WO2020138557A1 - Pharmaceutical composition and health functional food for preventing or treating fatty liver - Google Patents

Abstract

The high-fat diet causes non-alcoholic fatty liver, and due to the fatty liver, excessive mitophagy occurs, and thus mitochondrial function is lost. Accordingly, hepatocytes receive oxidative stress, causing cell death, and liver damage occurs. A pharmaceutical composition comprising alpha-lipoic acid or a pharmaceutically acceptable salt thereof according to the present invention inhibits same, and thus can prevent or treat fatty liver and liver damage, and a health functional food comprising alpha-lipoic acid or a sitologically acceptable salt thereof according to the present invention can prevent or ameliorate fatty liver and liver damage.

Description

Pharmaceutical composition and health functional food for prevention or treatment of fatty liver

The present invention relates to a pharmaceutical composition and a health functional food for preventing or treating fatty liver.

The liver is one of the most active organs in the body that metabolizes in vivo, and is a very important organ responsible for various metabolism, detoxification, decomposition, synthesis, and secretion. Specifically, all nutrients absorbed from food are metabolized as substances capable of producing energy in the liver and supplied or stored in the body, and about 2,000 enzymes, albumin, serum proteins of coagulation factors, bile acids, phospholipids, cholesterol, and other fats The back is synthesized, stored and distributed in the liver. In addition, as the detoxification and decomposition functions, drugs, alcohol, and toxic substances are detoxified in the liver, so liver cells are easily damaged by these substances, and thus there is a high possibility of liver disease caused by drugs, alcohol, or toxicity.

The liver is one of the organs with excellent regenerative capacity, but if the damage persists, recovery of the liver to a normal liver is difficult due to complete destruction of liver tissue and deterioration of liver function. When the liver damage becomes chronic, the probability of progression to liver fibrosis, cirrhosis, or liver cancer increases regardless of the cause. Therefore, treating such liver damage before liver damage becomes chronic is very important in inhibiting liver fibrosis, cirrhosis or progression to liver cancer.

Among them, the fatty liver is unable to metabolize normal fat due to causes such as excessive intake of fat, increased accumulation and synthesis in the liver, and decreased discharge, resulting in excessive accumulation of lipids, especially triglycerides, and the proportion of fat in the liver is 5%. It means the case exceeding. Recently, as the nutritional status improves and the number of adult diseases increases, the number of patients with fatty liver increases.

Alcohol and obesity are the main causes of fatty liver, and it may be accompanied by diseases such as hyperlipidemia and diabetes, which have high levels of lipids in the blood, and drugs such as adrenal cortical hormones (steroids) and female hormones may also be the cause. have. In addition, fatty liver can be caused by severe malnutrition. Symptoms of fatty liver may vary depending on the degree of accumulation and duration of fat, and the presence or absence of other diseases.

On the other hand, in the case of a normal liver, as shown in Figure 1, in the presence of abnormal mitochondria mitophagy to remove the damaged mitochondria. However, in the case of fatty liver, the mitophagy system does not work, resulting in liver damage. That is, mitophagy should be properly activated, but mitophagy is reduced/inhibited/inactive, but it is not good to be excessively active.

To date, polyenphosphatidylcholine has been used clinically as a therapeutic agent for fatty liver, and it is known that fibrate-based drugs and statin-based drugs represented by clofibrate, which are hyperlipidemic agents, have an effect on fatty liver. However, fibrate-based drugs are known to improve lipid metabolism, such as increasing high-density lipoprotein cholesterol through fatty liver beta-oxidation enzymes, but on the other hand, they cause side effects that cause liver dysfunction, so they have excellent therapeutic effects and side effects There is a need to develop a therapeutic agent for the prevention and treatment of new fatty liver that does not cause problems such as or resistance.

In addition, mitophagy is a high-fat diet involved in the formation of fatty liver, and by inhibiting excessive mitophagy, there is a need to develop a therapeutic substance that can protect liver damage caused by high-fat diet.

