KR102611924B1 - Composition comprising syneilesis aconitifolia (bunge) maxim extract thereof for preventing and treating of periodontitis - Google Patents

Abstract

The present invention relates to a composition for the prevention and treatment of periodontal disease containing an extract of Elephant chinensis as an active ingredient, which prevents inflammation by inhibiting the release of tumor necrosis factor-alpha (TNF-α) and proliferation and differentiation of osteoblasts. Providing a composition for the prevention and treatment of periodontal disease containing as an active ingredient the extract of Achyranthes ginseng, which promotes the formation and activity of osteoclasts, inhibits the destruction of alveolar bone, and increases the proliferation and cell activity of periodontal ligament cells. can do.

Description

Composition for preventing and treating periodontal disease containing extract of Acronisum ginseng as an active ingredient

The present invention relates to a composition for the prevention and treatment of periodontal disease containing an extract of Elephant chinensis as an active ingredient, which prevents inflammation by inhibiting the release of tumor necrosis factor-alpha (TNF-α) and proliferation and differentiation of osteoblasts. Providing a composition for the prevention and treatment of periodontal disease containing as an active ingredient the extract of Achyranthes ginseng, which promotes the formation and activity of osteoclasts, inhibits the destruction of alveolar bone, and increases the proliferation and cell activity of periodontal ligament cells. can do.

Periodontal tissue consists of the alveolar bone, gingiva (gum), and periodontal ligament. The gums are part of the tooth supporting tissue that we can see in the mouth, and this is where the disease (gingivitis) begins. As the disease progresses and spreads deeper into the supporting tissue, the periodontal ligament, which is attached to the root surface of the tooth and the bone tissue surrounding the tooth, As the alveolar bone surrounding the teeth is destroyed, periodontal disease occurs.

Periodontal disease, including gingivitis and periodontitis, is an inflammatory condition of tooth-supporting tissues caused by bacteria and is a disease that results in tooth loss due to bleeding, formation of periodontal pockets, and destruction of alveolar bone. This periodontal disease progresses through the process of bacterial colonization, bacterial invasion of periodontal tissue, and destruction of periodontal tissue. Specifically, when oral bacteria colonize due to poor oral hygiene, a bacterial film is formed, causing inflammation, sometimes bleeding gums, and bad breath. If this condition progresses further, the gap between the teeth and gums becomes deeper, forming periodontal pockets. Bacteria that cause periodontal disease multiply here, resulting in periodontitis. As periodontitis progresses, gums bleed and swell even with mild stimulation, such as toothbrushing, and often turn into acute inflammation, causing pain. This inflammation reduces the function of bone formation and increases the function of bone absorption, causing the alveolar bone to gradually disappear and be destroyed, ultimately leading to tooth loss.

There are many causes of periodontal disease, but when plaque accumulates in the periodontal pockets due to local factors, it becomes a habitat for anaerobic gram-negative bacteria existing in the surrounding area and proliferates deep into the periodontal pockets. At this time, the toxins and all products of the proliferated anaerobic Gram-negative bacteria directly destroy the tissue or stimulate the immune system, causing destruction of periodontal tissue and inflammation through various actions of the stimulated immune system. As a defense mechanism against this, the function of polymorphonuclear leukocytes and immune response act as systemic factors.

In other words, periodontal tissue destruction and inflammation are caused by cytotoxins such as hydrogen sulfide, ammonia, and toxic amines, which are metabolites of anaerobic Gram-negative bacteria, and endotoxins such as lipopolysaccharide, a cell wall component of anaerobic Gram-negative bacteria. Active oxygen, prostaglandin, leukotrien, histamine, and interleuckin secreted outside the cell by the action of the humoral and cellular immune system stimulated by direct tissue destruction or stimulation of the biological immune system. It is caused by various cytokines such as ). In addition, collagen, the matrix of periodontal tissue, is broken down by enzymes such as collagenase secreted from bacteria and white blood cells, causing gum recession, and if left untreated, periodontal disease progresses.

