KR101569078B1 - Composition comprising extract of Phragmites spp. as an effective component for prevention or treatment of metabolic bone disease - Google Patents

Abstract

The present invention relates to a composition for prevention or treatment of bone metabolic diseases such as osteoporosis, osteoporosis or fracture, which comprises a reed extract as an active ingredient. The reed extract of the present invention is a composition for preventing or treating osteoblast differentiation, which comprises ALP, osterix, RUNX2 (BMD) in an ovariectomized osteoporosis animal model, and reduce fat cells in bone marrow, and can be used as medicines or health functional foods useful for bone metabolism diseases.

Description

[0001] The present invention relates to a composition for preventing or treating bone metabolism diseases, which comprises a reed extract as an active ingredient. As an effective component for prevention or treatment of metabolic bone disease,

The present invention relates to a composition for the prevention and treatment of bone metabolic diseases comprising an extract of Phragmites spp. As an active ingredient . The composition of the present invention can be used for the production of health functional foods and medicines for the prevention and treatment of osteoporosis have.

Osteoporosis refers to a skeletal disorder in which the risk of fracture and bone fragility are increased due to low bone mass and bone destruction. Osteoporosis is a particularly frequent occurrence in postmenopausal women, a disease in which the amount of bone is markedly reduced by the decreased secretion of estrogen. The decrease in the amount of bone may be due to individual differences or various other causes, but if the amount of bone excessively decreases and falls below a predetermined value, fracture easily occurs even in a small impact. Osteoporosis can not lead to a healthy life by restricting long-term activities due to various fractures caused by weakening of the bones, especially femur fracture or vertebral fracture, rather than the symptom itself, resulting in 15% of the elderly death It is known.

Osteodystrophy is also called bone dystrophy and is a bone disease caused by chronic renal failure. It is born by abnormally abnormal kidney function, and when the kidney is weak, it does not dialyse. This kind of bone disease is called Renal Osteodystrophy. Bone diseases associated with osteophagia include osteomalacia fibrous osteitis (osteitis fibrosa).

On the other hand, reed ( Phragmites australis, Phragmites communis Trinius.) Distributed in Korea is distributed in all regions of Korea, from Hantan to Burma. It grows in a wet place such as a river, a den, a waterway, etc. It is about 1 ~ 3m in size and it blooms from August to September. The fruit is a spirit and the seed has a tangent, which is easily blown away in the wind and reproduced by seeds and undergrowth. Leaves are broad and long, pointed end, 20 ~ 50cm in length, 2 ~ 4cm in width, sheaths are hairless, short tusks, hairy on edge, feather-shaped flower buds, stems are straight and smooth. The roots stretch long to the ground and have yellow beard roots in the nodes.

The root of the root used in oriental medicine has been used for drying the roots of reeds and has been used for hemostasis, gastric motility, and diuretic effect. However, the effect on the bone turnover disease is not yet known.

It is an object of the present invention to provide a composition for the prevention or treatment of bone metabolic diseases such as osteoporosis, osteoporosis or fracture derived from a natural product having no toxicity.

The composition for preventing or treating bone metabolic diseases of the present invention contains a reed extract as an active ingredient.

The reed extract of the present invention is selected from reeds ( Phragmites communis Trin. Or Phragmites australis ), Phragmites japonica Steud., Phragmites karka (Retz. Trin. Ex Steud.) And their inbred plants It is an extract of one or more plants, preferably a reed ( Phragmites communis Trin. Or Phragmites australis ).

The term "reeds" or "reed extracts" used in the present invention means, unless specifically limited, reptiles ( Phragmites spp. ) Or their extracts.

In the present invention, the reed extract may be an extract of a reed stalk, a part of leaves and roots, or an outpost, preferably a reed root extract.

As used herein, the term "comprising as an active ingredient" means an amount sufficient to achieve the efficacy or activity of the reed extract.

The bone metabolic diseases of the present invention include osteoporosis, bone formation disorder or fracture.

