KR20170026240A - Health functional food composition for improving liver function - Google Patents

Abstract

The present invention relates to a functional health food composition for improving liver functions, containing vitamin D, vitamin B12, vitamin B6, vitamin B9, and choline.

Description

{Health functional food composition for improving liver function}

The present invention relates to a health functional food composition for improving liver function comprising vitamin D, vitamin B12, vitamin B6, vitamin B9 and choline.

Liver disease is becoming an important cause of adult disease and death in Korea. The liver is an organ with a large buffer capacity. Since the liver is not aware of the symptoms at an early stage of the disease, the disease is often found in a state of advanced disease, and effective therapeutic agents and diagnostic methods are lacking. The causes of liver disease include alcohol, drugs, chemicals, hepatitis viruses, metabolic diseases such as obesity and insulin resistance, and autoimmune diseases. In addition, the liver is the center of the blood storage and circulation, blood volume control and detoxification in the human body. Recently, as the industrialization has increased opportunities for exposure to harmful substances, the liver has been suffering from continuous detoxification. Especially, And the liver damage caused by the causes of the immune system is caused by abnormalities, as well as other diseases are increasing the cause.

In most of the liver diseases, complications and liver cancer, which have a great influence on the prognosis of the disease, are mostly developed after hepatic fibrosis and cirrhosis, so they understand the progress of liver disease and inhibit its progress. And the development of a liver function-improving agent.

Korean Patent Laid-Open No. 10-2014-0095167 discloses a health food having an effect of improving liver function, Honkiool and Magnolol are described as effective ingredients, and Korean Patent Laid-Open No. 10-2014-0064507 discloses a health food having liver function And Korean Patent Laid-Open No. 10-2014-0019456 discloses a composition for relieving hangover comprising an extract of Omija. However, the development of a more economical, safe and effective agent for improving liver function is urgently required.

Under these circumstances, the present inventors have completed the present invention by developing a new composite composition containing five active ingredients and confirming that the composition is effective for improving liver function.

Korean Patent Publication No. 10-2014-0095167 Korean Patent Publication No. 10-2014-0064507 Korean Patent Publication No. 10-2014-0019456

It is an object of the present invention to provide a health functional food composition for improving liver function comprising vitamin D, vitamin B12, vitamin B6, vitamin B9 and choline.

It is also an object of the present invention to provide a health functional food composition for preventing or ameliorating liver diseases including vitamin D, vitamin B12, vitamin B6, vitamin B9 and choline.

In one aspect of the present invention, there is provided a health functional food composition for improving liver function comprising vitamin D, vitamin B12, vitamin B6, vitamin B9 and choline.

In the present invention, 'vitamin D' is a fat-soluble vitamin, and vitamin D2 (ergocalciferol) is synthesized from yeast and plant sterol ergosterol. Vitamin D3 (cholecalciferol) is synthesized from cholesterol precursor in skin cells, ≪ / RTI >

In the present invention, 'vitamin B12' refers to an anti-anemic agent, also called cyanocobalamine.

In the present invention, 'vitamin B6' is a substance present in the form of pyridoxine, pyridoxal, pyridoxamine or phosphorylated form in food, and three kinds of the above-mentioned pyridoxine, pyridoxal and pyridoxamine It is known that it is able to convert in vivo, is a component of enzymes important for protein metabolism, and is involved in the hem synthesis process, which is a component of hemoglobin.

In the present invention, 'vitamin B9' refers to folic acid which is a water-soluble vitamin of the vitamin B group.

In the present invention, 'choline' is a type of vitamin B complex, and is a lipotropic factor. It is a component of complex lipids such as lecithin and plasmarogens, and is a component of acetylcholine, It is known.

In the present invention, the vitamin D is cholecalcidiol, cholecalcitriol or ergocalciferol, vitamin B12 is a cobalamine derivative, vitamin B6 is pyridoxal or pyridoxamine, vitamin B9 Can be replaced by folate salts, choline can be replaced by lecithin or methionine, and can be used interchangeably.

In the present invention, the vitamin D, vitamin B12, vitamin B6, vitamin B9 and choline may be used in the free base form, derivatives thereof, or salts known in the art.

In the present invention, vitamin D, vitamin B12, vitamin B6, vitamin B9, and choline are commercially available, synthesized by methods known in the art, used in nature or cultivated in nature, The obtained plants may be used, but not limited thereto, obtained by treating them with a method known in the art.

For the purpose of the present invention, the above-mentioned vitamin D, vitamin B12, vitamin B6, vitamin B9 and choline are used for improving liver function.

The vitamin D of the present invention is preferably vitamin D3. The vitamin D3 is preferably selected from the group consisting of 900 IU to 20,000 IU, 1000 IU to 20,000 IU, 1100 IU to 20,000 IU, 1500 IU to 20,000 IU, 1800 IU to 20,000 IU, 2000 IU to 20,000 IU, IU to 20,000 IU, or 10,000 IU to 20,000 IU, more preferably 1000 IU to 20,000 IU, 2000 IU to 20,000 IU, 5000 IU to 20,000 IU, particularly preferably 1000 IU to 10,000 IU, or 2000 IU To 10,000 IU, or 1000 IU, 2000 IU, 5000 IU, 10,000 IU, or 20,000 IU.

The composition of the present invention, which contains vitamin D, specifically vitamin D3 in the above contents, is extremely excellent in improving liver function such as effectively lowering ALT, AST, and BUN values upon administration.