Therefore, antibacterial action against anaerobic Gram-negative bacteria, which are the fundamental causative factors of periodontal disease, removal of bacterial toxic products, and restoration of lost periodontal tissue are important keys in the prevention and treatment of periodontal disease.

Currently, the treatment of periodontal disease includes improving the patient's oral hygiene, non-surgical treatment, and surgical treatment (calculus removal, root planing, gingival curettage, periodontal tissue regeneration using new attachment). The surgical treatment method, which is the most effective treatment, is not effective in treating periodontal disease due to the inconvenience of having to go to the dentist for treatment and the limitation that it is performed when the disease has progressed to a certain extent rather than preventing the disease. Periodontal disease is mostly a chronic disease. It proceeds as follows. Systemic antibiotics and topical sustained-release preparations have been used as additional treatments, but side effects are caused by too much drug being delivered to unnecessary areas, and cases of isolation of periodontal disease bacteria that are resistant to recently used antibiotics have been reported. It has serious problems.

In order to compensate for the limitations of surgical treatment and the problems of antibiotic use and to improve the effectiveness of periodontal disease prevention and treatment, there is an urgent need for the development of an agent that has the effect of restoring destroyed and lost periodontal tissue to its original state.

KR 10-2017-0162217 A

The object of the present invention is to provide a composition for preventing and treating periodontal disease containing an extract of Elephant chinensis as an active ingredient, which prevents inflammation and proliferation of osteoblasts by inhibiting the release of tumor necrosis factor-alpha (TNF-α). and a composition for the prevention and treatment of periodontal disease containing as an active ingredient an extract of Aedum ginseng, which promotes differentiation, reduces the formation and activity of osteoclasts, inhibits the destruction of alveolar bone, and increases the proliferation and cell activity of periodontal ligament cells. can be provided.

A composition for preventing and treating periodontal disease according to an embodiment of the present invention includes Syneilesis aconitifolia (Bunge) Maxim extract as an active ingredient.

The extract is extracted using an extraction solvent selected from the group consisting of water, C ₁ to C ₆ lower alcohols, and mixtures thereof.

The periodontal disease includes diseases selected from the group consisting of gingivitis, periodontitis, and mixtures thereof.

The composition shows the effect of preventing and improving dry mouth.

The dry mouth is a group consisting of acid erosion, oral candidiasis, dysgeusia, dysphagia, oral dysesthesia, intraoral halitosis, and a combination thereof. Includes diseases selected from.

A toothpaste for preventing periodontal disease according to another embodiment of the present invention includes the composition for preventing and treating periodontal disease.

A mouthwash for preventing periodontal disease according to another embodiment of the present invention includes the composition for preventing and treating periodontal disease.

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

The term “prevention” used in the present invention refers to any action that suppresses the symptoms or delays the progression of a specific disease by administering the composition of the present invention.

The term “treatment” used in the present invention refers to any action that improves or beneficially changes the symptoms of a specific disease by administering the composition of the present invention.

As used herein, the term “improvement” refers to any action that reduces at least the severity of a parameter, such as a symptom, related to the condition being treated.

A composition for preventing and treating periodontal disease according to an embodiment of the present invention includes Syneilesis aconitifolia (Bunge) Maxim extract as an active ingredient.

Syneilesis aconitifolia (Bunge) Maxim is a perennial plant of the Asteraceae family with a height of 70-120 cm. The roots have short rhizomes that extend to the sides, and the main stem is purple and has no branches and has two leaves. The first leaf is round, 20-30cm in diameter, divided into long shapes, has 7-9 lobes, and is divided into two rows 2-3 times. The second leaf is slightly smaller, 12-24cm in diameter, the lobe is 4-5cm wide, and the petiole is 2-6cm long. The leaf is round, 20-30cm in diameter, divided into long shapes, and flowers bloom from July to August.