In the present invention, 'osteoporosis' includes clinical classification according to the measurement of bone mineral density (BMD), ie, osteopenia, osteoporosis, and severe osteoporosis. It also includes primary osteoporosis type 1 (postmenopausal osteoporosis type 1), type 2 senile osteoporosis type 2 (secondary osteoporosis), and secondary osteoporosis.

In the present invention, 'osteodystrophy' includes osteomalacia, osteitis fibrosa and the like.

In the present invention, 'fracture' refers to a state in which the continuity of bone or cartilage is completely or incompletely lost or a linear deformation occurs. Fractures are pathologic fractures caused by osteoporosis and tumor osteomyelitis, which are anatomical location, extent of fracture, direction of fracture, presence of open window, number of fractures, stability, dislocation of fracture, And fatigue fractures caused by being applied, and the term "fracture" includes all of them.

The extract of the present invention may be an extract of purified water, water, an organic solvent or a mixed solvent thereof. Preferably, the organic solvent may be at least one solvent selected from the group consisting of alcohols having 1 to 4 carbon atoms, hexane, dichloromethane, ethyl acetate, acetone, chloroform and diethyl ether, more preferably methanol, ethanol or methanol And an aqueous solution of ethanol. In addition to the organic solvent, polar solvents such as acetic acid, dimethyl-formamide (DMFO) and dimethyl sulfoxide (DMSO), acetonitrile, ethyl acetate, methyl acetate, fluoroalkane, pentane, 2,2,4-trimethylpentane, decane, Butene, 1-chloropentane, o-xylene, diisopropyl ether, 2-chloropropane, toluene, 1-chloropropane, chlorobenzene, Non-polar solvents such as benzene, diethyl ether, diethyl sulfide, 1,2-dichloroethane, aniline, diethylamine, ether, carbon tetrachloride and THF (Tetrahydrofuran) may be used.

The term 'extract' used in the present invention includes fractions obtained by further fractionating the above extract. That is, not only the water, the lower alcohol having 1 to 4 carbon atoms, the nonpolar solvent, or the mixed solvent thereof, but also the fractions fractionated by the above-mentioned solvents, or the ultrafiltration membrane having various molecular weight cutoff values And fractions obtained.

The method for producing the extract of the present invention can be carried out by, for example, shredding reed roots and crushing the reed roots, and then adding water, an organic solvent or a mixture thereof to the dried sample in an amount of about 1 to 50 times, preferably about 10 to 40 times, Mixing extraction, hot water extraction, reflux cooling extraction, ultrasonic extraction or supercritical extraction at 20 to 100 ° C, preferably 40 to 60 ° C for about 24 hours, or 2 to 4 hours, The extract of the present invention can be obtained by filtration, concentration under reduced pressure, or drying.

The reed extract of the present invention exhibits an osteoblast differentiation increasing activity and a bone density increasing activity, and contains the reeds extract in an amount of 0.1 to 50% by weight based on the total weight of the composition. However, the composition is not limited thereto, and may vary depending on the condition of the patient, the type of disease, and the progress of the disease.

The composition for prevention or treatment of bone metabolic diseases including the reed extract of the present invention may further comprise suitable carriers, excipients and diluents conventionally used in the production of pharmaceutical compositions.

The composition containing the extract according to the present invention may be formulated in the form of powders, granules, tablets, capsules, oral preparations such as suspensions, emulsions, syrups and aerosols, external preparations, suppositories and sterilized injection solutions, Can be used.

Examples of carriers, excipients and diluents that can be contained in the composition containing the reed extract include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, Calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.

In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid formulations for oral administration include tablets, pills, powders, granules, capsules and the like, which may contain at least one excipient such as starch, calcium carbonate, sucrose ), Lactose, gelatin and the like. In addition to simple excipients, lubricants such as magnesium stearate talc are also used. Examples of the liquid preparation for oral use include suspensions, solutions, emulsions, and syrups. In addition to water and liquid paraffin, simple diluents commonly used, various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included . Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Examples of the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. Examples of the suppository base include witepsol, macrogol, tween 61, cacao butter, laurin, glycerogelatin and the like.

