KR102527480B1 - Composition for preventing, improving or treating osteoporosis comprising camellia leaf extract - Google Patents

Abstract

The present invention relates to a composition for preventing, alleviating or treating osteoporosis, and more specifically, to a composition for preventing, alleviating or treating osteoporosis containing a Camellia japonica leaf extract, and a method for manufacturing the composition. The composition according to the present invention contains the Camellia japonica leaf extract, so that it is possible to provide the composition with excellent effects for preventing, alleviating, or treating osteoporosis, and a method for manufacturing the composition.

Description

Composition for preventing, improving or treating osteoporosis comprising camellia leaf extract

The present invention relates to a composition for preventing, improving or treating osteoporosis, and more specifically, to a composition for preventing, improving or treating osteoporosis comprising a camellia leaf extract and a method for preparing the composition.

Camellia, the same genus as tea, has been mainly used as a horticultural resource such as ornamental. By part, the seeds containing high-quality fat have traditionally been used as a resource for cooking oil and cosmetic oil, and the stem is used as a raw material for high-quality charcoal. However, it has not yet been used as a food or medicinal resource, and only some research is being conducted. Camellia grows in the beaches and islands of Ulleungdo, Jeju Island, and southern regions of Korea. The flowering period is December to May, and the fruiting period is September to October. Dried camellia flowers used as medicine are flexible and have a good scent. Those with red color, no mold, and large buds that have not yet bloomed have the highest grade. The taste and properties of sandalwood are sweet, slightly spicy and cool. As for the efficacy of camellia flower, it is known that it enters liver cirrhosis and menopause, and is effective for yang blood, hemostasis, and acid fish blood.

On the other hand, osteoporosis, rather than the symptom itself, is easily caused by bone weakness, and various fractures, especially femur fractures or vertebral fractures, restrict activity for a long time and cannot lead a healthy life, resulting in about 15% of deaths among the elderly. It is known to provide a cause. Bone mass is influenced by many factors, including genetic factors, nutritional intake, hormonal changes, and differences in exercise and lifestyle. Ovariectomy and the like are known.

Although there are individual differences, blacks have higher bone mass than whites because their bone resorption level is lower. Bone mass usually peaks between the ages of 14 and 18 and decreases by about 1% per year in old age. In particular, in the case of women, bone loss continues to progress after the age of 30, and bone loss rapidly progresses due to hormonal changes when reaching menopause. As described above, osteoporosis is an unavoidable symptom in the elderly, especially postmenopausal women, although the degree varies. As the population ages in developed countries, interest in osteoporosis and its treatment is gradually increasing, but camellia, camellia flower, and camellia leaf There is no study on the prevention or treatment of osteoporosis using the back.

Registered Patent Publication No. 10-1838354 Registered Patent Publication No. 10-0702327

In order to solve the above problems, the present invention is to provide a composition for preventing, improving or treating osteoporosis containing a camellia leaf extract.

In addition, it is intended to provide a method for preparing the camellia leaf extract and a method for preparing a composition for preventing, improving or treating osteoporosis containing the camellia leaf extract.

In order to achieve the above object, the present invention provides a composition containing a camellia leaf extract as an active ingredient and excellent in preventing, improving, and treating osteoporosis.

The camellia leaf extract according to the present invention can be prepared through a manufacturing method comprising the following steps:

(a) Camellia leaf preparation step;

(b) pre-treating the dried camellia leaves;

(c) extracting the pretreated camellia leaves.

The (a) camellia leaf preparation step is a step of preparing camellia leaves to be used for extraction, and fresh or dried leaves may be used as the camellia leaves. When dried leaves are used, they are dried at 25 to 35°C, preferably at 30°C for 2 to 7 days, preferably 3 to 5 days. If the temperature and period are less than the above ranges, the camellia leaves are not dried properly, and if the temperature and period are exceeded, the leaves are too dry, and the leaves may be broken in the subsequent steaming and roasting steps.

