KR20180080834A - Composition for antiinflammatory and inflammatory neurodegenerative diseases comprising hizikia fusiforme extract - Google Patents

Abstract

The present invention relates to a composition for preventing or treating antiinflammatory and inflammatory neurodegenerative diseases comprising a Sargassum fusiforme extract.

Description

Technical Field [0001] The present invention relates to a composition for preventing or treating inflammatory and inflammatory neurodegenerative diseases, which comprises an extract of Chamaecyparis obtusa,

The present invention relates to a composition for preventing or treating anti-inflammatory and inflammatory neurodegenerative diseases, which comprises an extract of Chrysanthemum morifolium.

In addition to kelp, Hizikia fusiformis is a neutral polysaccharide known to have functionalities such as amino acids (glutamic acid and aspartic acid), blood coagulation action, and immunity enhancing action, and laminaran, It is known to contain a lot of fucoidan. Studies on the physiological activity of Ttot have been reported mainly on the chemical characteristics of Fucoidan extracted from Tart, the antimicrobial activity of Tartori extracts, and the effect of Tartar extract on lipid metabolism and hepatic enzymatic activity, Antihyperlipidemic, anti-cholesterol and antioxidant effects have been studied.

On the other hand, as the age of the aging society increases, the development of degenerative brain diseases such as Alzheimer's disease, Parkinson's disease, and ischemic brain diseases is greatly increased. Therefore, many researches are being carried out in the world on the development of therapies and medicines have.

In general, neuroinflammation is accompanied by the onset of various neurological and neurodegenerative diseases such as cerebral ischemia, Alzheimer's disease, Parkinson's disease, Huntington's disease and amyotrophic lateral sclerosis. Microglia, a unique immune cell in the brain, play a particularly important role in brain inflammation through the production of proinflammatory cytokines, nitric oxide (NO) and other neurotoxic factors. The neuron neurons can be directly damaged by activated cytoskeletal stimulation of tumor necrosis factor (TNF-a) from microglial cells.

Therefore, inhibition of cell signaling by microglial cells may lead to improvement of neuroinflammatory diseases.

Lipopolysaccharide (LPS) has been widely used as an active model immunostimulant in vivo and in vitro studies. Previous studies have shown that LPS stimulates microglia to induce cytokines such as inducible nitric oxide synthase (iNOS), NO and interleukin (IL) -1, IL-6 and tumor necrosis factor (TNF-a) It was proposed to produce.

Excessive production of NO has harmful effects on the body, resulting in cell damage, and is an important cause of degenerative diseases such as inflammation, meningitis, Alzheimer's disease and Parkinson's disease. TNF- Or chronic inflammatory diseases, and their regulation is recognized as an important goal in the treatment of inflammatory diseases.

Korean Patent Laid-Open No. 10-2012-0011981 relates to a dermal extract having anti-inflammatory effect and a composition for external application for skin comprising the same, wherein the dermal extract is characterized in that the expression of hemeoxygenase-1 and sirtuin-1 And has an anti-inflammatory effect.

Korean Patent Publication No. 10-2012-0011981

The present invention aims to provide a composition for treating or preventing an inflammatory neurodegenerative disease.

The present invention also aims to provide an anti-inflammatory composition.

In order to achieve the above object,

The present invention provides a pharmaceutical composition for the prevention or treatment of inflammatory neurodegenerative diseases comprising an extract of Chrysanthemum morifolium as an active ingredient.

In addition, the present invention provides a food composition for relieving inflammatory neurodegenerative diseases, comprising an extract of Chrysanthemum morifolium as an active ingredient.

The present invention also provides a pharmaceutical composition for antiinflammation comprising an extract of Chrysanthemum morifolium as an active ingredient.

In addition, the present invention provides a functional food composition for anti-inflammation, comprising an extract of Chrysanthemum morifolium as an active ingredient.

The present invention also provides a composition for antioxidation comprising an extract of Chrysanthemum morifolium as an active ingredient.

The present invention also provides a method for inhibiting the concentration of nitrogen monoxide produced in glial cells in vitro by treating an alcohol extract of the extract from a plant with a lipopolysaccharide-stimulated glial cell.

The composition comprising the extract of the present invention as an active ingredient can completely weaken the production of LPS-induced inducible nitric oxide synthase (iNOS) and nitrogen monoxide (NO) in BV-2 cells.