The preferred dose of the reed extract of the present invention varies depending on the condition and body weight of the patient, the degree of disease, the drug form, the administration route and the period, but can be appropriately selected by the therapist. However, for the desired effect, the extract of the present invention is preferably administered at a dose of 0.01 mg / kg to 10 g / kg per day, preferably 1 mg / kg to 1 g / kg per day. The administration may be carried out once a day or divided into several doses. Accordingly, the dosage is not limited in any way to the scope of the present invention.

The composition of the present invention may be administered to mammals such as rats, mice, livestock, humans, and the like in various routes. All modes of administration may be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intra-uterine or intracerebroventricular injections.

The present invention provides a health functional food for preventing or ameliorating bone metabolic diseases comprising a reed extract and a food acceptable food supplementary additive exhibiting osteoblast differentiation increasing activity and bone mineral density increasing activity.

The health functional food of the present invention includes various foods, gums, tea, vitamin complex, health supplement foods and the like, and can be used as powder, granule, tablet, capsule or beverage.

Since the reed extract of the present invention has little toxicity and side effects, it can be safely used for prolonged use even for prophylactic purposes.

When the reed extract of the present invention is added to a food or beverage for the purpose of preventing or improving bone metabolic diseases, the amount of the extract in the food or beverage generally ranges from 0.01 to 15% by weight of the total food weight of the health food composition of the present invention And the health beverage composition may be added at a ratio of 0.02 to 10 g, preferably 0.3 to 1 g, based on 100 ml.

The health beverage composition of the present invention may contain various flavors or natural carbohydrates such as ordinary beverages as an additional ingredient, as long as it contains the extract as an essential ingredient at the indicated ratio, and there is no particular limitation to the liquid ingredient. Examples of the above-mentioned natural carbohydrates include monosaccharides such as disaccharides such as glucose and fructose such as maltose, sucrose and the like and polysaccharides such as dextrin, cyclodextrin and the like Sugar, and sugar alcohols such as xylitol, sorbitol, and erythritol. Natural flavors (tau martin, stevia extracts (e.g., rebaudioside A, glycyrrhizin, etc.) and synthetic flavors (saccharin, aspartame, etc.) can be advantageously used as flavors other than those described above The ratio of the natural carbohydrate is generally about 1 to 20 g, preferably about 5 to 12 g per 100 ml of the composition of the present invention.

In addition to the above, the health functional food of the present invention may contain flavorings such as various nutrients, vitamins, minerals (electrolytes), synthetic flavors and natural flavors, colorants and heavies (cheese, chocolate etc.), pectic acid and its salts, And salts thereof, organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonating agents used in carbonated drinks, and the like. In addition, the health functional foods of the present invention 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. The proportion of such additives is not so important, but is generally selected in the range of 0.1 to about 20 parts by weight per 100 parts by weight of the health functional food of the present invention.

The present invention relates to a composition for prevention or treatment of bone metabolic diseases such as osteoporosis, osteoporosis or fracture, which comprises a reed extract as an active ingredient. The reed extract of the present invention is a composition for preventing or treating osteoblast differentiation, which comprises ALP, osterix, RUNX2 (BMD) in an ovariectomized osteoporosis animal model, and reduce fat cells in bone marrow, and can be used as medicines or health functional foods useful for bone metabolism diseases.

