fraction and cytosolic fraction. The microsomal fraction was resuspended in 0.25M sucrose solution containing 0.1M CaCl ₂ and used for measuring MEOS activity. The cytosolic fraction was used as an enzyme source for measuring ADH activity.

Alcohol dehydrogenase

Alcohol dehydrogenase activity was measured at 340 nm to change the absorbance according to NADH production. The reaction solution was mixed with 0.05M Glycine-NAOH buffer (pH 9.6), 0.58% ethanol, 13.3mM NAD sol 'and 1mg protein cytosol, stabilized at 37 ° C for 5 minutes, and the absorbance was measured. Activity was calculated | required by the change amount.

Aldehyde dehydrogenase

Aldehyde dehydrogenase activity was measured at 340 nm to change the absorbance according to NADH production. The reaction was initiated using propionaldehyde as a substrate, and the composition of the ALDH reaction solution was mixed with 70 mM Na₄P₂O ₇ · 10 H₂O 2, 16.7 mM pyrazole, 60 mM propionaldehyde, 13.3 mM NAD, and 1 mg protein mitochondria as an enzyme source for 5 minutes at 37 ° C. After stabilization, the absorbance was measured and the activity was indicated by the amount of change.

MEOS activity measurement

MEOS activity was measured at a ratio of 1: 1 with 3uM NADP, 5uM MgCl ₂ , 20uM nicotinamide, 8uM glucose-6-phosphate, 50uM ethanol, and 80uM KP buffer (PH 7.4) in the out well of the Erlenmeyer flask type reaction vessel. One microsome and a cytosol fraction were added, 0.16M potassium phosphate buffer containing 0.015M semicarbazide HCl was added to the center well and reacted at 37 ° C. for 30 minutes. The absorbance of acetaldehyde-semicarbazone produced by standing for 24 hours was measured at 224 nm.

Blood alcohol level

Blood alcohol was analyzed using Kit (enzyme ethanol assay kit, BioChain Institute, Inc).

Protein Quantitation

Protein quantitation was measured using bovine serum albumin (Sigma) as a standard according to the method of Lowry et al.

Alcohol dehydrogenase, aldehyde dehydrogenase and MEOS activity were measured in the livers of experimental animals. Dehydrogenase activity was increased by 5% in the chitosan oligosaccharide administration group (Fig. 1). Aldehyde dehydrogenase activity was increased by 10% in the deep seawater and chitosan oligosaccharide mixed administration group (Fig. 2). 45% increase was observed compared to the control group (Fig. 3). Blood alcohol concentration was decreased by 20% (p <0.05) in the deep seawater treatment group and 47% (p <0.01) in the seawater and chitosan oligosaccharide mixed group (p <0.01). 4). According to the present invention, it was determined that the treatment of hangover and liver disease could be prevented by reducing the blood alcohol concentration by administration of the deep seawater-containing chitosan oligosaccharide.

The deep sea water-containing chitosan oligosaccharide of the present invention has an excellent effect on alcohol decomposition and metabolism in experimental animals to which alcohol is administered, and thus can be used as a substance for preventing and treating alcoholic liver disease in the human body.

Figure 1 It is a graph showing the effect of deep seawater-containing chitosan oligosaccharides on the ADH activity of the liver of the experimental animal mice (ICR mouse).

2 is It is a graph showing the effect of deep seawater-containing chitosan oligosaccharides on ALDH activity of the liver of the experimental animal mice (ICR mouse).

Figure 3 is a graph showing the effect of deep seawater containing chitosan oligosaccharides on the MEOS activity of the liver of the experimental animal mice (ICR mouse) of the present invention.

Figure 4 is a graph showing the effect of deep seawater containing chitosan oligosaccharides on the blood alcohol concentration of the experimental animal rats (ICR mouse) of the present invention.

Claims (1)

A deep sea water containing chitosan oligosaccharide as an active ingredient for alcohol detoxification and hangover relief composition.

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