The brachyurysis extract is extracted using an extraction solvent selected from the group consisting of water, C ₁ to C ₆ lower alcohols, and mixtures thereof.

Specifically, in order to produce an extract of the vegetable extract, the steps include washing the vegetable extract; Drying after washing; Grinding the baby umbrella greens after drying; leaching the pulverized material using an organic solvent; Leaching and drying the sample; Leaching using water; And, including the step of leaching, an extract of Aerobium chinensis can be obtained.

The extract extracted using the organic solvent may further include the step of performing fractionation using the organic solvent, and the method of preparing the extract may be a conventional extraction method in the art, such as ultrasonic extraction, leaching, and reflux extraction. You can.

Specifically, it may be an extract obtained by extracting a Papola shoot extract from which foreign substances have been removed by washing and drying with water, alcohol having 1 to 6 carbon atoms, or a mixed solvent thereof, and may be an extract obtained by sequentially applying the solvents to the sample.

The reflux extraction method is based on 100 mL of water and alcohol with 1 to 6 carbon atoms, 10 to 30 g of pulverized natural product, reflux time of 1 to 3 hours, and 50 to 100% of alcohol or water with 1 to 6 carbon atoms. More specifically, based on 100 mL of alcohol with 1 to 6 carbon atoms or 100 mL of water, 10 to 20 g of pulverized natural product, reflux time of 1 to 2 hours, and 70 to 90% of alcohol or water with 1 to 4 carbon atoms. .

The leaching method is carried out at 15 to 30°C for 24 to 72 hours, and water or 50 to 100% alcohol with 1 to 6 carbon atoms is used as the extraction solvent. More specifically, the extraction is performed at 20 to 25°C for 30 to 54 hours, and the extraction solvent is water or 70 to 80% alcohol with 1 to 6 carbon atoms.

The ultrasonic extraction method proceeds at 30 to 50°C for 0.5 to 2.5 hours, and the extraction solvent is water or 50 to 100% alcohol with 1 to 6 carbon atoms. Specifically, extraction is performed at 40 to 50°C for 1 to 2.5 hours, and the extraction solvent is water or 70 to 80% alcohol with 1 to 6 carbon atoms.

The extraction solvent can be used in an amount of 2 to 50 times, more specifically, 2 to 20 times the weight of the sample. For extraction, the sample may be left in the extraction solvent for 1 to 72 hours, more specifically, 24 to 48 hours.

After extraction, the extract can be fractionated by sequentially applying a new fractionation solvent. The fractionating solvent used during fractionation is at least one selected from the group consisting of water, hexane, butanol, ethyl acetic acid, ethyl acetate, methylene chloride, and mixtures thereof, and is preferably ethyl acetate or methylene chloride. After obtaining the extract or fraction, additional methods such as concentration or freeze-drying can be used.

The periodontal disease includes diseases selected from the group consisting of gingivitis, periodontitis, and mixtures thereof.

Periodontal disease is divided into gingivitis and periodontitis depending on the severity of the disease. It is a relatively mild and fast-recoverable form of periodontal disease. When it is limited to the gums, or soft tissue, it is called gingivitis. When this inflammation progresses to the area around the gums and gum bones, it is called periodontitis. Through a dental examination and periodontal examination, the condition of the teeth and gingiva can be checked to make a diagnosis of gingivitis and periodontitis, and a radiological examination can be performed to check the degree of destruction of the alveolar bone.

Gingivitis is a stage in which an inflammatory response caused by bacteria appears only in the superficial soft tissue, and periodontitis attacks the part below the gum line in the sulcus, damaging the periodontal ligament and surrounding bone tissue. As inflammation progresses and more periodontal ligament tissue is damaged, the narrow, shallow sulcus deepens and develops into a periodontal pocket. As periodontitis becomes more severe, the periodontal pockets deepen and the alveolar bone surrounding the tooth roots is lost. As the inflammation progresses, the space between the gums and teeth becomes wider and eventually the teeth become loose. If left untreated, the teeth may fall out on their own.