In the composition of the present invention, vitamin B12 is contained in an amount of 0.003 mg to 3 mg, vitamin B6 is contained in an amount of 0.1 mg to 500 mg, vitamin B9 is contained in an amount of 0.05 mg to 4 mg, ≪ / RTI >

In the present invention, vitamin B12, vitamin B6, vitamin B9 and choline have a common function as a group, and their typical functions are as follows: 1) Acceleration of action of liver regeneration genes, 2) (3) DNA methylation, (5) repair of injury genes, (6) synthesis of red blood cells and white blood cells, (7) synthesis of proteins, fats and carbohydrates into polymers, (8) The synthesis of fatty acids, 9) the elimination of homocysteine, which causes deterioration of hepatitis, liver cirrhosis or liver cancer in liver disease, and 10) the detoxification by methylating toxic substances.

The composition of the present invention preferably further comprises at least one selected from vitamin E, vitamin C, milk-thistle extract and Omija extract.

For the purpose of the present invention, the vitamin E, vitamin C, milk-sis extract and omija extract are used for improving liver function.

Preferably, the composition of the present invention may comprise vitamin D, vitamin B12, vitamin B6, vitamin B9, choline, vitamin E and vitamin C.

Preferably, the composition of the present invention may include vitamin D, vitamin B12, vitamin B6, vitamin B9, choline, vitamin E, vitamin C, milk shise extract and omija extract.

In the present invention, 'vitamin E' is a fat-soluble vitamin and is a tocol derivative. There are eight kinds of α-, β-, γ-, and δ-tocopherol and α-, β-, γ-, and δ-tocotrienols in nature.

In the present invention, 'vitamin C' includes all of the reduced type ascorbic acid and oxidized form dehydroascorbic acid which can be converted to each other, and is related to the formation of connective tissues and supporting tissues, It is known to be a substance.

A representative function of the above-mentioned vitamins E and C is a continuous oxidative stress relieving action of ROS (Reactive oxygen species).

The liver has more than 500 functions in a single cell, and over 2,000 biochemical reactions are continuously developed. A myriad of biochemical reactions are mainly redox reactions, resulting in very high levels of ROS in normal conditions. ROS depletes an electron from a nearby enzyme, protein or lipid of the cell membrane, oxidizes and destroys the enzyme, oxidizes and destroys the protein, and oxidizes the lipid to lipid peroxide. Vitamins E and C effectively eliminate oxidative stress caused by ROS, thereby preventing and treating liver diseases, 2) preventing and treating hepatitis, 3) preventing and treating cirrhosis, and 4) preventing and treating liver cancer. .

"Milk Thistle" In the present invention, Silybum marianum Refers to a substance obtained by separating from milk thistle. Specifically, it is a conventional extraction solvent known in the art, for example, water, C1-4 alcohol (for example, : Methanol, ethanol, butanol, etc.), or a mixture of alcohol and water, and the like.

The milk-syrup extract of the present invention can be obtained by various known extraction methods such as hot water extraction, organic solvent extraction, supercritical extraction, and the like, and can be obtained by an additional process such as vacuum distillation or freeze- .

The milk thistle extract of the present invention contains silymarin as an active ingredient. Preferably the milk thistle extract of the present invention contains 65% by weight of silymarin and other active ingredients (e.g., silibinin (sylibin: sylvin A + sylibin B), iso-sylvinin, By weight to 80% by weight.

It is known that the silymarin stabilizes the hepatocyte membrane, inhibits intracellular inflow of harmful substances, activates protein synthesis of hepatocytes, and promotes regeneration of hepatocytes. Especially, it has a strong antioxidative effect, and decreases phospholipid synthesis, excessive accumulation of neutrophil , Reduction of serum LDL, anti-inflammation, anticancer, acute viral hepatitis, chronic hepatitis, liver cirrhosis complication, and it is known that the treatment of various liver diseases caused by smoking, drinking, Have been widely applied.

In the present invention, Schisandra chinensis, Schisandra is chinensis (Turcz.) Baill. "Omija extract" means a substance obtained by separating from Omija. Specifically, it is a conventional extraction solvent known in the art, for example, water, C1-4 Means a substance obtained by using an alcohol (for example, methanol, ethanol, butanol, etc.) or a mixed solvent of the alcohol and water.

The Omiza extract of the present invention can be obtained by various known extraction methods such as hot water extraction, organic solvent extraction, supercritical extraction, etc., and can be produced in powder form by an additional process such as vacuum distillation, freeze drying, .

The Schizandrin extract of the present invention can be used as an active ingredient in combination with a schisandrin-based compound such as schisandrin A, schisandrin B, schisandrin C, , Gomisin A or gomicin N. The term " gomicin "

The above-mentioned Schizandra chinensis extract has antioxidant activity, fatty liver improvement effect, normalizing action of ALT elevation, and is known to help prevent and treat viral hepatitis, chronic hepatitis and liver cancer.

In the present invention, the milk-thistle extract can be replaced with silymarin, the omija extract can be replaced with biphenyldimethyldicarboxylate, and can be used interchangeably.

In the present invention, the vitamin E and vitamin C may be used in the free base form, derivatives thereof, or salts known in the art.

In the present invention, vitamin E, vitamin C, milk shise extract, and omija extract are commercially available, or they are synthesized by methods known in the art, used in nature or cultivated, The plant obtained by treating the plant with a method known in the art can be used, but is not limited thereto.