An object of the present invention is to provide a pharmaceutical composition for preventing or treating fatty liver, comprising alpha lipoic acid or a pharmaceutically acceptable salt thereof.

In addition, an object of the present invention is to provide a health functional food for preventing or improving fatty liver containing alpha lipoic acid or a food-acceptable salt thereof.

1. Alpha-lipoic acid (α-Lipoic Acid) or a pharmaceutical composition for preventing or treating fatty liver comprising a pharmaceutically acceptable salt thereof.

2. In the above 1, wherein the fatty liver is a non-alcoholic fatty liver (Non-Alcoholic Fatty Liver Disease, NAFLD), pharmaceutical composition.

3. In the above 1, wherein the composition is to prevent or treat liver damage caused by fatty liver, pharmaceutical composition.

4. Alpha-lipoic acid (α-Lipoic Acid) or a health functional food for preventing or improving fatty liver containing a food-acceptable salt thereof.

5. In the above 4, the fatty liver is a non-alcoholic fatty liver (Non-Alcoholic Fatty Liver Disease, NAFLD), health functional food.

6. In the above 4, the health functional food is to prevent or improve liver damage caused by fatty liver, health functional food.

High-fat diets cause non-alcoholic fatty liver, and excessive mitophagy from fatty liver is created, resulting in loss of mitochondrial function. As a result, hepatocytes undergo oxidative stress and cause apoptosis, and liver damage occurs. The pharmaceutical composition of the present invention can prevent or treat fatty liver and liver damage by inhibiting it, and the health functional food of the present invention can prevent or improve fatty liver and liver damage.

1 is a diagram showing liver damage due to mitophagy function in normal liver and mitophagy overactivity in fatty liver.

Figure 2 is a control (control), high-fat diet group (HF), high-fat diet + alfaic acid treatment group (HF + ALA) is a diagram showing the weight, liver weight, fat accumulation in the body.

3 is a result of staining the fat in the liver tissue with Oil Red O.

4 is a diagram showing abnormal liver tissue through H&E staining.

5 is a diagram showing the fibrosis around the blood vessels in the liver tissue through MT staining.

6 is a view showing the degree of oxidative stress in the liver through 8-OHdG staining.

7 is a diagram showing apoptosis in liver tissue through TUNEL staining.

8 is a view showing the expression of mitophagy-related factors responsible for the functional abnormality of the organelle mitochondria in cells in adipose tissue.

9 is a diagram confirming the expression levels of Parkin, Pink1, VDAC1, and b-actin in mitochondria.

10 is a view showing the results of confirming the expression of Parkin with an electron microscope.

11 is a view showing the results of confirming the expression of Pink1 with an electron microscope.

Hereinafter, the present invention will be described in detail.

The present invention relates to a pharmaceutical composition for the prevention or treatment of fatty liver comprising alpha-Lipoic Acid.

The alpha lipoic acid is a compound represented by the following Chemical Formula 1, the chemical formula is C ₈ H ₁₄ O ₂ S ₂ , the molar mass is 206.33 g/mol, and the name of IUPAC is 5-(1,2-dithiolan-3-yl)pentanoic acid to be:

[Formula 1]

.

The alpha lipoic acid may be derived from nature, or may be synthesized using a known organic synthetic method.

The alpha lipoic acid may be a product obtained by culturing non-protein compounds, peptides, plant-derived tissue or cell extracts, and microorganisms (eg, bacteria or fungi, and especially yeast).

The pharmaceutical composition according to the present invention may include an active ingredient alone or may be provided as a pharmaceutical composition including one or more pharmaceutically acceptable carriers, excipients, or diluents.

"Pharmaceutically acceptable" in the present invention means that it exhibits properties that are not toxic to cells or humans exposed to the composition.