Accordingly, the composition comprising the extract of the present invention as an active ingredient can be used as an anti-inflammatory agent and a therapeutic agent for neurodegenerative diseases.

Fig. 1 is a graph showing the antioxidative effect of the extract of Chrysanthemum morifolium.
FIG. 2 is a graph showing the cell survival rate of the extract of Chrysanthemum morifolium.
FIG. 3 is a graph showing inhibition of nitrogen monoxide (NO) production of the extract from the extract.
FIG. 4 is a graph showing the efficacy against inflammatory proteins caused by the extracts of Chrysanthemum morifolium.

Hereinafter, the present invention will be described in more detail.

The present invention relates to a pharmaceutical composition for preventing or treating an inflammatory neurodegenerative disease comprising an extract of Chrysanthemon hull extract as an active ingredient.

In one embodiment of the present invention, the composition desirably reduces the production of inductive nitrogen oxide synthase (iNOS) and nitrogen monoxide (NO), but is not limited thereto.

In another embodiment of the present invention, the extraction solvent of the extract is preferably an alcohol, and most preferably, ethanol is not limited thereto.

In another embodiment of the present invention, the composition preferably inhibits microglial cell activity in microglial cells, but is not limited thereto.

Further, the present invention relates to a pharmaceutical composition for antiinflammation comprising an extract of Chrysanthemum morifolium as an active ingredient.

In one embodiment of the present invention, the composition desirably reduces the production of inductive nitrogen oxide synthase (iNOS) and nitrogen monoxide (NO), but is not limited thereto.

In another embodiment of the present invention, the extraction solvent of the extract is preferably an alcohol, and most preferably, ethanol is not limited thereto.

In another embodiment of the present invention, the composition preferably inhibits microglial cell activity in microglial cells, but is not limited thereto.

The pharmaceutical composition for the prevention or treatment of inflammatory neurodegenerative diseases and the pharmaceutical composition for anti-inflammation may be prepared by using pharmaceutically acceptable and physiologically acceptable adjuvants in addition to the above-mentioned active ingredients. Examples of the adjuvants include excipients, A sweetener, a binder, a coating agent, a swelling agent, a lubricant, a lubricant or a flavoring agent.

The pharmaceutical composition may be formulated into a pharmaceutical composition containing at least one pharmaceutically acceptable carrier in addition to the above-described active ingredients for administration.

The pharmaceutical composition may be in the form of granules, powders, tablets, coated tablets, capsules, suppositories, liquids, syrups, juices, suspensions, emulsions, drops or injectable solutions. For example, for formulation into tablets or capsules, the active ingredient may be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, and the like. Also, if desired or necessary, suitable binders, lubricants, disintegrants and coloring agents may also be included as a mixture. Suitable binders include, but are not limited to, natural sugars such as starch, gelatin, glucose or beta-lactose, natural and synthetic gums such as corn sweeteners, acacia, tracker candles or sodium oleate, sodium stearate, magnesium stearate, sodium Benzoate, sodium acetate, sodium chloride, and the like. Disintegrants include, but are not limited to, starch, methyl cellulose, agar, bentonite, xanthan gum and the like. Acceptable pharmaceutical carriers for compositions that are formulated into a liquid solution include sterile solutions suitable for the living body such as saline, sterile water, Ringer's solution, buffered saline, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, One or more of these components may be mixed and used. If necessary, other conventional additives such as an antioxidant, a buffer, and a bacteriostatic agent may be added. In addition, diluents, dispersants, surfactants, binders, and lubricants may be additionally added to formulate into injectable solutions, pills, capsules, granules or tablets such as aqueous solutions, suspensions, emulsions and the like. Further, it can be suitably formulated according to each disease or ingredient, using the method disclosed in Remington's Pharmaceutical Science, Mack Publishing Company, Easton PA as an appropriate method in the field.

The present invention also provides a method of preventing or treating an inflammatory neurodegenerative or anti-inflammatory disease comprising administering a therapeutically effective amount of a top extract to an experimental subject other than a human.

It is preferable that the subject to be tested is a mammal to be treated or observed.

As used herein, the term "therapeutically effective amount " refers to the amount of active ingredient or pharmaceutical composition that induces a biological or medical response in a tissue system, animal or human, as contemplated by a researcher, veterinarian, This includes an amount that induces relief of the symptoms of the disease or disorder being treated. It will be apparent to those skilled in the art that the therapeutically effective dose and the number of administrations of the active ingredient of the present invention will vary depending on the desired effect. Thus, the optimal dosage to be administered can be readily determined by those skilled in the art and will vary with the nature of the disease, the severity of the disease, the amount of active and other ingredients contained in the composition, the type of formulation, and the age, The age, body weight, sex, diet, time of administration, route of administration and fraction of the composition, duration of treatment, concurrent medication, and the like.