The camellia leaves are not limited, and adult leaves or young leaves may be used. When using a young leaf, it may preferably be a young leaf of 1 cm or more and less than 18 cm, and when using a young leaf, the effect of preventing, improving or treating osteoporosis may be further increased.

Step (b) is a step of pre-processing the prepared camellia leaves by performing one or more of the following: mindfulness of rubbing, roasting and rubbing, steaming of leaves, and steaming/steaming of steaming and rubbing. The roasting step is 180 to 260 ° C, preferably 200 to 240 ° C for 1 to 10 minutes, preferably 3 to 5 minutes, and the boiled camellia leaves are cooled at room temperature (20 to 25 ° C) for 3 to 5 minutes. give. At this time, it should be cooled in the shade without sunlight.

The step of steaming the camellia leaves is steamed at 90 to 110 ° C, preferably 100 ° C for 10 to 120 minutes, preferably 30 to 60 minutes. Outside the above temperature range and time, the effect of preventing, improving or treating osteoporosis may be reduced.

The steaming/roasting process may be performed under the conditions of the steaming process and the roasting process, respectively.

Step (c) is a step of obtaining a camellia leaf extract by extracting the camellia leaves pretreated in step (b).

The solvent used for the extract is not limited, but may preferably be extracted using water or purified water as a solvent.

In addition, the extract may be obtained by extracting camellia leaves at 60 to 80° C. for 3 to 10 minutes, preferably 5 to 10 minutes, and more preferably 5 minutes. When extracted under conditions other than the above temperature and time, active ingredients may not be sufficiently extracted.

The present invention also provides a composition for preventing, improving or treating osteoporosis comprising a camellia leaf extract.

The composition for preventing, improving or treating osteoporosis may be provided as a pharmaceutical composition.

Alternatively, when the composition is provided as a food composition, it is provided as a food composition for preventing or improving osteoporosis.

In the pharmaceutical composition or food composition of the present invention, the camellia leaf extract according to the present invention may be included in a concentration of greater than 0, preferably 10ug/mL or more, but is not limited thereto.

In addition, the pharmaceutical composition may further include a pharmaceutically acceptable carrier, binder, lubricant, disintegrant, excipient, solubilizer, dispersant, stabilizer, suspending agent, etc., for example, lactose, dex Trose, sucrose, sorbitol, mannitol, xylitol, erythritol, malditol, starch, alginate, calcium phosphate, calcium silicate, maltodextrin, silicon dioxide, cellulose, methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, methylhydride It may be hydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, etc., but is not limited thereto.

In addition, the pharmaceutical composition may be prepared in a formulation such as a tablet, capsule, sustained-release preparation, elixir, suspension, syrup, wafer, unit dose injection ampoule, suspension, etc. by a method commonly used for formulation in the art.

The administration method of the pharmaceutical composition is oral administration, intravenous administration, intramuscular administration, intraarterial administration, intramedullary administration, intrathecal administration, intracardiac administration, transdermal administration, subcutaneous administration, intraperitoneal administration, intranasal administration, enteral administration , It may be administered by methods such as topical administration, rectal administration, topical administration, dermal administration, mucosal administration, intravertebral administration, including oral and sublingual administration, preferably oral administration, but is not limited thereto. In addition, at the time of administration, it may be formulated and administered in a formulation suitable for each administration method, and the formulation may be formulated by a method widely known in the art.

The pharmaceutical composition may be appropriately selected according to the age, weight, health condition, sex, administration time, administration route, excretion rate, active ingredient content, severity of a specific disease to be prevented or treated, etc., and the number of administrations is 1 It may be administered once or twice a day, every other day, every other week, or every other month.

In addition, the food composition may additionally include flavoring agents, flavoring agents, stabilizers, thickeners, preservatives, etc. commonly used in the art to prepare food, for example, lactose, dextrose, water chlorose, sorbitol, mannitol, xylitol, erythritol, malditol, starch, alginate, calcium phosphate, calcium silicate, maltodextrin, silicon dioxide, cellulose, methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, methylhydroxybenzoate, It may be propyl hydroxybenzoate, talc, magnesium stearate, etc., but is not limited thereto.