The term "pharmaceutically acceptable salt" of the present invention means a salt prepared by using a specific compound according to the present invention and a relatively non-toxic acid or base. When the compounds of the present invention contain relatively acidic functional groups, base addition salts can be obtained by contacting a sufficient amount of base in a neutral form of such a compound in a pure solution or a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amines, or salts of magnesium or similar salts. When the compounds of the present invention contain relatively basic functional groups, acid addition salts can be obtained by contacting a sufficient amount of acid in a neat solution or in a suitable inert solvent with the neutral form of these compounds. Pharmaceutically acceptable acid addition salts are salts of inorganic acids such as hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, hydrogen carbonate ion, phosphoric acid, monohydrogen phosphate, dihydrogen phosphate, sulfuric acid, hydrogen sulfate ion, hydroiodic acid or phosphorous acid, And salts of organic acids such as acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-tolylsulfonic acid, citric acid, tartaric acid, methanesulfonic acid Examples thereof include salts of amino acids (for example, arginine) and salts of organic acids such as glucuronic acid.

The pharmaceutically acceptable salt of the present invention can be synthesized by a conventional chemical method from a parent compound containing an acidic or basic moiety. Generally, these salts are prepared by reacting the form of the free acid or base of these compounds with a stoichiometrically appropriate base or acid in water or in an organic solvent or in a mixture of the two. In general, non-aqueous media such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile are preferred.

Furthermore, the pharmaceutical composition of the present invention may be provided by mixing with a composition for preventing or treating fatty liver known in the art. That is, the pharmaceutical composition of the present invention can be administered in parallel with a known compound having a prophylactic or therapeutic effect on fatty liver.

In the present invention, "administration" refers to the introduction of a predetermined substance to an individual by an appropriate method, and "individual" refers to all living organisms such as rats, mice, livestock, etc., including humans capable of retaining fatty liver. As a specific example, it may be a mammal, including a human.

If necessary, the pharmaceutical composition of the present invention may further include a composition having a known fatty liver prevention or treatment effect.

As a composition for preventing or treating fatty liver, a composite of extract of yangha and vitamin tree leaf extract, nalmefen and naltrexon or derivatives thereof, 1',4-dihydroxy-3,3',5'-trimethoxy- 7'8'9'-triol-8,4'-oxyneorigna-7,9-diol, curcumin derivatives, fat extracts of Guamegi, and the like, but are not limited thereto.

In the present invention, the route of administration of the pharmaceutical composition is, but is not limited to, oral, intravenous, intramuscular, intraarterial, intramedullary, intrathecal, intracardiac, transdermal, subcutaneous, intraperitoneal, intranasal, intestinal, Topical, sublingual or rectal.

The composition of the present invention may be administered orally or parenterally, and when parenterally administered, it is preferable to select an external injection or intraperitoneal injection, intrarectal injection, subcutaneous injection, intravenous injection, intramuscular injection, or intrathoracic injection injection method. , But is not limited thereto.

The preferred dosage of the composition of the present invention depends on the patient's condition and body weight, the degree of disease, the drug form, the route and duration of administration, but can be appropriately selected by those skilled in the art. However, for the desired effect, the composition is preferably administered at 0.01 to 1000 mg/kg/day, preferably 0.1 to 500 mg/kg/day, but is not limited thereto. The 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.

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 include at least one excipient in the extract, for example, starch, calcium carbonate, sucrose. Or it is prepared by mixing 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 commonly used simple 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 suppository base, witepsol, macrogol, tween 61, cacao butter, laurin butter, and glycerogelatin may be used.

The fatty liver may be, for example, non-alcoholic fatty liver (NAFLD) or alcoholic fatty liver (Alcoholic Fatty Liver Disease, AFLD), preferably non-alcoholic fatty liver, but is not limited thereto. .

The alcoholic fatty liver may be caused by drinking alcohol, and the non-alcoholic fatty liver may be caused by obesity due to a high-fat diet, but is not limited thereto.

The composition may be capable of preventing or treating liver damage caused by fatty liver. In the case of non-alcoholic fatty liver, mitophagy becomes overactive, and thus, the function of mitochondria is lost, and accordingly, the liver cells may be damaged by oxidative stress and apoptosis, thereby preventing or treating fatty liver by inhibiting the alpha lipoic acid. The effect can be achieved, but is not limited thereto.