The dose of the top extract as an active ingredient in the pharmaceutical composition of the present invention may be 6 to 30 mL per kg of body weight once or three times per day depending on the patient's age, sex, symptom, method of administration, or prevention purpose . Dosage levels for patients exhibiting the specific symptoms may vary depending on the patient's body weight, age, sex, health condition, diet, time of administration, administration method, excretion rate, severity of disease, and the like.

In the treatment method of the present invention, the composition comprising the extract of the present invention as an active ingredient can be administered orally, rectally, intravenously, intraarterally, intraperitoneally, intramuscularly, intrasternally, transdermally, topically, Lt; / RTI >

In addition, the present invention relates to a food composition for relieving inflammatory neurodegenerative diseases comprising an extract of Chrysanthemum morifolium as an active ingredient.

The present invention also relates to a functional food composition for antiinflammation comprising an extract of Chrysanthemum morifolium as an active ingredient.

The food composition of the present invention can be formulated and used as a functional food functionalized with a food composition for relieving inflammatory neurodegenerative diseases or as a functional food composition for antiinflammation, or can be added to various foods.

The form of the preparation may be formulated in the same manner as the pharmaceutical composition and used as a functional food or may be added to various foods.

Examples of foods to which the food composition for relieving inflammatory neurodegenerative diseases or the functional food composition for anti-inflammation of the present invention can be added include foods, beverages, meat, chocolates, foods, confections, pizza, ramen, Ice cream, alcoholic beverages, vitamin complexes, and health supplement foods.

The present invention also relates to a composition for antioxidation comprising an extract of Chrysanthemum morifolium as an active ingredient.

The present invention also relates to a method for inhibiting the concentration of nitrogen monoxide produced in glial cells in vitro by treating an alcohol extract of the extract of Liliaceae with lipopolysaccharide-stimulated glial cells.

The alcohol is preferably ethanol, and the effective amount of the extract is preferably 10 to 100 μg / mL.

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these examples are for illustrative purposes only and that the scope of the present invention is not construed as being limited by these examples.

Example 1. Preparation of Hizikia Fusiforma Extract, HFE

The extracts were prepared by using ethanol as an extraction solvent, and 100 g of starch was added to 1 L of ethanol and extracted at room temperature for 72 hours.

Experimental Example 1. Antioxidant capacity measurement

The DPPH radical scavenging activity of the extract prepared in Example 1 was determined by adding 30 μL of the extract solution and 30 μL of the 60 μM DPPH solution dissolved in ethanol, and transferring it to the capillary tube at room temperature for 2 minutes. The electron spin resonance (ESR) spectrometer (JES-PX 2300, JEOL, Japan).

In this case, the measurement conditions of the ESR spectrophotometer are 336.5 ± 5 mT for the magnetic field, 5 mW for the microwave power, and 9.41 GHz for the modulation frequency. The DPPH free radical scavenging activity (%) for the antioxidative sample is given by the following equation (1).

[Equation 1]

(ESR signal intensity for the control medium)

As a result, it was found that DPPH radical scavenging activity was 20% or more at a concentration of 0.1 mg / mL and DPPH radical scavenging activity was 80% or more at a concentration of 100 μg / mL (FIG. 1).

Experimental Example 2. Measurement of cell viability

In order to examine the effect of LPS (lipopolysaccharide) and extracts on cell survival in BV-2 cells stimulated with LPS, cell biability was measured using 3- [4,5-Dimethylthiazol-2-yl] -2,5-diphenyl-tetrazolium bromide (MTT) assay.

Cells (1 × 10 ⁵ cells / ml) were seeded on a 96-well plate in a volume of 180 μL and cultured in a CO ₂ incubator for 12 hours or longer. After culturing, the culture medium was removed and 200 μL of medium containing 0.5 mg / mL MTT was added, followed by incubation for 4 hours to allow MTT reduction. After removing the culture medium, 100 μL of dimethylsulfoxide (DMSO) was added and the resulting formazone crystals were dissolved. The absorbance was measured at 540 nm using an ELISA reader, and the cell viability was expressed as a percentage (%) as compared with the control .