The most basic thing in the treatment of periodontal disease is to cleanly remove bacterial plaque and tartar to avoid creating an environment favorable for the growth of bacteria. Oral gum treatment medication does not help much in treatment. A mouthwash called chlorhexidine may be used, a special medication may be placed between the gums and teeth, and antibiotics may be used to eradicate special bacteria in the gums.

The main cytokines associated with bone loss, inflammation, and connective tissue destruction associated with periodontal disease are known to be interleukin-1 (IL-1) and tumor necrosis factor.

The composition shows the effect of preventing and improving dry mouth.

Healthy adults secrete 1 to 1.5 liters of saliva per day, but if less saliva is produced, the mouth feels dry. Alternatively, when the moisture in your mouth evaporates while breathing through your mouth, you may subjectively feel dry mouth.

If the dryness of the mouth is severe, it may become difficult to chew and swallow food and speech. Additionally, because saliva plays a role in digestion, digestive problems may occur, and eating spicy or irritating foods may cause pain in the mouth. Gingivitis and cavities are also more likely to occur.

The dry mouth is a group consisting of acid erosion, oral candidiasis, dysgeusia, dysphagia, oral dysesthesia, intraoral halitosis, and a combination thereof. Includes diseases selected from.

Xerostomia is the most common clinical symptom related to the salivary glands. Dry mouth is diagnosed when the amount of saliva secreted when not stimulated is less than 0.1 mL per minute. The cause of dry mouth is mainly the side effects of various drugs, such as antidepressant, antidiabetic, and antiallergic drugs. Dry mouth caused by natural aging occurs when the parenchymal tissue of the salivary glands decreases and is replaced by fatty tissue or the volume of the glands decreases. In addition, it may occur when the salivary glands are destroyed or blood flow to the salivary glands is blocked due to Sjögren's syndrome, an autoimmune disease, or radiation therapy. When the amount of saliva in the mouth is reduced, cavities, acid erosion, oral candidiasis, dysgeusia, dysphagia, oral dysesthesia, and intraoral halitosis occur. It is accompanied.

Preferably, the composition for preventing and treating periodontal disease of the present invention may additionally include natural extracts in addition to the extract of Aerobium chinensis.

The natural extract is selected from the group consisting of Viburnum erosum Thunb. extract, Primula PJsoana var. pubescens extract, Cryptotaenia japonica extract, and mixtures thereof.

The Viburnum erosum Thunb. is a deciduous broad-leaved shrub belonging to the genus of L. vulgaris and is commonly found in low mountain areas south of Gyeonggi-do. It is a short tree of the honeysuckle family, reaching a height of 2 to 3 m. Its white flowers in spring and red fruits in fall are beautiful, so it is often planted as a park tree. The leaves are oval in shape and grow opposite to each other. The fur on the front and back is thick, giving it a soft feel when touched. The stem is characterized by splitting into several pieces and growing in bundles, and the flowers bloom in April to May with white bisexual flowers forming a round shape at the ends of new branches. The small red bean-like fruits ripen red in September or October and are good food for birds. It grows in forests with moderate sunlight and likes soil with good moisture retention and drainage. It has very strong cold resistance, grows well in both sunny and shaded areas, and has the characteristic of growing well even in dry environments. Meanwhile, the name Lesser Pheasant Tree appears to be related to pheasants. Specifically, it is presumed that it was called a wild pheasant tree, meaning that it has fruits that pheasants in the field like, and then became a lesser pheasant tree. The young leaves can be eaten as a vegetable, and the old leaves and stems are used as a medicine for stomatitis or itching in oriental medicine.