1A is a photograph of ALP staining of C3H10T1 / 2 cell line in Experimental Example 1 after 9 days of treatment with a reed root ethanol extract of Preparation Example 1 in an osteoblast differentiation medium. Fig. 1B is a photograph of ALP staining of C3H10T1 / 2 cell line in the solvent fraction of reed root ethanol fraction . The results are shown in FIG.
FIG. 2A is a graph showing the relative mRNA expression levels of genes related to osteoblast differentiation after treating C3H10T1 / 2 cell line with osteogenic differentiation medium for 9 days with the reed root ethanol extract of Preparation Example 1 in Experimental Example 2. FIG. 1 is a graph showing the relative mRNA expression levels of genes involved in osteoblast differentiation after primary mesenchymal stem cells were treated with the solvent fraction of the reed root ethanol fraction of Preparation Example 2 for 9 days in the osteoblast differentiation medium.
FIG. 3 is a graph showing the results of an experiment in which ovariectomized experimental group (Sham,)), ovariectomized experimental group (Ovx,)), and ovariectomized experimental group 1, reed root ethanol extract (Et-OH) (Ovx + Sample 100 mg / kg, ▴).
FIG. 4 is a graph showing the bone mineral density of each experimental group when the rat became 19 weeks old in Experimental Example 3. FIG.
FIG. 5 is a graph showing relative mRNA expression levels of genes involved in osteoblast differentiation in each experimental group in Experimental Example 3. FIG.
FIG. 6 is a photograph of a tibial section of each experimental group in Experimental Example 3, histologically analyzed by H & E staining.

Hereinafter, the present invention will be described in detail with reference to Examples and Experimental Examples.

However, the following examples and experimental examples are illustrative of the present invention, and the present invention is not limited thereto.

Preparation Example 1 Preparation of Extracts of Reeds ( Phragmites communis Trin.)

Reeds ( Phragmites communis Trin.) Were purchased from the medicines of Kyungdong market and reed roots ethanol extracts were prepared.

The roots of the purchased reed were completely dried for 2 days at room temperature and finely pulverized to obtain reed root powder samples. Then, 200 ml of 99.9 (v / v)% ethanol was added per 10 g of the powder sample, rpm and 40 ° C for 24 hours. The supernatant was filtered with Watman No.1 filter paper, and 99.9% ethanol was added to the remaining reed root samples, and 200 ml per 10 g of the sample was added. Lt; / RTI > The filtrate was concentrated under reduced pressure at 45 ° C using a rotary vacuum evaporator to completely remove the solvent to prepare a reed root ethanol extract (Et-OH).

Production Example 2: Reed ( Phragmites communis  Preparation of Solvent Fraction of Trin.

The reed root ethanol extract (Et-OH) was fractionated stepwise using solvents with different polarity.

The reed root ethanol extract of Preparation Example 1 (Et-OH), hexane and water were mixed at a weight ratio of 1:20:20, and the mixture was extracted and concentrated to obtain a reed root hexane fraction (Hex). The aqueous fraction was further fractionated with dichloromethane, ethyl acetate and butanol to obtain a dichloromethane fraction (DCM), ethyl acetate fraction (EtOAC), butanol fraction (Bu-OH) and water fraction (H ₂ O) Concentrated, and lyophilized to use.

Production Example 3: Phragmites japonica  Steud.) Preparation of extract

( Phragmites japonica Steud.) Was purchased from the medicinal product of Kyungdong market and the extract of the rootstock root roots ethanol was prepared in the same manner as in Preparation Example 1.

After the roots of the purchased rootstock were completely dried at room temperature for 2 days and finely ground to obtain a sample of rootstock root root powder, 200 ml of 99.9 (v / v)% ethanol was added per 10 g of the powder sample, , And the supernatant was filtered with Watman No.1 filter paper. 100 ml of 99.9% ethanol per 10 g of sample was added to the remaining rootstock root extract, and the extract was extracted again for 12 hours. The filtration was carried out in the same manner as described above. The filtrate was concentrated under reduced pressure at 45 [deg.] C using a rotary vacuum evaporator to completely remove the solvent to prepare an ethanol extract of the rootstock root (Et-OH).