The food composition may be prepared in the form of beverages, gum, tea, vitamin complexes, powders, granules, tablets, capsules, confectionery, and the like.

The food composition may be appropriately selected according to the age, weight, health condition, sex, administration time, content of the active ingredient, severity of the disease, etc. of the subject to be consumed, and the number of intakes is once or twice a day, every other day, You can take it every other week, every other month, etc.

The present invention can provide a composition containing a camellia leaf extract that has excellent effects on preventing, improving or treating osteoporosis, and the method for preparing the camellia leaf extract and the composition for preventing, improving or treating osteoporosis containing the camellia leaf extract A manufacturing method can be provided.

1 shows the results of evaluating the osteoblast differentiation improvement efficacy according to the concentration of the young camellia leaf extract prepared according to the present invention.
Figure 2 shows the results of osteoblast differentiation improvement efficacy evaluation according to the concentration of the young camellia leaf extract prepared according to the present invention.
Figure 3 shows the osteoblast differentiation improvement efficacy evaluation results according to the concentration of the camellia seongyeop extract prepared according to the present invention.
Figure 4 shows the results of the evaluation of the osteoblast differentiation improvement efficacy over time of the extract of young camellia steamed / boiled leaves prepared according to the present invention.

Hereinafter, the present invention will be described in detail by examples and experimental examples.

However, the following Examples and Experimental Examples are merely illustrative of the present invention, and the contents of the present invention are not limited to the following Examples and Experimental Examples.

<Examples 1 to 3> Preparation of fresh camellia leaf extract according to the present invention

Fresh leaves were used to prepare camellia leaf tea, and among the used camellia leaves, young leaves were collected based on newly grown less than 18 cm. Saengyeop (saengsinyeop) was prepared under the conditions of roasting and rubbing (steaming), steaming conditions (steaming), and steaming and rubbing conditions (steaming/steaming). For mindfulness, camellia leaves were stacked on a cotton cloth and rubbed, and for 3 minutes at 200℃~230℃ for 3 minutes, the cooling cycle was repeated twice. The steam was steamed at 100 ° C for 30 minutes and dried, and the steam / roast was steamed at 100 ° C for 30 minutes, cooled, then steamed at 200 ° C to 230 ° C for 3 minutes, and the cooling cycle was repeated twice.

1 g of each camellia leaf obtained by the above process was put in a tea rim bag, soaked in water at about 60 ° C to 80 ° C for 5 minutes, and then the tea rim bag was removed. After the camellia leaf extract was centrifuged, the supernatant was filtered using a 0.45 μm syringe filter, stored in a -70 ° C ultra-low temperature freezer for 24 hours, and then freeze-dried. Freeze-dried samples were stored at room temperature.

<Examples 4 to 7> Preparation of dried camellia leaf extract according to the present invention

Preparation of dried camellia leaf extract according to the present invention Newly grown young camellia leaves of less than 18 cm were collected, washed for drying, and then dried at 30 ° C. for 5 days.

As in Examples 1 to 3, the dried new leaves were prepared under the condition of only rubbing (mind), the condition of rubbing and rubbing (steaming), the condition of steaming (steaming), and the condition of steaming, rubbing and rubbing (steaming/steaming). . For mindfulness, camellia leaves were stacked on a cotton cloth and rubbed, and for 3 minutes at 200℃~230℃ for 3 minutes, the cooling cycle was repeated twice. The steam was steamed at 100 ° C for 30 minutes and dried, and the steam / roast was steamed at 100 ° C for 30 minutes, cooled, then steamed at 200 ° C to 230 ° C for 3 minutes, and the cooling cycle was repeated twice.