The hairy primrose (Primula PJsoana var. pubescens) is a perennial of the primrose family of the primrose family of dicotyledonous plants. Compared to the primrose, it has more long hairs on the petioles and flower stems, and has a rhizome extending sideways without a main stem. The flowers are whole, reddish purple flowers with a diameter of 1.5 to 2.5 cm that bloom in July to August. The fruit produces long oval capsules measuring 7 to 12 mm in length. Distributed in Korea, Japan, China, etc.

Cryptotaenia japonica is an herb that grows in mountainous areas in Korea, also known as fireweed. The height is about 30 to 60 cm. The stem is erect, slightly split, hairless, and fragrant. The stem and leaves are green and the roots are thick. The leaves are alternate, have long petioles of 5 to 17 cm, and consist of three small leaves, which are characterized by a glossy appearance on the back. Meanwhile, the small leaf at the end has no petiole and is egg-shaped, 3 to 8 cm long and 2 to 6 cm wide. The end is sharp, the bottom narrows sharply, and there are sharp, irregular teeth at the edge. The small leaves on the sides are the same size and shape, have no petioles, and sometimes have irregular sawtooth edges that are shallowly divided into 2 or 3 pieces. The flowers are white or light purple and grow in compound umbels at the ends of branches from June to July. There are 1 to 4 small peduncles, either long or short, measuring 3 to 15 mm in length and standing upright. The small gun pieces are string-shaped and short. There are 5 petals, the ends of which are curved inward, 5 sepals, and 5 stamens. The fruit is a meristematic fruit that is 3 to 4 mm long, oval in shape, has no hairs, and ripens in August to September. Young leaves are edible and grown as a vegetable. Those collected in the summer are dried in the shade and used as medicine for goiter, etc.

More preferably, the composition for preventing and treating periodontal disease of the present invention comprises 10 to 20 parts by weight of Lesser Chrysanthemum extract, 10 to 20 parts by weight of Hairy Primula extract, and 10 to 20 parts by weight of Paddeuk sprouts, based on 100 parts by weight of the Euphorbia chinensis extract. It is done.

The composition for preventing and treating periodontal disease of the present invention, when the composition further includes the Euphorbiaceae extract, the Euphorbia primrose extract, and the Euphorbia chinensis extract, is superior to the tumor necrosis factor compared to the case where it contains only the Euphorbiaceae extract. -It can inhibit the release of alpha (TNF-α), promote the proliferation and differentiation of osteoblasts, and reduce the formation and activity of osteoclasts, leading to increased activity to suppress destruction of alveolar bone.

A toothpaste for preventing periodontal disease according to another embodiment of the present invention includes the composition for preventing and treating periodontal disease.

When using the composition for preventing and treating periodontal disease as a toothpaste, the composition can be added as is to the toothpaste or mixed or added with functional food or functional ingredients, and can be used appropriately according to conventional methods. The mixing amount of the active ingredient can be appropriately determined depending on the purpose of use.

A mouthwash for preventing periodontal disease according to another embodiment of the present invention includes the composition for preventing and treating periodontal disease.

The mouthwash is defined as including all compositions that can gargle the oral cavity and other materials for cleaning the oral cavity. Meanwhile, the mouthwash to achieve the purpose of the present invention should be an oil-based composition, and it is preferable not to mix surfactants or alcohols. The specific formulation is defined as including all compositions and mixing ranges that can be selected and mixed by a person with ordinary knowledge in the relevant field.

The present invention relates to a composition for the prevention and treatment of periodontal disease containing an extract of Elephant chinensis as an active ingredient, which prevents inflammation by inhibiting the release of tumor necrosis factor-alpha (TNF-α) and proliferation and differentiation of osteoblasts. Providing a composition for the prevention and treatment of periodontal disease containing as an active ingredient the extract of Achyranthes ginseng, which promotes the formation and activity of osteoclasts, inhibits the destruction of alveolar bone, and increases the proliferation and cell activity of periodontal ligament cells. can do.

Hereinafter, embodiments of the present invention will be described in detail so that those skilled in the art can easily implement it. However, the present invention may be implemented in many different forms and is not limited to the embodiments described herein.