Preparation Example 4: Preparation of a large rootstock pull ( Phragmites carcass (Retz.) Trin. ex Steud.) Preparation of extract

( Phragmites karka (Retz.) Trin. Ex Steud.) Was purchased from the medicinal product of Kyungdong market and a large rootstock root ethanol extract was prepared in the same manner as in Preparation Example 1.

The roots of the above-purchased large rootstocks were thoroughly dried at room temperature for 2 days and finely pulverized to obtain a large rootstock root root powder sample. Then, 200 ml of 99.9 (v / v)% ethanol was added per 10 g of the powder sample, The supernatant was filtered with Watman No.1 filter paper, and 99.9% ethanol was added to the remaining large rootstock roots. 100 ml per 10 g of the sample was added again, and then 12 Time was extracted and filtered in the same manner as described above. The filtrate was concentrated under reduced pressure at 45 ° C using a rotary vacuum evaporator to completely remove the solvent to prepare a large rootstock ethanol extract (Et-OH).

Experimental Example 1. Measurement of ALP activity in C3H10T1 / 2 cells

(1) Measurement of ALP (alkaline phosphatase) activity of the extracts of Preparation Examples 1, 3 and 4

The C3H10T1 / 2 cell line derived from mouse embryonic fibroblasts is a pluripotent stem cell line that can differentiate into various cell lines including osteoblasts. One of the characteristics of the osteoblast is that it exhibits ALP (Alkaline phosphatase) activity. Therefore, the osteoblast differentiation effect was measured by ALP activity of C3H10T1 / 2 cell lines.

The C3H10T1 / 2 cell line was cultured in DMEM medium supplemented with 10% FBS, 1% penicillin and streptomycin at 37 ° C in a 5% CO 2 environment. The C3H10T1 / 2 cells were cultured on a 6-well plate with a medium containing 10 mM glycerophosphate and 50 / / ml ascorbic acid for osteoclast differentiation at a concentration of 2.5 ㅧ 10⁴ / ml, 10 μg / ml, 20 μg / ml and 40 μg / ml of the root ethanol extract (Et-OH) and 40 μg / ml of the extracts of Preparation Examples 3 and 4 were added, Lt; / RTI >

The results of staining for confirmation of ALP activity according to the concentration of the reed root ethanol extract of Preparation Example 1 and the negative control group are shown in FIG.

The ALP activity was measured by measuring the change in absorbance at 405 nm using p-nitrophenylphosphate decomposed into p-nitrophenol and phosphate, and the negative control was set to 0 % And the positive control group was 100%, the relative activity of ALP was measured and shown in Table 1. The negative control (Ctrl) was 10% FBS-DMEM and the positive control was NaF (0.001 ㎍ / ml).

division Concentration (/ / ml) Relative activity (%) for positive control Negative control group 0 0 Positive control group 0.001 100 Production Example 1 10 83 Production Example 1 20 85 Production Example 1 40 88 Production Example 3 40 80 Production Example 4 40 80

Although the ethanol extract of reed roots (Et-OH) in Preparation Example 1 is lower than that of the positive control, it can be confirmed that the osteoblast differentiation activity is increased in a concentration-dependent manner by the relative activity of ALP. In the same 40 ㎍ / ml, It was confirmed that osteoblast differentiation activity was excellent in reed compared with large.

(2) Manufacturing example  2 of Fraction ALP ( alkaline 포스화제 ) Active measurement

(Hex), a dichloromethane fraction (DCM), an ethyl acetate fraction (EtOAC), a butanol fraction (Bu-OH) and a water fraction (H ₂ O) was added at a concentration of 10 / / ml. After 9 days of differentiation, the result of staining for ALP activity was shown in FIG. 1B. The relative activity of ALP was measured when the negative control was 0% and the positive control was 100% Are shown in Table 2.

division Concentration (/ / ml) Relative activity (%) for positive control Negative control group 0 0 Positive control group 0.001 100 Et-OH 10 83 Hex 10 70 DCM 10 98 EtOAC 10 55 Bu-OH 10 0 H ₂ O 10 0

In the dichloromethane fraction (DCM), the ALP relative activity was significantly higher than that of the reed root ethanol extract (Et-OH) of Preparation Example 1. The hexane fraction (Hex) and the ethyl acetate fraction (EtOAC) , But the butanol fraction (Bu-OH) and the water fraction (H ₂ O) did not show ALP activity.