In the pharmaceutical composition of the present invention, the vitamin E is contained in an amount of 10 IU to 1,000 IU, the vitamin C is contained in an amount of 10 mg to 2,000 mg, the milk syrup extract is contained in an amount of 50 mg to 5,000 mg, May be included in an amount of 5,000 mg.

The health functional food composition of the present invention is used as a liver function improving agent.

In the present invention, 'improvement' is meant to include treatment, alleviation and prevention of symptoms. In the present invention, it means that the liver function, liver condition, liver disease and liver disease are treated, alleviated or prevented.

In the health functional food composition of the present invention, vitamin B12, vitamin B6, vitamin B9, vitamin D, vitamin E, vitamin C, milk syrup extract and omija extract are used for the purpose of formulation, May be included in an arbitrary amount (effective amount).

The term " effective amount " as used herein refers to the amount of active ingredient capable of inducing liver function improving effect. Such effective amounts can be determined experimentally within the ordinary skill of those skilled in the art.

In another aspect, the present invention provides a health functional food composition for preventing or ameliorating liver disease comprising vitamin D, vitamin B12, vitamin B6, vitamin B9 and choline.

The composition of the present invention preferably further comprises at least one selected from vitamin E, vitamin C, milk-thistle extract and Omija extract.

The above-mentioned vitamin D, vitamin B12, vitamin B6, vitamin B9, choline, vitamin E, vitamin C, milk shise extract and omija extract are as described above.

The term "liver disease" as used herein includes, but not limited to, non-alcoholic fatty liver disease, alcoholic fatty liver disease, hepatitis, liver cirrhosis, liver cancer, etc., which can be treated by the composition of the present invention.

The fatty liver refers to a phenomenon in which abnormally deposited fat cells appear in liver cells unlike the case of normal triglycerides. In normal liver, about 5% is composed of adipose tissue, and triglyceride, fatty acid, phospholipid, cholesterol and cholesterol ester are the major components of fat. However, once fatty liver occurs, most of the components are replaced by triglycerides. If the liver weight is 5% or more, it is diagnosed as fatty liver. Fatty liver is caused by fat metabolism disorder in hepatocytes or defects in the process of transporting excess fat, and it is mainly caused by fat metabolism disorder in the liver. Most of the fat accumulated in the liver is triglyceride, and fatty liver can be roughly divided into non-alcoholic fatty liver due to obesity, diabetes, hyperlipidemia, drug, and alcoholic fatty liver due to excessive drinking.

The nonalcoholic fatty liver refers to a case of fatty liver without a history of alcohol consumption and is known to be associated with metabolic diseases such as obesity, diabetes and hyperlipidemia. Non-alcoholic steatohepatitis or end-stage fibrosis liver disease include not only the accumulation of fat in the liver, but also non-alcoholic fatty liver disease.

In addition, the alcoholic fatty liver means that a large amount of alcohol is consumed to promote fatty acid synthesis in the liver and normal energy metabolism is not performed. Alcohol is not stored in the body and is completely eliminated by oxidation in the liver. Specifically, the alcohol in the liver is largely divided into the alcohol dehydrogenase (ADH) pathway, the microsomal ethanol oxidizing system (MEOS ) Pathway and the catalase pathway, which are then converted to acetaldehyde, which in turn is metabolized to the acetyl salt by dehydrogenase (ALDH). At this time, acetaldehyde is poisonous and can damage hepatocytes. Also, as a result of such alcohol metabolism, a large amount of fatty acid is produced, and liver fat accumulates. Especially, the toxic oxygen (ROS) rapidly increasing due to alcohol destroys the lipid metabolizing enzyme And lipid peroxidation accumulates. The fatty liver accumulated in the liver due to the above-mentioned causes is referred to as an alcoholic fatty liver.

In addition, cirrhosis refers to all forms of liver disease characterized by significant loss of hepatocytes, proliferation of fibrous tissue, and regenerative nodules. The main cause of cirrhosis is hepatitis virus, alcohol, and the harmful oxygen (RO) caused by metabolic syndrome. Therefore, the gene of the hepatic stellate cell with storage function is altered to produce Type 1 collagen fiber Liver cirrhosis is caused by fibrosing of cirrhotic cells, and cirrhosis is induced by antiretroviral treatment, abstinence or nutritional supply and mass therapy.

The above-mentioned liver cancer refers to a disease caused by a patient suffering from chronic liver disease caused by a liver cirrhosis patient, alcohol or the like, and is diagnosed as a liver cancer if a typical hepatic mass is found or an AFP is increased in a blood test.

The cause of all liver diseases may vary from patient to patient, but chronic hepatitis (ROS) increases rapidly with hepatitis, liver cirrhosis, and liver cancer.

The composition of the present invention contains a complex active ingredient of vitamin D, vitamin B12, vitamin B6, vitamin B9, and choline, thereby preventing ROS from increasing in liver disease and producing a destroyed enzyme. And healing effects, and can be used as a preventive and remedy agent for various liver diseases.

In addition, the composition of the present invention is useful for the treatment of Aspartate Aminotransferase (AST), Alanine Aminotransferase (ALT), and Blood Urea Nitrogen (BUN) (SOD activity), and thus can be useful as a remedy for liver damage and a liver function restorer.

In the present invention, " prevention " refers to any action that inhibits or delays the onset of liver disease by the administration of the composition according to the present invention.