[Preparation Example 1: Preparation of natural extract]

1. Preparation of Umbrella extract

The baby radish sprouts were washed, dried, and then pulverized. The ground product was mixed with ethanol, maintained at 98 to 100° C. for 2 hours, cooled, and filtered through Whatman filter paper to obtain the filtrate. This was prepared with Elephant herb extract (ESA).

2. Preparation of other natural extracts

Using the same method as the method for producing the above-mentioned Elephant herbacea extract (ESA), EVE extract (EVE), EPP extract, and Elephant primrose extract (ECJ) were prepared.

3. Preparation of mixed extract

A mixed extract was prepared by mixing the prepared above-mentioned Elephant eucalyptus extract (ESA), Elephant eucalyptus extract (EVE), Elephant primrose extract (EPP), and Elephant euphorbia extract (ECJ) within the weight range shown in Table 1 below.

ME1 ME2 ME3 ME4 ME5 ESA 100 100 100 100 100 EVE - 5 10 20 25 EPP - 5 10 20 25 ECJ - 5 10 20 25

(Unit: parts by weight)

[Example 1: Experiment on inhibition of secretion of TNF-α]

The effect of the Achyrantheus extract (ESA, ME1) and the mixed extract ME2 to ME5 prepared in Preparation Example 1 on the inhibition of the release of cytokine TNF-α released from THP-1 cells, a human monocytic cell line. To determine the effect, the following experiment was conducted.

THP-1 cells (ATCC No. TIB-202), a human-derived cell line, were purchased from ATCC (Rockville, USA), and 10% FBS (fetal bovine serum) was added to RPMI 1640 (Gibco, BRL, USA). After culturing in , it was used in experiments. The cell line was distributed in a 96-well plate at 5×10 ⁵ cells per ml, and lipopolysaccharide was added to activate the cells to secrete TNF-α.

Simultaneously with LPS, the experimental group was treated with Elephant herbacea extract (ESA) and mixed extracts ME2 to ME5, and after treatment, the amount of TNF-α in the cell culture was measured by enzyme-linked immunosorbent assay (ELISA), and the results are shown in Table 2 below. indicated.

ESA, ME1 ME2 ME3 ME4 ME5 TNF-α secretion amount 94.16±2.32 77.97±1.34 61.79±5.63 47.92±0.82 69.49±2.07

(Unit: mg/ml)

As shown in Table 2, as a result of treatment of the Euphorbiaceae extract (ESA, ME1) and the mixed extract ME2 to ME5 of the present invention, compared to ME1 containing only the Euphorbiaceae extract (ESA), the EVE extract ), it was confirmed that the secretion amount of TNF-α was significantly suppressed in a concentration-dependent manner in ME2 to ME5 further containing hairy primrose extract (EPP) and papilla extract (ECJ).

In particular, it was confirmed that the secretion amount of TNF-α was significantly suppressed by the mixed extract ME3 to ME4 according to the present invention compared to ME2 and ME5.

Therefore, it was found that the composition according to the present invention effectively inhibits the secretion of TNF-α.

[Example 2: Osteoblast proliferation experiment]

In order to determine the effect of the Elephant cerulean extract (ESA, ME1) and the mixed extracts ME2 to ME5 prepared in Preparation Example 1 on the proliferation of osteoblasts, the following experiments were performed.

As mouse osteoblasts, MC3T3-E1 cells (ATCC No. CRL-2593), a mouse calvarial cell line, were purchased from ATCC (Rockville, USA) and grown in DMEM (Gibco, BRL, USA) with 10% FBS (Fetal bovine serum). After culturing in the added medium, it was used in the experiment.