Specifically, the alpha lipoic acid can reduce body fat, fat accumulation in the body, and more specifically, reduce the expression of the fat synthesis inducer gene Dgat2 in adipose tissue, and increase the expression of the fat synthesis inhibitory gene Pnpla 2 However, it is not limited thereto.

In addition, the alpha lipoic acid may reduce blood sugar levels, reduce oxidative stress and apoptosis of liver tissue, more specifically restore abnormal liver tissue, and reduce fibrosis around blood vessels in liver tissue. However, it is not limited thereto.

In addition, the alpha-lipoic acid may decrease the activity of mitophages in liver tissue, and more specifically, may reduce the expression of the mitophages markers Ambra1, PINK, Parkin, BNIP3, and Parkin, Pink1, VDAC1 in mitochondria. , It is possible to reduce the expression level of b-actin, but is not limited thereto.

In addition, the present invention relates to a health functional food for preventing or improving fatty liver containing alpha-Lipoic Acid.

The fatty liver may be alcoholic fatty liver and non-alcoholic fatty liver as described above, preferably non-alcoholic fatty liver, but is not limited thereto.

The efficacy and mechanism of action of the alpha lipoic acid may be within the aforementioned range.

In the present invention, "food-tolerant" means that it exhibits non-toxic properties to cells or humans exposed to the compound.

In the present invention, "food-acceptable salt" means a salt prepared by using a specific compound according to the present invention and a relatively non-toxic acid or base, and may be within the above-mentioned range for "salt".

The health functional food of the present invention may be formulated as one selected from the group consisting of tablets, pills, powders, granules, powders, capsules, and liquid formulations, further comprising one or more of carriers, diluents, excipients, and additives. Foods to which the extract of the present invention can be added include various foods, powders, granules, tablets, capsules, syrups, drinks, gums, teas, vitamin complexes, and health functional foods.

Additives that may be further included in the present invention include natural carbohydrates, flavoring agents, nutrients, vitamins, minerals (electrolytes), flavoring agents (synthetic flavoring agents, natural flavoring agents, etc.), coloring agents, fillers (cheese, chocolate, etc.), Factic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusting agents, stabilizers, preservatives, antioxidants, glycerin, alcohols, carbonizing agents and one or more components selected from the group consisting of flesh can be used. .

Examples of the natural carbohydrates described above include monosaccharides, such as glucose, fructose, and the like; Disaccharides such as maltose, sucrose, etc.; And polysaccharides, for example, conventional sugars such as dextrin, cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As the flavoring agent, natural flavoring agents (taumatine, stevia extract (for example, rebaudioside A, glycyrrhizine, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used.

In addition to the above, the health functional food of the present invention includes various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and neutralizing agents (cheese, chocolate, etc.), pectic acid and salts thereof, alginic acid And salts, organic acids, protective colloidal thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonic acid used in carbonated beverages, and the like. In addition, the composition according to the present invention may contain natural fruit juice and pulp for the production of vegetable beverages. These ingredients can be used independently or in combination.

Specific examples of the carrier, excipients, diluents and additives include, but are not limited to, lactose, dextrose, sucrose, sorbitol, mannitol, erythritol, starch, acacia rubber, calcium phosphate, alginate, gelatin, calcium phosphate, calcium Silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, polyvinylpyrrolidone, methylcellulose, water, sugar syrup, methylcellulose, methyl hydroxy benzoate, propylhydroxy benzoate, talc, magnesium stearate And it is preferred that one or more selected from the group consisting of mineral oil is used.

When formulating the health functional food of the present invention, it is prepared using diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, surfactants, etc., which are usually used.

Hereinafter, examples will be described in detail to specifically describe the present invention.

Example

1. Experimental method

(1) Experimental rat and control rat

3 weeks old C57BL/6J male rats were prepared and treated with a high fat diet (HFD) for 4 to 10 weeks to treat the test drug (alphalipoic acid, α-Lipoic Acid, ALA) and to compare with the control group. The experiment was conducted. A total of 10 C57BL/6 rats were fed a high-fat diet, and a total of 10 C57BL/6 rats were fed a high-fat diet and alphalipoic acid was administered at 200 mg/kg/day for 10 weeks. In addition, a total of 5 C57BL/6 mice were subjected to a normal diet (ND).