The health functional food composition of the present invention is a food prepared and processed in the form of tablets, capsules, powders, granules, liquids, and rings using raw materials and components having useful functions in the human body Article 3 (1) of the Act). The term "functional" as used herein refers to the structure and function of the human body, which has a beneficial effect on health uses such as controlling nutrients or physiological actions. In other words, it means that it can be useful for the health use of healthy people or half-health people.

The health functional food composition may contain various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, colorants and heavies (cheese, chocolate etc.), pectic acid and its salts, alginic acid and its salts , Organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonating agents used in carbonated drinks, and the like. In addition, it may contain natural fruit juice and pulp for the production of fruit juice drinks and vegetable drinks. These components may be used independently or in combination. In addition, the health functional food may be in the form of any one of meat, sausage, bread, chocolate, candy, snack, confectionery, pizza, ramen, gum, ice cream, soup, beverage, tea, functional water, Lt; / RTI >

In addition, the above food composition or health functional food composition may further include a food additive, and the suitability of the additive as a 'food additive' is not limited to the General Rules for Food Additives approved by the Food and Drug Administration, It shall be determined according to the standards and standards for the item.

Examples of the above-mentioned 'food additives' include natural products such as ketones, chemical products such as glycine, potassium citrate, nicotinic acid, cinnamic acid, coloring matter, licorice extract, crystalline cellulose, guar gum, Sodium laurate, sodium glutamate preparation, noodles-added alkaline agent, preservative agent, tar pigment preparation and the like.

The health functional food composition of the present invention may contain additives for nutritional supplementation, additives for improving functionalities and formulations within the range not impairing the effects of the present invention.

The health functional food composition of the present invention is excellent in the effect of protecting and healing from damage of liver cells including a complex active ingredient of vitamin D, vitamin B12, vitamin B6, vitamin B9 and choline, And can be widely used as an improving agent.

Figure 1 is a graph of elution in water of tablets according to Example 1;
FIG. 2 is a graph showing changes in body weight according to administration of a single composition according to Example 5,
Figure 3 is a graph comparing liver weights according to administration of a single composition according to Example 5,
4 is a graph comparing the weights of kidneys according to administration of a single composition according to Example 5,
FIG. 5 is a graph comparing the weights of hearts according to the administration of a single composition according to Example 5. FIG. (* And ** mean statistical significance with G2, representing P <0.05 and P <0.01, respectively).
FIG. 6 is a graph comparing the values of the amino acid transferase (AST) according to administration of the single composition according to Example 5,
7 is a graph comparing the values of alanine aminotransferase (ALT) according to administration of a single composition according to Example 5,
8 is a graph comparing the antioxidant activity (SOD activity) of liver according to administration of a single composition according to Example 5. Fig.

Hereinafter, preferred embodiments of the present invention will be described in order to facilitate understanding of the present invention. However, the following examples are provided to further understand the present invention, and the present invention is not limited by the examples.

Reference Example 1: Materials

Vitamin B12 in DSM (Switzerland) or LYCORED, vitamin B6 in DSM (Germany), vitamin B9 in DSM (Switzerland), hydrogen stearate choline in Balcehm (Italy) or Algry, vitamin D in DMS , Vitamin E in DSM (Switzerland), Vitamin C in DSM (UK), Milk Thistle extract in Monteloeder (Spain), Silymarin in Teva (Israel) or Monteloeder, Omiza X in Kosice Biphenyl Dimethyl Dicarboxylate was purchased from Zhejiang Hisoar and used in Joe Foodtek (Korea). The sterilized water for injection of 0.5% carboxymethylcellulose (CMC) was purchased from CMC Co., Ltd. and sterilized water for injection was purchased from Korea Pharmaceutical Industry Co., Ltd. Also, D-galactosamine was purchased from Sigma-Aldrich Ltd and used.

The vitamin D used in Examples 1 to 4 was 90,000 ~ 110,000 IU / g of vitamin D3 and the vitamin D was used according to the content described in Example 5. Also, vitamin B12 (1% powder) is a product obtained by dissolving vitamin B12 with an excipient (calcium phosphate), which means that 1% by weight of vitamin B12 is included in the vitamin B12 (1% powder), and vitamin E mixed powder E 50%) means that 50% by weight of vitamin E is contained in the mixed powder.

Reference Example 2: Preparation of an animal model

Seven-week-old Sprague-Dawley rats (SD) were purchased from Orient Bio (Seongnam, Korea) and used for the experiment. (W235 * L380 * H175, 3 animals / breeding box) of the Gyeonggi Bio Center Animal Breeding Area No. 5 of the Gyeonggi Bio Center where the test was carried out for 7 days. The breeding box was changed at least once a week.

The conditioning and test conditions were maintained at a temperature of 23 ± 3 ° C, a relative humidity of 55 ± 15%, a ventilation frequency of 10 to 20 times / hour, a lighting time of 12 hours (8:00 am to 8:00 pm off) Respectively. Temperature and relative humidity were measured every hour using a computer system, and the number of times of ventilation and illumination were measured periodically.

The feed was fed free from feedstuffs (Teklad certified irradiated global 18% protein rodent diet; 2918C, ENVIGO, UK) supplied by Doosan Biotech Co., Ltd. in a feeder.

The water was sterilized with ultraviolet sterilizer and microfiltration device and placed in a polycarbonate water bottle for free ingestion. The water quality test was carried out by Gyeonggi Provincial Health and Environment Research Institute and it was confirmed that it conformed to the water quality standard of drinking water. Negative bottles were replaced at least once a week.

The ethical rules of experimental animals were approved by the Experimental Animal Care Committee of the Gyeonggi Bio Center (Serial No .: 2016-04-0010).

