KR20190125802A - Composition for Preventing or Treating Bone Disorders Comprising Extraction of Gentiana lutea - Google Patents

Abstract

The present invention relates to a composition for preventing or treating bone diseases containing a Gentiana lutea extract as an active component. In an embodiment of the present invention, the Gentiana lutea extract inhibits the differentiation of mononuclear cells isolated from mouse hematopoietic stem cells into osteoclasts, and exhibits an effect of inhibiting the expression of osteoarthritis-inducing factor in an osteoarthritis cell model. Therefore, the Gentiana lutea extract can inhibit bone absorption, which destroys bones, thereby being able to effectively prevent, alleviate or treat osteoporosis. In addition, the Gentiana lutea extract can inhibit the expression of osteoarthritis-inducing factors, thereby being able to effectively prevent, alleviate or treat osteoarthritis.

Description

Composition for Preventing or Treating Bone Disorders Comprising Extraction of Gentiana lutea}

The present invention relates to a pharmaceutical composition for preventing or treating bone disease, comprising a Gentiana lutea extract as an active ingredient, and a health functional food composition for preventing or improving bone disease.

Osteoclast, a multinucleated large cell, has the function of destroying and absorbing bone tissue, and is known to play a role in destroying bone matrix and degrading bone minerals. Activated osteoclasts have three or more nuclei. To differentiate from osteoclast progenitor cells into mature multinucleated osteoclasts, macrophage colony stimulating factor (MCSF) and receptor activator of nuclear factor-kappa B ligand (RANKL) It requires a variety of hormones and factors such as

The osteoclasts cause disruption and absorption of abnormal bone tissue due to imbalance with osteoblasts in the bone, thereby causing osteoporosis, which causes bone mass and bone density to decrease, osteoporosis, and lime in the bone. Is known to cause osteomalacia, bone marrow fibrous ostitis, periodontitis in which alveolar bone is lost, and rheumatoid arthritis, which causes joint destruction and deformation. have.

Osteoporosis is a condition in which bone mass decreases due to various causes and the risk of fracture is continuously increased due to the degeneration of the microstructure of bone tissue. As a result, the balance of aggregate formation is broken, resulting in an increase in osteoclasts than osteoblasts. Normal bone inside has a dense structure like a mesh, but in osteoporosis patients, the gap between the bone structures becomes wider and the microstructure becomes thinner and weaker, so that the bones can be easily broken even in the small impact.

Currently, the treatment of bone damage caused by osteoclasts such as osteoporosis is widely used for the treatment of bisphosphonate-based drugs such as Fosamax (component name: aledronate), Actonel (component name: risedronate), and Zometa (component name: zoledronate). Although these agents are mostly used to induce the death of osteoclasts to delay or inhibit the loss of bone, and thus have various side effects such as osteonecrosis, severe atrial fibrillation, incapacitation of bones or joints, and pain in musculoskeletal pain. There is a problem that causes. Therefore, there is a need for the development of medicines that can compensate for the shortcomings of existing bone disease treatment drugs.

Osteoarthritis, on the other hand, is a disease mainly characterized by the loss of proteoglaycan (PG), a component of articular cartilage, which causes the destruction of cells and components of articular cartilage, leading to joint dysfunction. to be. The expression of MMP-3, MMP-9, and MMP-13 increases when osteoarthritis develops, and it is known that the increase of MMPs aggravates degenerative arthritis by damaging the collagen matrix constituting the cartilage.

Nonsteroidal anti-inflammatory drugs (NSAIDs), analgesics, hyaluronan, glucosamine (glucosamine), and chondroitin (chondroitin) are currently used to treat osteoarthritis. Some NSAIDs are known to inhibit the proteoglaycan synthesis of articular cartilage, which tends to exacerbate osteoarthritis symptoms.

Therefore, there is an urgent need to develop alternative usages that can protect joint cartilage itself or directly treat damage. In this regard, efforts to find a natural osteoarthritis-improving agent has been tried in various ways. In particular, in recent years, efforts to find a new therapeutic agent having an improved effect on osteoarthritis by using foods or ingredients derived from natural products are in progress. It is becoming.

Korean Patent Publication No. 10-2016-0043844 (published Apr. 22, 2016)

It is an object of the present invention to provide a pharmaceutical composition that can effectively prevent or treat bone disease.

Another object of the present invention to provide a nutraceutical composition that can effectively prevent or improve bone diseases.

In order to achieve the above object, the present invention provides a pharmaceutical composition for the prevention or treatment of bone disease, including Gentian lutea extract (Gentiana lutea) as an active ingredient.