1 g of each camellia leaf obtained by the above process was put in a tea rinsing bag, soaked in water at about 60 to 80 ° C for 5 minutes, and then the tea rinsing bag was removed. After the camellia leaf extract was centrifuged, the supernatant was filtered using a 0.45 μm syringe filter, stored in a -70 ° C ultra-low temperature freezer for 24 hours, and then freeze-dried. Freeze-dried samples were stored at room temperature.

<Examples 8 to 10> Preparation of camellia sage leaf extract according to the present invention

It was prepared in the same manner as in Examples 1 to 3, but the used camellia leaves were adult leaves, not young leaves.

<Examples 11 to 14> Preparation 2 of dried camellia sex leaf extract according to the present invention

It was prepared in the same way as in Examples 4 to 7, but the camellia leaves used were adult leaves collected in May, not young leaves.

<Examples 15 to 18> Preparation 3 of dried camellia sex leaf extract according to the present invention

It was prepared in the same way as in Examples 4 to 7, but the camellia leaves used were adult leaves collected in September, not young leaves.

<Experimental Example 1> Evaluation of the osteoblast differentiation improvement effect according to the concentration of camellia young leaf tea 1

The concentrations of the extracts of Examples 1 to 7 were set to 10 μg/mL and 20 μg/mL, respectively, and the efficacy of improving osteoblast differentiation was evaluated using this.

In order to analyze the degree of osteoblast differentiation, an ALP staining kit, TRACP & ALP double-stain kit (MK300, TaKaRa) was used. After 6 days, the culture medium was removed, washed with PBS, and 200 μl of the fixative was added to each well, followed by reaction for 5 minutes to fix. After washing twice with 2ml of sterile water, 200μl of ALP substrate was added and reacted at 37℃ for 30 minutes.

The evaluation results after 6 days of differentiation are shown in FIG. 1, and the darker the staining, the higher the ALP activity and the better the differentiation into osteoblasts.

As a result of the evaluation, it was confirmed that when MG-63 cells, which are osteoblast-like cells, were differentiated and treated with the camellia young leaf extract, respectively, osteoblast differentiation was better than that of the differentiation culture medium when the extract was treated. In particular, the steamed/roasted camellia leaf extract induced differentiation the best compared to other extracts. Through these results, it is judged that the camellia leaf extract can prevent, improve or treat osteoporosis.

<Experimental Example 2> Evaluation of the osteoblast differentiation improvement effect according to the concentration of camellia young leaf tea 2

Concentrations of the extracts of Examples 4 to 7 were set to 10 μg/mL and 20 μg/mL, respectively, and the efficacy of improving osteoblast differentiation was evaluated using this.

In order to analyze the degree of osteoblast differentiation, ALP activity was performed using TRACP & ALP Assay Kit (MK301, TaKaRa). After 6 days, the culture medium was removed, washed with PBS, and 50 μl of the cell lysate was added to each well to lyse the cells. After adding 40 μl of p -nitro-phenyl phosphate substrate to 40 μl of the cell lysate and reacting at 37° C. for 1 hour, 40 μl of 0.5N NaOH was added to stop the reaction, and the absorbance was measured at 405 nm.

The evaluation results after 6 days of differentiation are shown in Figure 2, * p <0.05, ** p <0.01.

As a result of the evaluation, when MG-63 cells, which are osteoblast-like cells, were differentiated and treated with camellia young leaf extract, respectively, ALP activity was higher than that of the differentiation culture medium when the extract was treated, confirming that osteoblast differentiation was better. . However, unlike the darkest ALP staining in the steamed/roasted extract, there was no difference in ALP activity between the steamed/roasted extract and the steamed/roasted extract, and all of the extracts are considered to be capable of preventing, improving or treating osteoporosis.

<Experimental Example 3> Evaluation of the osteoblast differentiation improvement effect according to the concentration of camellia seongyeop tea

The concentrations of the extracts of Examples 8 to 18 were set to 10 μg/mL and 20 μg/mL, respectively, and the efficacy of improving osteoblast differentiation was evaluated using this.