Reference Example 3: Statistical Analysis

Statistical analysis was performed using SPSS statistics 22 for medical science, and the results were analyzed using ONE-WAY ANOVA and Student's t-test. Normal group and single composition administration group were compared with excipient control group, and P <0.05 was considered statistically significant.

Example 1: Preparation of a single tablet

As shown in Table 1 below, the mixture was mixed with microcrystalline cellulose mixed with vitamin B12, vitamin B6, vitamin B9, hydrogen choline starch hydrolyzate, vitamin D, vitamin E, vitamin C, milk cisse extract and Omija extract, , Germany). Separately, hydroxypropylcellulose was dissolved in a 1: 1 mixture of water and ethanol, and the resulting binding solution was fed to a high-speed mixing unit containing the mixture. After coalescence, the mixture was granulated using a 25-point tumbler, dried at 60 ° C using a dryer (Jayutec, Korea), and then dried again with a No. 25 sieve after drying. The thus-prepared granules were mixed with a V-type mixer (Erweka, Germany), and sodium starch glycolate and silicon dioxide were mixed with the mixture. Magnesium stearate was added thereto and mixed with a V-type mixer. The mixture was compressed into 980 mg of a tablet using a rotary tablet machine (Sejong, Korea), and then a film coating layer was formed using a high-coater (Sejong, Korea) to prepare a single tablet of 1,020 mg per tablet.

Example  2: Preparation of a single tablet

As shown in Table 1 below, the mixture was mixed with microcrystalline cellulose mixed with vitamin B12, vitamin B6, vitamin B9, hydrogen choline starch hydrolyzate, vitamin D, vitamin E, vitamin C, milk cisse extract and Omija extract, , Germany). Separately, hydroxypropylcellulose was dissolved in a 1: 1 mixture of water and ethanol, and the resulting binding solution was fed to a high-speed mixing unit containing the mixture. After the association, the mixture was granulated using a 25-neck tumbler, dried at 60 ° C using a drier (Jayutec, Korea), and then dried with a No. 25 sieve. After drying, the granulated product was placed in a V-type mixer (Erweka, Germany) , Mixed with croscarmellose sodium and silicon dioxide, and then magnesium stearate was added thereto and mixed in a V-type mixer. The mixture was compressed into 980 mg of a tablet using a rotary tablet machine (Sejong, Korea), and a film coating layer was formed using a high-coater (Sejong, Korea) to prepare a tablet of 1,020 mg per tablet.

Example 3: Preparation of a single tablet

1) Preparation of primary granules

Vitamin B12, vitamin B6, vitamin B9, and hydrogenated choline powder were mixed with microcrystalline cellulose and sieved through a No. 35 sieve and mixed in a high-speed mixer (Diosna, Germany) as shown in Table 2 below. Separately, hydroxypropylcellulose was dissolved in water to prepare a binding solution, which was then combined with the mixture in the high-speed mixer. After coalescence, the mixture was granulated using a 25-point tumbler, dried at 60 ° C using a dryer (Jayutec, Korea), and then dried again with a No. 25 sieve after drying.

2) Manufacture of secondary granules

Vitamin D, vitamin E, vitamin C, milk syrup extract and Omija extract were mixed with microcrystalline cellulose, sieved through a No. 35 sieve, and mixed in a high-speed mixer (Diosna, Germany) as shown in Table 2 below. Separately, hydroxypropylcellulose was dissolved in water to prepare a binding solution, which was then combined with the mixture in the high-speed mixer. After coalescence, the mixture was granulated using a 25-point tumbler, dried at 60 ° C using a dryer (Jayutec, Korea), and then dried again with a No. 25 sieve after drying.

3) After mixing, tableting and coating

The primary granules and the secondary granules prepared in the above steps 1) and 2) were mixed with a V-type mixer (Erweka, Germany), and sodium starch glycolate and silicon dioxide were mixed therein. Then, magnesium stearate And finally mixed with a V-type mixer. The final mixture was tableted to 960 mg per tablet using a rotary tablet machine (Sejong, Korea), and then a film coating layer was formed using a high-coater (Sejong, Korea) to prepare a single tablet of 1,000 mg per tablet.

Example 4: Preparation of a single tablet

1) Preparation of primary granules

Vitamin B12, vitamin B6, vitamin B9, and hydrogenated choline powder were mixed with microcrystalline cellulose and sieved through a No. 35 sieve and mixed in a high-speed mixer (Diosna, Germany) as shown in Table 2 below. Separately, polyvinylpyrrolidone was dissolved in water to prepare a binding solution, which was then combined with the mixture in the high-speed mixer. After coalescence, the mixture was granulated using a 25-point tumbler, dried at 60 ° C using a dryer (Jayutec, Korea), and then dried again with a No. 25 sieve after drying.

2) Manufacture of secondary granules

Vitamin D, vitamin E, vitamin C, milk syrup extract and Omija extract were mixed with microcrystalline cellulose, sieved through a No. 35 sieve, and mixed in a high-speed mixer (Diosna, Germany) as shown in Table 2 below. Separately, polyvinylpyrrolidone was dissolved in water to prepare a binding solution, which was then combined with the mixture in the high-speed mixer. After coalescence, the mixture was granulated using a 25-point tumbler, dried at 60 ° C using a dryer (Jayutec, Korea), and then dried again with a No. 25 sieve after drying.