In order to achieve the above another object, the present invention provides a health functional food composition for preventing or improving bone diseases comprising Gentian lutea extract (Gentiana lutea) as an active ingredient.

The present invention relates to a composition for preventing, improving or treating bone diseases containing Gentiana lutea extract as an active ingredient. According to the present invention, Gentiana Lutea extract inhibits differentiation into osteoclasts and destroys bone. As a composition for the prevention, improvement and treatment of bone diseases including osteoporosis and osteoarthritis, because it can inhibit bone absorption, inhibit the factors causing joint inflammation and cartilage degeneration, thereby preventing or treating cartilage degeneration. It can be useful.

1 is a result confirming the osteoclast differentiation inhibitory effect of the gentian lutea extract.
Figure 2 is a result of confirming the inhibitory effect of the expression of COX-2, MMP-3 and MMP-13 factors that cause arthritis and cartilage degeneration of gentiana lutea extract in osteoarthritis cell model.

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

The present invention provides a pharmaceutical composition for preventing or treating bone disease, comprising an extract of Gentiana lutea as an active ingredient.

The gentiana lutea extract may be prepared by extracting the root of gentiana lutea with 20% to 50% ethanol aqueous solution, and more preferably, by extracting with 30% ethanol aqueous solution. Bar, if extracted under such conditions is preferable because the effect of preventing or treating bone disease is maximized.

Preferably, the bone disease may be osteoporosis or osteoarthritis, that is, the gentian lutea extract may prevent or treat osteoporosis by inhibiting the differentiation of mononuclear cells into osteoclasts, which may cause osteoporosis. In addition, osteoarthritis can be prevented or treated by suppressing the expression of COX-2, MMP-3 and MMP-13, which are factors of osteoarthritis and factors inducing joint inflammation and cartilage degeneration.

More preferably, the osteoporosis may be postmenopausal osteoporosis, but is not limited thereto.

As used herein, the term "prevention" refers to any action that inhibits or delays a disease by administration of a composition containing the gentian or lutea extract. In addition, the term "treatment" as used in the present invention means all the actions that improve or cure the symptoms of the disease by administration of the composition containing the gentian lutea extract.

The pharmaceutical composition of the present invention may include a pharmaceutically acceptable carrier, excipient or diluent in addition to the above-mentioned active ingredient for administration. The carrier, excipient and diluent may include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, Polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.

The pharmaceutical compositions of the present invention may be formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, oral formulations, external preparations, suppositories, or sterile injectable solutions according to conventional methods. . In detail, when formulated, it may be prepared by using diluents or excipients such as fillers, weights, binders, wetting agents, disintegrating agents, and surfactants which are commonly used. Solid preparations for oral administration include, but are not limited to, tablets, pills, powders, granules, capsules, and the like. Such solid preparations may be prepared by mixing at least one excipient such as starch, calcium carbonate, sucrose, lactose, gelatin and the like with the gentiana lutea extract. In addition to simple excipients, lubricants such as magnesium stearate, talc can also be used. It may be prepared by adding various excipients such as humectants, sweeteners, fragrances, preservatives and the like in addition to liquid oral liquids or liquid paraffin for oral use. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized formulations and tasks. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, injectable esters such as ethyl oleate, and the like can be used. As the base of the suppository, utopsol, macrosol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

Suitable dosages of the compositions of the present invention vary depending on the condition and weight of the patient, the extent of the disease, the form of the drug, and the time of day, but may be appropriately selected by those skilled in the art, and the daily dosage of the gentiana lutea extract is preferred. Preferably it is 1 mg / kg to 500 mg / kg, it can be administered once or divided into several times as needed.

Furthermore, the present invention provides a health functional food composition for preventing or improving bone disease, including Gentian lutea extract as an active ingredient.

The gentiana lutea extract may be prepared by extracting the root of gentiana lutea with 20% to 50% ethanol aqueous solution, and more preferably, by extracting with 30% ethanol aqueous solution. Bar, if extracted under such conditions is preferred because it maximizes the effect of preventing or improving bone disease.

More preferably, the bone disease may be osteoporosis or osteoarthritis, that is, the Gentiana lutea extract prevents or ameliorates osteoporosis by inhibiting the differentiation of mononuclear cells into osteoclasts, which may cause osteoporosis. In addition, osteoarthritis can be prevented or improved by inhibiting the expression of COX-2, MMP-3 and MMP-13, which are factors of osteoarthritis and factors inducing joint inflammation and cartilage degeneration.

More preferably, the osteoporosis may be postmenopausal osteoporosis, but is not limited thereto.