The cell viability can be determined by measuring the number of active cells and determining the intracellular toxicity of the extract.

As a result, it was confirmed that the cell survival rate was not changed in all the experimental groups treated with LPS or the extract alone or in the same manner (FIG. 2). This means that inflammation inducers, LPS, and Zygomyk and melon mixed extract do not affect cell survival.

Experimental Example  3. By LPS  Concentration-dependent inhibition of NO production in the extracts of goat extracts in activated glial cells

In order to analyze the anti - inflammatory effect of the extracts, LPS, which is an inflammation inducer, was examined to determine whether it was dependent on the concentration of nitrogen monoxide (NO) produced in stimulated glial cells.

The NO concentration was determined by seeding 5 × 10 ⁵ cells / well in a 24-well plate, adding LPS and extracts at different concentrations, reacting in an incubator for 24 hours, and then adding 50 μL each of Griess reagent (1% sulfanilamide / 0.1% N- (1-naphthyl) -ethylenediamine dihydrochloride / 2.5% H ₃ PO ₄ ) and the value was measured using a Sunrise basic 96-well microplate spectrophotometer (TECAN AUSTRIA) with a wavelength of 540 nm.

The NO concentration induced by LPS (1 ㎍ / mL) in the glial cells was about 36.5 μM, which was about 13 times higher than that of the control group (2.8 μM). As a result, 80, and 100 ㎍ / mL, the NO production was reduced in a concentration-dependent manner (FIG. 3).

Experimental Example  4. After treatment of root extracts in glial cells LPS  due to INOS induced  dependent inhibition of inducible nitric oxide synthase protein expression

Changes in iNOS protein expression patterns were observed after administration of the extract.

Protein expression immunoblot analysis was performed using BV-2 cells. The cells were washed twice with 1 × PBS and lysed with 1% Triton X-100, 1% deoxycholate, 0.1% NaN3 Respectively.

The protein was quantitatively analyzed by the bradford method of the BCA assay kit (bio-rad CA). The same amount of protein was electrophoresed on 10% SDS-PAGE and transferred from the gel using 0.45 polyvinylidene fluoride (PVDF: Millipore) Antibody was diluted to 1/1000 of anti-iNOS and 1/5000 of beta -actin as the primary antibody. O / N (over-night) was used at 4 and Horseradish peroxidase (HRP) The antibodies were reacted at room temperature for 1 hour and then reacted with supersignal (Pierce, CA), and the results were confirmed using LAS-3000 (Fuji Co, Japan).

Immunoblot analysis showed that the expression level of iNOS protein in the control group not treated with LPS was almost zero and the expression of iNOS protein in the experimental group treated with LPS was significantly increased. The concentration - dependent decrease of the expression level was observed after the treatment with the extracts. Beta-actin was used as a control for protein quantitation (Fig. 4).

Claims (12)

A pharmaceutical composition for preventing or treating an inflammatory neurodegenerative disease, comprising an extract of Chrysanthemum morifolium as an active ingredient. The pharmaceutical composition according to claim 1, wherein the composition weakens inductive nitrogen oxide synthase (iNOS) and nitrogen monoxide (NO) production. [Claim 2] The pharmaceutical composition according to claim 1, wherein the extracting solvent of the extract is an alcohol. The pharmaceutical composition for preventing or treating inflammatory neurodegenerative diseases according to claim 1, wherein the composition inhibits microglial cell activity in microglial cells. A food composition for relieving inflammatory neurodegenerative diseases comprising an extract of Chrysanthemum morifolium as an active ingredient. The food composition according to claim 5, wherein the food is beverage, jelly, candy, gum or dairy product. A pharmaceutical composition for antiinflammatory, comprising an extract of Chrysanthemum morifolium as an active ingredient. 8. The pharmaceutical composition according to claim 7, wherein the composition weakens inductive nitrogen oxide synthase (iNOS) and nitrogen monoxide (NO) production. A functional food composition for antiinflammatory, comprising an extract of Chrysanthemum morifolium as an active ingredient. An antioxidative composition comprising an extract of Chrysanthemum morifolium as an active ingredient. A method of inhibiting the concentration of nitric oxide produced in glial cells in vitro by treating an alcohol extract of the extract from a glycoprotein stimulated with lipopolysaccharide. [Claim 12] The method according to claim 11, wherein the effective amount of the extract is 10 to 100 [mu] g / mL.

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