3) After mixing, tableting and coating

The primary granules and the secondary granules prepared in the above steps 1) and 2) were mixed with a V-type mixer (Erweka, Germany), mixed with croscarmellose sodium and silicon dioxide, and then magnesium stearate was added V-type mixer. The mixture was compressed to 960 mg per tablet using a rotary tablet machine (Sejong, Korea), and then a film coating layer was formed using a high-coater (Sejong, Korea) to prepare a single tablet of 1,000 mg per tablet.

Example  5: Preparation of a single composition

The following components of Table 3 were weighed and mixed in the stated amounts to prepare a single composition (A1-A3).

Component (unit) A1 A2 A3 Vitamin D3
(DSM, Germany) (IU) 2,000 5,000 10,000 Vitamin B12
(Lycored) (mg) One One One Vitamin B6
(DSM, Germany) (mg) 60 60 60 Vitamin B9
(DSM, Switzerland) (mg) 0.4 0.4 0.4 Hydrogen stearate choline
(Balchem, Italy) (mg) 50 50 50 Vitamin E
(DSM, Switzerland) (IU) 100 100 100 Vitamin C
(DSM, UK) (mg) 100 100 100 Milk Thistle Extract
(Monteloeder, Spain) (mg) 130 130 130 Omija extract
(Cho Eun Foodtec, Korea) (mg) 400 400 400

In addition, the following components of Table 4 were weighed and mixed in the stated amounts to prepare a single composition (B1-B3).

Component (unit) B1 B2 B3 Vitamin D3
(DSM, Germany) (IU) 2,000 5,000 10,000 Vitamin B12
(Lycored) (mg) One One One Vitamin B6
(DSM, Germany) (mg) 60 60 60 Vitamin B9
(DSM, Switzerland) (mg) 0.4 0.4 0.4 Hydrogen stearate choline
(Algry) (mg) 50 50 50 Vitamin E
(DSM, Switzerland) (IU) 100 100 100 Vitamin C
(DSM, UK) (mg) 100 100 100 Silymarin
(Teva) (mg) 420 420 420 Biphenyldimethyldicarboxylate
(Zhejiang Hisoar) (mg) 150 150 150

The A1-A3 and B1-B3 compositions thus prepared were weighed according to the dosages in Table 5 and dissolved in sterile water for injection of 0.5% carboxymethylcellulose (CMC) to prepare dosage compositions, which were then refrigerated. In the case of the above dose, 10 times the dose based on mg per kg (kg) of the animal without conversion of the body surface area was set at the dosage as follows.

A1 A2 A3 B1 B2 B3 Dose
(mg / head / day) 152.9 154.7 157.7 235.1 276.6 315.5 Dosage amount
(ml / head / day) 3 3 3 3 3 3

Experimental Example 1: Single tablet elution test

A dissolution test of the single tablet prepared according to Example 1 was carried out. Vitamin B6 (pyridoxine hydrochloride) and vitamin D (cholecalciferol) were used as indicators for the dissolution test in water.

[Vitamin B6 (pyridoxine) test method]

1) Dissolution test basis: In the 11th revision of Korea Pharmacopoeia, the dissolution test method of General Test Methods

2) Elution test method: Paddle method, 50 revolutions per minute

3) Elution test liquid: Water

4) Analysis method: HPLC method

- Detector: Ultraviolet absorption spectrophotometer (detection wavelength = maximum 270 nm)

- Column: Column with a diameter of about 4.6 mm and a length of about 25 cm and filled with porous silica particles of 5 to 10 μm in diameter

- Mobile phase: Add 0.4 mL of triethylamine, 15.0 mL of acetic acid (100), and 350 mL of methanol into a 2000 mL volumetric flask, add 0.008 mol / L sodium hexanesulfonate to 2000 mL,

[Vitamin D (cholecalciferol) test method]

1) Dissolution test basis: In the 11th revision of Korea Pharmacopoeia, the dissolution test method of General Test Methods

2) Elution test method: Paddle method, 50 revolutions per minute

3) Elution test liquid: Water

4) Analysis method: HPLC method

- Detector: Ultraviolet absorptiometer (detection wavelength = max 265nm)

- Column: Column packed with aminopropylsilyl porous silica gel for liquid chromatography of 10 μm in stainless steel pipe with inner diameter of about 4.6 mm and length of about 15 cm

- mobile phase: n-hexane isopropanol (99: 1)

As a result, as shown in Fig. 1, the dissolution rate of vitamin B6 was 95% at 30 minutes and that of vitamin D3 was 83% at 30 minutes.

Experimental Example  2: side effects due to the administration of a single composition Occurrence  And the damage between the damaged

1) administration of single composition and induction of liver damage

Animals that had undergone a one-week purification period as in Reference Example 2 were divided into groups as shown in Table 6, and the single compositions A1-A3 and B1-B3 prepared according to Example 5 were orally administered (G3-G8) . The dose was administered once / day (7 times / week) for 42 days. All doses were performed before 15:00 and autopsy was performed after 6 weeks. The weight of the animals determined to be healthy during the refinement period was ranked and randomly distributed so that the average weight of each group was evenly distributed.

The excipient to be administered to the G1 and G2 groups was sterile injectable water of 0.5% carboxymethylcellulose (CMC) used for preparing the composition in Example 5.