The dietary supplement composition includes various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, coloring and neutralizing agents (such as cheese and chocolate), pectic acid and salts thereof, alginic acid and salts thereof. , Organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated drinks, and the like. Others may contain pulp for the production of natural fruit juices, synthetic fruit juices and vegetable drinks. These components can be used independently or in combination. In addition, the nutraceutical composition is in the form of meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, gum, ice cream, soup, beverages, tea, functional water, drinks, alcohol and vitamin complexes Can be.

In addition, the health functional food composition may further include a food additive, the suitability as a "food additive" is the item according to the General Regulations and General Test Methods of the Food Additives Code approved by the Food and Drug Administration unless otherwise specified It is determined by the standard and the standard.

Items listed in the "Food Additives Code" include, for example, chemical compounds such as ketones, glycine, potassium citrate, nicotinic acid, cinnamon acid, natural additives such as color pigments, licorice extract, crystalline cellulose, high-quench pigments, guar gum, L Mixed preparations, such as a sodium glutamate preparation, a noodles addition alkali agent, a preservative preparation, and a tar pigment preparation, etc. are mentioned.

At this time, the gentian lutea extract according to the present invention added to the food in the process of preparing the health functional food composition can be appropriately added or subtracted as needed, preferably 1 part by weight to 15 parts by weight of food It is preferable to add so as to include by weight.

In addition, the present invention provides a reagent composition that inhibits the differentiation of mononuclear cells into osteoclasts in vitro, including gentiana lutea extract as an active ingredient.

The present invention also provides a reagent composition that inhibits the expression of COX-2, MMP-3 and MMP-13 in vitro, including Gentiana lutea extract as an active ingredient.

Furthermore, the present invention provides a method for inhibiting differentiation of mononuclear cells into osteoclasts, comprising the step of treating the gentian or lutea extract to an animal except a human.

The present invention also provides a method for inhibiting expression of COX-2, MMP-3 and MMP-13, comprising the step of treating a gentian or lutea extract to an animal other than a human.

Hereinafter, examples will be described in detail to help understand the present invention. However, the following examples are provided to more fully explain the present invention to those skilled in the art, and the scope of the present invention is not limited to the following examples, only to illustrate the contents of the present invention. no.

Example 1 Cell Experiment for Confirming the Effect of Osteoporosis Improvement

Osteoclasts are derived from hematopoietic stem cells and are responsible for bone resorption, which destroys aged bone, and maintains bone remodeling through a balanced action of bone formation by osteoblasts and bone resorption by osteoclasts. . On the other hand, osteoporosis refers to a disease in which osteoblasts increase bone resorption activity compared with osteoblasts, and as a result, total bone mass decreases and fractures are induced even with a slight impact. To date, there is no effective treatment for the cure of osteoporosis and prevention is emphasized.

Thus, the inventors of the present invention obtained hematopoietic stem cells in the mouse thighs to confirm the osteoporosis treatment effect of the gentian lutea extract, and to separate the mononuclear cells that are osteoclast precursors from the hematopoietic stem cells, as shown in Figure 1a , FACS was performed. At this time, CD11b (purchased by Sigma), a mononuclear cell positive marker antibody, was used to identify and isolate the presence of primary monocytes.

In addition, 50 g of Gentiana lutea root was added to 0.5 L of 30% ethanol aqueous solution, followed by two steps of cooling for 24 hours, followed by filtration and concentration under reduced pressure to prepare 2.3 g of Gentiana Lutea extract.

Thereafter, the mononuclear cells obtained above were cultured in alpha-MEM medium and treated with 30 ng / mL M-CSF and 50 ng / mL RANKL to induce differentiation for 3 days, and the gentiana lutea extract prepared above was Treated at concentrations of 2 μg / ml (GL2), 10 μg / ml (GL10) and 50 μg / ml (GL50), the tartrate-resistant acid phosphatase (TRAP), an osteoclast differentiation marker Activity was measured. At this time, after washing the medium containing the cells with physiological saline, by using the Acid-Phosphatase Kit to measure the absorbance at a wavelength of 405 nm to the cells of the medium was confirmed the TRAP activity, TRAP staining under a microscope The osteoclasts were observed through.

As a result, as shown in Figures 1a and 1b, from the hematopoietic stem cells isolated from the mouse femoral monocytes (Monocyte), the mother of osteoclasts was well separated, it can be confirmed that the mononuclear cells separated into osteoclasts through the inducer. there was. In addition, Gentiana Lutein was treated with 2 μg / ml (GL2), 10 μg / ml (GL10), and 50 μg / ml (GL50) concentrations, and significantly improved in the 50 μg / ml treatment group (GL50). As the activity was reduced, it was confirmed that osteoclast differentiation activity was significantly suppressed.