The osteoblasts were placed in a 24-well plate with 20,000 cells per well, cultured in a 5% CO ₂ incubator for 48 hours, and then treated with the Brachypodium herb extract (ESA, ME1) and the mixed extract ME2 to ME5 according to the present invention. was replaced with culture medium added at various mixing ratios, and cultured for an additional 48 hours. After removing the culture medium, trypsin-EDTA was added, and the number of cells was counted using a hemacytometer and shown as a percentage in Table 3 below.

ESA, ME1 ME2 ME3 ME4 ME5 Osteoblast proliferation 64 77 85 95 82

(unit: %)

As shown in Table 3 above, compared to the case containing only the Euphorbia chinensis extract (ESA, ME1), in ME4 further containing the Eucalyptus chinensis extract (EVE), the primrose extract (EPP), and the Eucalyptus herbacea extract (ECJ). It was confirmed that cell proliferation was promoted by up to 30%.

[Example 3: Inhibition of osteoclast activity]

In order to confirm the osteoclast growth and activity of the Aerobium chinensis extract (ESA, ME1) and the mixed extract ME2 to ME5 prepared in Preparation Example 1, osteoclast progenitors were placed on a plate (OAAS, OCT Inc.) coated with hydroxyapatite. Cells were cultured to observe TRAP activity and resorption, a marker enzyme for osteoclasts.

First, to isolate mouse bone marrow cells, a 7-9 week old male mouse was sacrificed by cervical torsion, the femur and tibia were removed aseptically, soft tissue was removed, both ends of the ilium were cut, and one end was cut using a 26G needle. Inject 1 ml of enzyme solution containing 0.1% Collagenase (Gibco), 0.05% trypsin, and 0.5 mM EDTA (Gibco) into the bone marrow cavity, remove the bone marrow, stir for 30 minutes, collect bone marrow cells, and 10% Non-adherent cells were obtained after pre-culturing for 24 hours in α-minimum essential medium (α-MEM) containing FBS. Non-adherent cells, which become osteoclast precursor cells, were distributed and cultured to obtain 2×10 ⁵ cells per well.

Among the cultured cells, the adherent cells were washed with PBS, fixed with citrate-acetateformaldehyde for 5 minutes, and incubated with Naphthol AS-BI Phosphate, Fast Garnet GBC solution, and TRAP staining was performed by culturing for 1 hour in acetate buffer (pH 5.0) at 37°C containing 7 mM tartrate buffer (pH 5). At this time, TRAP(+) multinucleated cells with three or more nuclei were considered osteoclasts.

To induce differentiation of osteoclasts, bone marrow known to contain osteoclast precursor cells was used. TRAP-positive (+) multinucleated cells were regarded as osteoclasts, and the osteoclast precursor cells were cultured.

The culture medium was removed from the cultured osteoclast progenitor cells. After cell culture, to remove attached cells, the OAAS plate was washed with distilled water, then 5% sodium hypochlorite solution was added and left for 5 minutes, then washed thoroughly with distilled water again, dried, and imaged of the absorbed area. Observation was made using Pro Plus (Image pro plus). That is, in order to test the activity of osteoclasts, which are mainly responsible for bone resorption within bone tissue, a plate coated with calcium and phosphate made to resemble the mineral part of bone tissue was used to culture osteoclast precursor cells according to the present invention. When treating the extracts (ESA, ME1) and the mixed extracts ME2 to ME5, it was observed whether there was a change in the absorption area.

The results are shown in Table 4 below.

ESA, ME1 ME2 ME3 ME4 ME5 Osteoclast activity inhibition rate 28 47 58 73 43

(unit: %)

As shown in Table 4, when the other extracts (ESA, ME1) and mixed extracts ME2 to ME5 according to the present invention were treated, osteoclast progenitor cells were cultured on a plate coated with calcium-phosphate to form osteoclasts. It was confirmed that activity and absorption were reduced.

In particular, in the case of the mixed extract ME3 to ME4 according to the present invention, an excellent inhibition rate on osteoclast activity was confirmed.