On the other hand, 200 mg / kg of D-galactosamine was intraperitoneally administered to all but the G1 group at 1 hour after administration of the first single composition to induce liver damage in the animal model.

group gender Number of animals Animal number Dosage amount
(mL / head / day) Dosing substance G1 cock 6 1-6 3 Excipient G2 cock 6 7-12 3 Excipient G3 cock 6 13-18 3 A1 G4 cock 6 19-24 3 A2 G5 cock 6 25-30 3 A3 G6 cock 6 31-36 3 B1 G7 cock 6 37-42 3 B2 G8 cock 6 43-48 3 B3

(G1: normal group, G2: vehicle control group, G3-8: single composition administration group)

2) Confirmation of occurrence of side effects by administration of single composition

To determine whether a single composition exhibited adverse effects on animal models, the presence or absence of abnormal symptoms, the date of onset, the severity of symptoms, and the mortality during the observation period were observed once a day when the single composition was administered. The first single composition administration day was set to Day 1, and autopsy was performed after 6 weeks of observation.

Weights of the animal models were measured at the time of arrival, at the time of separation, once / week, and at the date of the autopsy, and the weights of the liver, heart and kidney were measured after completion of blood collection at the date of autopsy.

Calcium ion (Ca ^{2 +} ) levels were measured by autopsy and blood biochemistry.

As a result of observing general symptoms during the test period, no animals died from the first administration day of the single composition until the day of the autopsy, and no animals exhibiting abnormal symptoms due to administration of the single composition were observed (Table 7).

Work Symptom group G1 G2 G3 G4 G5 G6 G7 G8 1-43 normal 6/6 6/6 6/6 6/6 6/6 6/6 6/6 6/6 43 Autopsy 6/6 6/6 6/6 6/6 6/6 6/6 6/6 6/6

Body weights of animal models during the test period were not significantly different in all groups (Table 8, Figure 2)

G1 G2 G3 G4 G5 G6 G7 G8 0 weeks 298.91 + - 8.54 300.66 + - 9.85 299.90 + - 8.74 301.39 ± 9.14 299.97 ± 5.97 298.93 + - 7.46 299.58 +/- 7.53 302.73 + - 5.43 1 week 348.76 + - 11.64 345.84 + - 9.98 345.23 + - 9.83 347.30 ± 15.26 344.12 + - 9.14 345.89 ± 18.23 349.36 ± 14.22 348.40 ± 8.64 2 weeks 386.73 + - 13.11 384.61 + - 10.15 384.17 + - 11.75 387.03 + - 20.19 389.56 ± 17.21 387.45 +/- 29.18 395.42 + - 24.18 381.68 + - 21.84 3 weeks 415.99 ± 9.94 412.34 + - 13.14 407.75 ± 12.88 418.66 ± 25.34 415.91 + - 23.98 411.55 + 33.54 420.50 ± 30.34 405.01 + - 20.55 4 weeks 445.82 + - 13.14 442.05 ± 15.29 431.18 ± 18.69 447.16 +/- 29.81 440.16 + 26.85 434.87 +/- 40.84 447.54 +/- 37.40 428.65 + - 24.44 5 weeks 468.66 ± 13.26 467.23 + - 18.65 451.16 + - 18.78 471.93 + - 31.67 464.22 + - 33.19 458.40 ± 49.08 475.01 + - 40.03 450.59 ± 26.25 6 weeks 487.67 ± 16.14 485.19 + - 24.52 469.57 ± 22.28 492.49 + - 34.82 481.72 + - 34.55 478.94 + - 53.61 493.57 ± 45.47 468.79 ± 29.51 Autopsy 459.25 + 15.91 457.74 ± 23.87 445.85 + - 22.36 463.70 ± 32.35 451.04 + - 33.80 448.76 ± 51.64 467.04 + - 43.11 444.49 ± 28.62

As a result of measuring the weight of the liver after the completion of the test, no significant difference was observed between the single composition administration groups and the excipient control group (FIG. 3 and Table 9) (Fig. 4, Table 9). As a result of measuring the weight of the heart, it was confirmed that there was no significant difference in all groups (Fig. 5 and Table 9).

Weight of organ (g) liver Height (left) Height (right) Heart G1 13.265 ± 1.277 ^** 1.652 + 0.142 1.692 + 0.122 1.552 + 0.086 G2 16.870 + 1.795 1.665 + 0.082 1.666 + 0.109 1.434 + 0.088 G3 15.545 ± 1.249 1.623 + 0.129 1.650 + 0.135 1.442 + 0.084 G4 17.185 ± 3.312 1.772 + 0.130 1.741 + 0.091 1.499 + 0.123 G5 16.194 ± 2.283 1.644 + 0.107 1.615 + 0.075 1.494 + 0.158 G6 16.156 ± 3.030 1.715 + 0.125 1.730 + 0.187 1.515 + 0.110 G7 16.927 + 1.450 1.767 + 0.155 1.814 + 0.177 1.522 + 0.165 G8 15.429 ± 2.228 1.873 + - 0.185 ^* 1.876 + - 0.236 ^* 1.441 + 0.180

(* And ** mean statistical significance with G2, representing P <0.05 and P <0.01, respectively)

Also, as a result of the measurement of the calcium ion level, it was confirmed that all the single composition administration group did not show any significant difference compared with the excipient control group (Table 10).

G1 G2 G3 G4 G5 G6 G7 G8 Calcium ion
(mg / dL) 10.02 0.08 9.92 + - 0.12 10.17 ± 0.16 10.25 + 0.09 10.02 ± 0.16 10.09 + - 0.11 10.17 ± 0.19 10.25 + - 0.10

Thus, it was confirmed that the single composition does not cause side effects such as hepatotoxicity even when administered in the body, and is a safe substance that does not cause toxic action even when containing a high dose of vitamin D in particular.