2. Experimental results

(1) Effects on body weight, epididymal fat weight, and liver tissue weight by alpha lipoic acid-Fatty liver reduction effect

The alpha-lipoic acid-treated group had less weight and less epididymal fat than the control group, but there was no difference in dietary amount between the control and alpha-lipoic acid-treated groups.

The fat synthesis-inducing gene Dgat2 in adipose tissue, an enzyme related to fattening, was decreased in alpha lipoic acid compared to the control group, and the fat synthesis inhibitory gene Pnpla 2 showed an effect of increasing expression (FIG. 2).

2, it can be seen that the treatment with alpha lipoic acid decreases body weight (left) and body fat (epididymal fat) accumulation (right) (statistically significant). However, the weight (middle) of liver tissue was not statistically significant in the alpha-lipoic acid treatment group, but increased.

In the alpha-lipoic acid treatment group, a reduction effect of fatty liver formation was observed compared to the control group, which was confirmed by Oil Red O fat staining (FIG. 3). 3, it was found that the fat in the liver tissue increased by the high-fat diet was reduced by treatment with alpha lipoic acid.

(2) Comparison of blood sugar levels

Blood sugar was significantly lower in the alpha-lipoic acid treatment group than in the control group.

(3) Effects on oxidative stress and apoptosis-suppressing liver tissue damage

As a result of H&E staining, it was confirmed that the appearance of abnormal liver tissue was normalized by alpha lipoic acid treatment with a high fat diet (FIG. 4 ).

In addition, as a result of MT staining, it was confirmed that fibrosis around the blood vessels in the liver tissue increased by a high fat diet was reduced by treatment with alpha lipoic acid (FIG. 5 ).

In the oxidative stress, the alpha lipoic acid treatment group showed a tendency to decrease compared to the control group, which was confirmed through 8-OHdG staining (FIG. 6 ).

In addition, apoptosis in the liver tissue increased by the high-fat diet showed a tendency to decrease in the alpha lipoic acid treated group compared to the control group, which was confirmed through TUNEL staining (FIG. 7 ).

(5) Mitochondrial damage reduction effect-Inhibit mitophagy activity

As a result of Western blot, it was confirmed that mitophagy activity of liver tissue increased by a high fat diet was reduced by treatment with alpha lipoic acid.

The expression of the mitophagy-related factors (mitophagy markers), Ambra1, PINK, Parkin, and BNIP3, which are responsible for functional abnormalities of the organelle mitochondria in adipose tissue, was reduced in the alphalipoic acid treatment group compared to the control group (FIG. 8).

In addition, as a result of confirming the expression levels of Parkin, Pink1, VDAC1, and b-actin in the mitochondria, it was confirmed that the decrease in the alphalipoic acid treatment group (FIG. 9).

Actually, the expression of Parkin was confirmed by optical microscopy after performing immunohistochemistry, and FIG. 11 shows the result of pink1 expression confirmed by optical microscopy after immunohistochemistry.

Claims (6)

1. A pharmaceutical composition for preventing or treating fatty liver, comprising alpha-Lipoic Acid or a pharmaceutically acceptable salt thereof.
2. The pharmaceutical composition of claim 1, wherein the fatty liver is Non-Alcoholic Fatty Liver Disease (NAFLD).
3. The method according to claim 1, wherein the composition is to prevent or treat liver damage caused by fatty liver, pharmaceutical composition.
4. Alpha-lipoic acid (α-Lipoic Acid) or a health functional food for preventing or improving fatty liver containing a food-acceptable salt thereof.
5. The method according to claim 4, The fatty liver is a non-alcoholic fatty liver (Non-Alcoholic Fatty Liver Disease, NAFLD), health functional food.
6. The method according to claim 4, The health functional food is to prevent or improve liver damage caused by fatty liver, health functional food.

Images