Based on these results, Gentiana lutea extract was able to effectively inhibit osteoclast differentiation, it was confirmed that it can be effectively used for the prevention, improvement and treatment of osteoporosis.

Example 2 Cell Test for Confirming the Effect of Osteoarthritis Improvement

Osteoarthritis (degenerative arthritis) is a disease that causes degeneration of articular cartilage and structural changes in the bone under the cartilage. The pathological cause of degenerative arthritis is not clear yet, but the main cause is cartilage due to repeated use of cartilage during aging. Wear, heredity, damage to cartilage tissue by impact, overweight pressure, muscle weakness around the knee. There are symptomatic therapies, such as pain control and surgery, and fundamental therapeutics have not yet been developed.

Accordingly, the inventors of the present invention, in order to confirm the arthritis-improving effect of Gentiana Lutea extract, isolated chondrocytes from cartilage of 5 days old mouse and cultured the cells in DMEM medium, and then interleukin- Osteoarthritis cell models were constructed by treating 1β (IL-1β). The negative control group (control) was used for cells that did not cause osteoarthritis.

Then, the gentiana lutea extract was prepared according to the method described in Example 1, and in order to verify whether the catabolic factor expression by the gentiana lutea extract was expressed at the cellular level, cartilage cells secrete It was confirmed whether catabolic elements were inhibited by the extract. That is, the cells were treated with gentiana lutein extract at concentrations of 2 μg / ml (GL2), 10 μg / ml (GL10) and 50 μg / ml (GL50), followed by COX-2, MMP-3 and MMP-. 13 was subjected to biochemical analysis. That is, the expression of the COX-2, MMP-3 and MMP-13 was measured via RT-PCR according to a conventionally known method.

As a result, as shown in Figure 2, when treated with gentiana lutein extract, expression of osteoarthritis-inducing factors COX-2, MMP-3 and MMP-13 was significantly inhibited, in particular gentiana lutein extract 50 ug A particularly good inhibitory effect was seen at the / ml concentration.

Based on these results, Gentiana lutea extract was able to effectively inhibit the expression of osteoarthritis-inducing factors, it was confirmed that it can be effectively used for the prevention, improvement and treatment of osteoarthritis.

Having described the specific part of the present invention in detail, it is obvious to those skilled in the art that such a specific description is only a preferred embodiment, thereby not limiting the scope of the present invention. Do. That is, the substantial scope of the present invention is defined by the appended claims and their equivalents.

Claims (10)

Gentiana Gentiana Lutea ) A pharmaceutical composition for preventing or treating bone diseases comprising the extract as an active ingredient. The pharmaceutical composition for preventing or treating bone disease according to claim 1, wherein the extract of Gentiana Lutea is extracted from 20% to 50% aqueous ethanol solution. The pharmaceutical composition for preventing or treating bone disease according to claim 1, wherein the bone disease is osteoporosis or osteoarthritis. The pharmaceutical composition for preventing or treating bone disease according to claim 3, wherein the gentiana lutea extract exhibits an osteoporosis prevention or treatment effect by inhibiting differentiation of mononuclear cells into osteoclasts. According to claim 3, The gentian lutea extract is characterized in that it exhibits an osteoarthritis prevention or treatment effect by inhibiting the expression of COX-2, MMP-3 and MMP-13 factors that cause joint inflammation and cartilage degeneration. Pharmaceutical composition for preventing or treating bone diseases. Gentiana Gentiana Lutea ) Health functional food composition for preventing or improving bone diseases comprising the extract as an active ingredient. According to claim 6, The gentiana lutea extract is a health functional food composition for preventing or improving bone disease, characterized in that the root of the gentian lutea extracted with 20% to 50% ethanol aqueous solution. According to claim 6, wherein the bone disease is osteoporosis or osteoarthritis bone disease prevention or improvement health functional food composition, characterized in that. According to claim 6, The gentian lutea extract is a health functional food composition for the prevention or improvement of bone diseases, characterized in that to prevent or improve osteoporosis by inhibiting the differentiation of mononuclear cells into osteoclasts. According to claim 6, The gentian lutea extract is characterized in that it exhibits an osteoarthritis prevention or improvement effect by inhibiting the expression of COX-2, MMP-3 and MMP-13 factors that cause joint inflammation and cartilage degeneration. Health functional food composition for preventing or improving bone disease.

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