Experimental Example 2 Measurement of Osteoblast Differentiation Factor Expression by Real-time PCR

(1) Measurement of osteoblast differentiation factor expression level of the extract of Preparation Example 1

After the C3H10T1 / 2 cells were differentiated for 9 days in Experimental Example 1 (1), the amount of mRNA expression of ALP and osteopontin, which are the differentiation factors involved in osteoblast formation, was confirmed by Real-time PCR, Table 3 shows the results.

division 0 ㎍ / ml 10 [mu] g / ml 20 ug / ml 40 [mu] g / ml ALP 0.3049 + 0.0095 0.3575 + 0.0480 0.4852 0.0718 0.9169 + 0.2271 Osteopontin 0.0378 0.0026 0.0546 ± 0.0099 0.0493 0.0075 0.0775 0.0021

The expression of ALP and osteopontin mRNA was higher in C3H10T1 / 2 cells at all concentrations of 10 ㎍ / ml, 20 ㎍ / ml and 40 ㎍ / ml in osteoblast differentiation compared to the negative control, especially at 40 ㎍ / ml , The expression of ALP and osteopontin mRNA was significantly increased.

(2) Measurement of osteoblast differentiation factor expression level of the fraction of Preparation Example 2

Primary mesenchymal stem cells collected from mice instead of C3H10T1 / 2 cells were treated with 10 占 퐂 / ml of the solvent fraction of Preparation Example 2 and cultured in medium (1) in Experimental Example 1 every 3 days , And the mRNA expression levels of ALP, OCN, Osterix and RUNX2, which are the differentiation factors involved in osteoblast formation, were confirmed by Real-time PCR, and they are shown in FIG. 2B and Table 4, respectively.

division Et-OH Hex DCM EtOAC ALP 0.01688 0.01452 0.02113 0.00959 OCN 0.00042 0.00039 0.00051 0.00029 Osterix 0.01506 0.01184 0.01740 0.00692 RUNX2 0.02410 0.02105 0.02667 0.00785

In the first mesenchymal stem cells, the differentiation factors related to osteoblast formation were increased similarly to C3H10T1 / 2 cells. In particular, in the dichloromethane fraction (DCM) of Preparation Example 2, the reed root ethanol extract of Preparation Example 1 (Et- OH, mRNA expression levels of ALP, OCN, Osterix and RUNX2 were all significantly increased.

Experimental Example 3: ovariectomy Test using an animal model of osteoporosis

To investigate the effect of reed root ethanol extract (Et-OH) on the treatment and prevention of osteoporosis in Preparation Example 1, bone mineral density was measured and histological analysis was performed after administration of the extract to ovariectomized rats.

(1) Animal breeding and ovariectomy

11-week-old female SD rats were purchased from BioLink for a week of purifying period. At 12 weeks of age, ovariectomies were performed and he had a one week recovery period. Experimental animals were housed in a single cage. The environmental conditions were adjusted to room temperature 24 ± 2 ℃, relative humidity 55 ± 10%, and illumination time 12 hours. Feed and water were freely available.

The experimental animals were divided into three groups. Ten experimental ovariectomized group (Sham,)), 10 ovariectomized group (Ovx,)), 10 ovariectomized, reed roots ethanol extract of preparation 1 after ovariectomy (Ovx + Sample 100 mg / kg, ▲) in the experimental group (Et-OH). Ovariectomy was performed by using anesthesia with Zoletil and rumpun, and then depilating and incising the skin, removing the ovaries and suturing. The test samples were orally administered for a total of 8 weeks.