[Example 4: Gingival fibroblast and periodontal ligament cell proliferation experiment]

In order to determine the effect of the Elephant sycamore extract (ESA, ME1) and the mixed extract ME2 to ME5 prepared in Preparation Example 1 on the proliferation and increase in cell activity of human periodontal ligament cells and gingival fibroblasts, the following experiments were performed. It was carried out.

To confirm osteoclast growth and activity, osteoclast progenitor cells were cultured on plates coated with hydroxyapatite (OAAS, OCT Inc.), and TRAP activity and resorption, a marker enzyme for osteoclasts, were observed.

The periodontal ligament portion of a premolar (molar) extracted for the purpose of orthodontic treatment from a healthy person was scraped aseptically using a curette, placed on a petri dish, and then mixed with DMEM (Gibco, BRL, USA) with 10% FBS (fetal bovine). A small amount of culture medium containing serum was added and cultured. Gingival fibroblasts were cultured in the same manner by removing part of the gingiva after extraction from the above premolar teeth. At this time, it is observed under a microscope while culturing to confirm that cells emerge from the tissue as an explant. After the cells grew in clusters, each tissue sample was removed, the cells were allowed to grow as a single layer, and then subcultured and used in experiments. Human periodontal ligament cells and gingival fibroblasts were placed in a 24-well plate with 20,000 cells per well, cultured in a 5% CO ₂ incubator for 48 hours, and then mixed with the Achyrantheus extract (ESA, ME1) of the present invention. Extracts ME2 to ME5 were treated and cultured for an additional 48 hours.

After removing the culture medium, trypsin-EDTA was added, and the number of cells was counted with a hemacytometer and shown as a percentage in Table 5 below.

ESA, ME1 ME2 ME3 ME4 ME5 Gingival fibroblast proliferation rate 88 82 78 73 80 Periodontal ligament cell proliferation rate 41 53 72 78 58

(unit: %)

As shown in Table 4 above, it was confirmed that the proliferation of gingival fibroblasts by the extract according to the present invention was maintained at a normal or low level compared to the case of treatment with the Brachypodium herb extract (ESA, ME1) alone.

In addition, it was confirmed that cell proliferation was promoted by up to 40% when periodontal ligament cells were treated with the mixed extract ME4, compared to when the periodontal ligament cells were treated with the Euphorbiaceae extract (ESA, ME1) alone.

Therefore, it was confirmed that the extract according to the present invention can effectively proliferate periodontal ligament cells, which play the most important role in the regeneration of periodontal tissue, without significantly affecting the proliferation of human gingival fibroblasts.

Although the preferred embodiments of the present invention have been described in detail above, the scope of the present invention is not limited thereto, and various modifications and improvements made by those skilled in the art using the basic concept of the present invention defined in the following claims are also possible. falls within the scope of rights.

Claims (7)

It is a composition for preventing and treating periodontal disease containing Syneilesis aconitifolia (Bunge) Maxim extract as an active ingredient,
The periodontal disease includes diseases selected from the group consisting of gingivitis, periodontitis, and mixtures thereof.
Composition for preventing and treating periodontal disease. delete According to paragraph 1,
The composition shows the effect of preventing and improving dry mouth.
Composition for preventing and treating periodontal disease. According to paragraph 3,
The dry mouth is a group consisting of acid erosion, oral candidiasis, dysgeusia, dysphagia, oral dysesthesia, intraoral halitosis, and a combination thereof. Including diseases selected from
Composition for preventing and treating periodontal disease. According to paragraph 1,
The extract is extracted using an extraction solvent selected from the group consisting of water, C ₁ to C ₆ lower alcohols, and mixtures thereof.
Composition for preventing and treating periodontal disease. Containing a composition for preventing and treating periodontal disease according to any one of claims 1, 3 to 5.
Toothpaste for preventing periodontal disease. Containing a composition for preventing and treating periodontal disease according to any one of claims 1, 3 to 5.
Mouthwash for preventing periodontal disease.