3) Confirmation of improvement of surface element

(AST), alanine aminotransferase (ALT), and blood urea nitrogen (BUN), which are components that increase expression levels in liver damage, are measured And the antioxidant activity (SOD activity) was measured in order to confirm the recovery effect of liver function.

Specifically, D-galactosamine was administered, followed by fasting for 24 hours. Jugular vein sampling was performed, and serum was separated using a vacutainer tube containing a clot activator. Blood biochemical tests for AST, ALT, and BUN were performed on the day of the autopsy.

In addition, after autopsy, some tissues were cut and the tissue was disrupted, and the SOD activity was measured using an SOD kit.

As a result of the AST measurement, it was confirmed that the AST level was increased in the excipient control group in which hepatic injury was induced compared to the normal control group. On the other hand, the single composition administration groups were found to have decreased AST levels compared with the vehicle control group. Specifically, the single composition A1-A3 treated groups (G3-G5) showed about 74.18, 80.62 and 77.90% improvement compared to the vehicle control group, and the single composition B1-B3 treated groups (G6-G8) , It was confirmed that the improvement rate was about 84.98, 77.42, and 89.34% (FIG. 6 and Table 11).

As a result of the ALT measurement, it was confirmed that the ALT level was increased in the excipient control group in which hepatic injury was induced compared to the normal group. On the other hand, the single composition administration groups were found to have decreased ALT levels compared with the vehicle control group. Specifically, the single composition A1-A3 administration groups (G3-G5) showed improvement rates of about 77.40, 82.90 and 82.15% compared to the vehicle control group and the single composition B1-B3 administration groups (G6-G8) , It was confirmed that the improvement rate was about 90.35, 88.35, and 92.95% (FIG. 7 and Table 11).

As a result of BUN measurement, it was confirmed that the BUN value of the excipient control group in which hepatic injury was induced was increased as compared with the normal group. On the other hand, the single composition administration groups were found to have decreased BUN levels compared to the vehicle control group (Table 11).

In addition, SOD measurement showed that SOD was decreased in the excipient control group induced liver damage compared with the normal group. On the other hand, it was confirmed that the single composition administration groups had an increase in SOD as compared with the vehicle control group (Fig. 8 and Table 11).

AST (U / L) ALT (U / L) BUN (mg / dL) SOD activity (%) G1 172.60 ± 6.19 ^* 34.55 ± 2.13 ^* 12.02 + - 0.25 ^* 52.90 + - 5.39 ^* G2 2936.42 + - 770.25 2000.60 ± 526.22 14.40 0.95 41.31 8.31 G3 758.42 + - 315.79 ^* 452.68 ± 203.78 ^* 12.68 ± 1.01 57.91 + - 2.32 ^** G4 569.17 ± 112.02 ^* 342.00 ± 132.52 ^* 11.80 ± 0.59 63.32 + - 6.55 ^** G5 649.86 + - 216.02 ^* 357.10 + - 141.14 ^* 13.82 ± 1.68 62.46 + - 6.45 ^** G6 441.22 + ^- 121.11 ^* 193.34 ± 70.66 ^* 11.44 + - 0.58 ^* 57.20 ± 5.66 ^** G7 663.15 ± 410.18 ^* 293.63 + ^- 217.14 ^* 11.02 + - 0.22 ^** 51.22 + - 8.13 G8 313.52 + - 158.91 ^* 141.93 + - 101.30 ^* 12.12 + - 0.44 49.17 ± 3.47

(* And ** mean statistical significance with G2, representing P <0.05 and P <0.01, respectively)

Through this, it was confirmed that the single composition showed an improvement effect on liver damage and a recovery effect of liver function.

Claims (15)

Vitamin D, vitamin B12, vitamin B6, vitamin B9 and choline. The health functional food composition according to claim 1, further comprising at least one selected from vitamin E, vitamin C, milk syrup extract and omija extract. The health functional food composition according to claim 1 or 2, wherein the vitamin D is vitamin D3. The health functional food composition according to claim 3, wherein the vitamin D3 is contained in an amount of 1,000 IU to 20,000 IU. The health functional food composition according to claim 3, wherein the vitamin D3 is contained in an amount of 2,000 IU to 20,000 IU. The health functional food composition according to claim 3, wherein the vitamin D3 is contained in an amount of 5,000 IU to 20,000 IU. The health functional food composition according to claim 3, wherein the vitamin D3 is contained in an amount of 2,000 IU to 10,000 IU. Vitamin D, vitamin B12, vitamin B6, vitamin B9 and choline. The health functional food composition according to claim 8, which further comprises at least one selected from vitamin E, vitamin C, milk syrup extract, and omija extract. The health functional food composition according to claim 8 or 9, wherein the vitamin D is vitamin D3. The health functional food composition according to claim 10, wherein the vitamin D3 is contained in an amount of 1,000 IU to 20,000 IU. The health functional food composition according to claim 10, wherein the vitamin D3 is contained in an amount of 2,000 IU to 20,000 IU. The health functional food composition according to claim 10, wherein the vitamin D3 is contained in an amount of 5,000 IU to 20,000 IU. The health functional food composition according to claim 10, wherein the vitamin D3 is contained in an amount of 2,000 IU to 10,000 IU. The health functional food composition according to claim 8 or 9, wherein the liver disease is any one of fatty liver, hepatitis, liver cirrhosis and liver cancer.

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