As a result, the ovariectomized group (Ovx, ■) showed a tendency to increase in body weight compared with the ovariectomized group (Sham, ◆) due to the decrease of estrogen secretion due to ovariectomy. (Ovx + Sample 100 mg / kg, ▴) treated with ethanol extract of black rice (Et-OH) showed a tendency to decrease in body weight compared to the ovariectomized group (OVX) .

(2) Measurement of BMD

When the animal was 19 weeks of age, the femur and tibia were separated and the femur was used to measure bone density. BMD was measured using pDEXA (Forearm: X-Ray, NORLAND, Bone Densitometer, USA).

As a result, the BMD of the ovariectomized group (OVX) was significantly lower than that of the non-ovariectomized group (Sham). After the ovariectomy, the reed root ethanol extract of Preparation Example 1 (Et- (BMD) of the experimental group (Ovx + Sample 100 mg / kg) administered with the ovariectomized group (OVX) was significantly higher than that of the ovariectomized group (OVX) (FIG.

(3) Measurement of osteoblast differentiation factor expression level

(Ovx + Sample) to which the ovariectomized experimental group (Sham), the ovariectomized experimental group (OVX) and the ovariectomized ovariectomized, and the reed root ethanol extract (Et-OH) And mRNA expression of ALP, OCN, Osterix and RUNX2, which are the differentiation factors involved in osteoblast formation, was confirmed by real-time PCR, and the results are shown in FIGS. 5 and 5 . The differentiation factor is in vivo The results were as follows.

division Sham OVX OVX + Sample ALP 0.0694 + 0.0042 0.0490 0.0048 0.0608 0.0066 OCN 0.3712 + 0.0190 0.3134 + 0.0114 0.3720 + 0.0147 Osterix 0.0547 0.0030 0.0404 0.0062 0.0519 ± 0.0037 RUNX2 0.0244 0.0009 0.0206 ± 0.0016 0.0237 0.0009

The administration of the reed root ethanol extract of Preparation Example 1 increased the expression levels of ALP, OCN, Osterix and RUNX2 mRNA expression levels, which are related to osteoblast-induced osteoblast differentiation, to levels similar to those of the ovariectomized experimental group.

(4) H & E staining

The tibia isolated from each animal was fixed in 10% formaldehyde and decalcified to produce paraffin blocks. Hematoxylin & eosin (H & E) was performed by removing the paraffin block with a thickness of 5 ㎛.

As a result, it was observed that the amount of adipocyte in the bone marrow was increased in the ovariectomized group (Ovx + control) than in the ovariectomized group (Sham) (Ovx + Sample 100mg / kg) treated with reed roots ethanol extract (Et-OH) in the ovariectomized group (ovx + control) (Fig. 6).

Hereinafter, formulation examples of the composition containing the extract of the present invention will be described, but the present invention is not intended to be limited thereto but is specifically described.

Preparation Example 1. Preparation of powder

20 mg of the extract of Preparation Example 1

Lactose 100 mg

Talc 10 mg

The above components are mixed and filled in airtight bags to prepare powders.

Formulation Example 2. Preparation of tablets

10 mg of the extract of Preparation Example 1

Corn starch 100 mg

Lactose 100 mg

Magnesium stearate 2 mg

After mixing the above components, tablets are prepared by tableting according to the usual preparation method of tablets.

Formulation Example 3. Preparation of capsules

10 mg of the extract of Preparation Example 1

Crystalline cellulose 3 mg

Lactose 14.8 mg

Magnesium stearate 0.2 mg

The above components are mixed according to a conventional capsule preparation method and filled in gelatin capsules to prepare capsules.

Formulation Example 4. Preparation of injection

10 mg of the extract of Preparation Example 1

180 mg mannitol

Sterile sterilized water for injection 2974 mg

Na ₂ HPO _4, 12H ₂ O 26 mg

(2 ml) per 1 ampoule according to the usual injection preparation method.