In order to analyze the degree of osteoblast differentiation, an ALP staining kit, TRACP & ALP double-stain kit (MK300, TaKaRa) was used. After 6 days, the culture medium was removed, washed with PBS, and 200 μl of the fixative was added to each well, followed by reaction for 5 minutes to fix. After washing twice with 2ml of sterile water, 200μl of ALP substrate was added and reacted at 37℃ for 30 minutes.

The evaluation results after 6 days of differentiation are shown in FIG. 3, and the darker the staining, the higher the ALP activity and the better the differentiation into osteoblasts.

As a result of the evaluation, when MG-63 cells, which are osteoblast-like cells, were differentiated and treated with the camellia seongyeop extract, respectively, it was confirmed that the differentiation of osteoblasts was better than that of the differentiated culture medium when the extract was treated. Unlike the young leaf extract, the steamed extract induced the best differentiation compared to other extracts. Through these results, it is judged that the camellia leaf extract has an excellent effect in preventing, improving or treating osteoporosis regardless of the treatment method.

<Experimental Example 4> Evaluation of osteoblast differentiation improvement effect according to steaming / roasting treatment of camellia young leaves

Among camellia young leaf extracts, the ability to improve differentiation over time was confirmed using the steamed/roasted extract of dried leaves, which had the highest ability to improve osteoblast differentiation (Example 7).

In order to analyze the degree of osteoblast differentiation, an ALP staining kit, TRACP & ALP double-stain kit (MK300, TaKaRa) was used. After 6 days, the culture medium was removed, washed with PBS, and 200 μl of the fixative was added to each well, followed by reaction for 5 minutes to fix. After washing twice with 2ml of sterile water, 200μl of ALP substrate was added and reacted at 37℃ for 30 minutes.

The darker the staining, the higher the ALP activity, and the evaluation results after 3 days and 6 days of differentiation are shown in FIG. 4 .

As a result of the evaluation, it was confirmed that osteoblast differentiation is better with time than in the differentiation culture medium when the extract is treated, and as a result, it is determined that osteoporosis can be prevented, improved, or treated using the camellia leaf extract.

Summarizing the above experimental results, all camellia leaf extracts according to the present invention proved to be effective in preventing and improving osteoporosis. , In particular, it was proved that steamed/roasted extracts may be more effective in preventing, improving, or treating osteoporosis.

Claims (8)

A pharmaceutical composition for preventing, improving or treating osteoporosis comprising a camellia leaf extract as an active ingredient,
The camellia leaves are pretreated by steaming / roasting dried young leaves of 1 cm or more and less than 18 cm,
A pharmaceutical composition for the prevention, improvement or treatment of osteoporosis, characterized by roasting at 200 to 240 ° C for 1 to 10 minutes, and steaming at 90 to 110 ° C for 30 to 60 minutes. A food composition for preventing or improving osteoporosis comprising a camellia leaf extract as an active ingredient,
The camellia leaves are pretreated by steaming / roasting dried young leaves of 1 cm or more and less than 18 cm,
Roasting is to roast at 200 to 240 ° C for 1 to 10 minutes, and steaming is a food composition for preventing or improving osteoporosis, characterized in that steaming at 90 to 110 ° C for 30 to 60 minutes. delete The composition according to claim 1 or 2, wherein the camellia leaf extract is extracted using water as a solvent. A method for producing the camellia leaf extract of claim 1 or 2, comprising the following steps:
(a) Camellia leaf preparation step;
(b) pre-treating the dried camellia leaves by steaming/roasting;
(c) extracting the pretreated camellia leaves;
The camellia leaves are dried young leaves of 1 cm or more and less than 18 cm,
The method of producing a camellia leaf extract, characterized in that the roasting is roasting at 200 to 240 ° C for 1 to 10 minutes, and the steaming is steaming at 90 to 110 ° C for 30 to 60 minutes. delete The method of claim 5, wherein in step (c), the camellia leaves are extracted for 5 to 10 minutes at 60 to 80° C. using water as a solvent. delete

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