Formulation Example 5. Preparation of a liquid preparation

20 mg of the extract of Preparation Example 1

10 g per isomer

5 g mannitol

Purified water quantity

Each component was added to purified water in accordance with the usual liquid preparation method and dissolved, and the lemon flavor was added in an appropriate amount. Then, the above components were mixed, and purified water was added thereto. The whole was adjusted to 100 ml with purified water, And sterilized to prepare a liquid preparation.

Formulation Example 6. Preparation of Healthy Foods

1,000 mg of the extract of Preparation Example 1

Vitamin mixture quantity

70 [mu] g of vitamin A acetate

Vitamin E 1.0 mg

0.13 mg vitamin B1

0.15 mg of vitamin B2

0.5 mg vitamin B6

0.2 [mu] g vitamin B12

10 mg vitamin C

Biotin 10 μg

Nicotinic acid amide 1.7 mg

50 ㎍ of folic acid

Calcium pantothenate 0.5 mg

Mineral mixture quantity

1.75 mg of ferrous sulfate

0.82 mg of zinc oxide

Magnesium carbonate 25.3 mg

15 mg of potassium phosphate monobasic

Secondary calcium phosphate 55 mg

Potassium citrate 90 mg

100 mg of calcium carbonate

24.8 mg of magnesium chloride

Although the composition ratio of the above-mentioned vitamin and mineral mixture is comparatively mixed with a composition suitable for health food as a preferred embodiment, the compounding ratio may be arbitrarily modified, and the above ingredients are mixed according to a conventional method for producing healthy foods , Granules can be prepared and used in the manufacture of health food compositions according to conventional methods.

Formulation Example 7. Preparation of health drink

1,000 mg of the extract of Preparation Example 1

Citric acid 1,000 mg

100 g of oligosaccharide

Plum concentrate 2 g

Taurine 1 g

Purified water was added to a total of 900 ml

The above components were mixed according to a conventional health drink manufacturing method, and the mixture was heated at 85 DEG C for about 1 hour with stirring, and the solution thus prepared was filtered to obtain a sterilized 2-liter container, which was sealed and sterilized, ≪ / RTI >

Although the composition ratio is a mixture of the components suitable for the preferred beverage as a preferred embodiment, the blending ratio may be arbitrarily varied according to the regional and national preferences such as the demand level, the demanding country, and the intended use.

Claims (9)

A method for the prevention or treatment of bone metabolic diseases selected from osteoporosis, osteoarthritis, and fracture, which comprises a reed root extract as an active ingredient.
The method according to claim 1, wherein the reed root extract is selected from the group consisting of reeds ( Phragmites communis Trin.), Phragmites japonica Steud., Phragmites karka (Retz. Tr. Ex Steud.), Wherein the pharmaceutical composition is an extract of at least one plant selected from the group consisting of plant extracts and plant extracts.
delete delete The pharmaceutical composition for prevention or treatment of bone metabolic diseases according to claim 1, wherein the reed root extract is an extract of water, an organic solvent or a mixed solvent thereof.
[Claim 6] The pharmaceutical composition according to claim 5, wherein the reed root extract is a fraction obtained by fractionating the extract with water, an organic solvent or a mixed solvent thereof.
The method according to claim 5 or 6, wherein the organic solvent is at least one solvent selected from the group consisting of alcohols having 1 to 4 carbon atoms, hexane, dichloromethane, ethyl acetate, acetone, chloroform and diethyl ether A pharmaceutical composition for preventing or treating bone metabolic diseases.
[Claim 6] The method according to claim 5, wherein the extract is obtained by stirring, extracting with hot water, refluxing, refluxing cooling, ultrasonic extraction, or supercritical extraction using an extraction solvent of 1 to 50 times the weight of dried reed root Or a pharmaceutically acceptable salt thereof.
A health functional food for prevention or improvement of bone metabolic diseases selected from osteoporosis, osteoarthritis, and fracture containing an extract of reed root as an active ingredient.

Images