WO2013147409A1 - Natural extract for preventing and treating oral disease - Google Patents

Abstract

The present invention relates to an extract having the activity of preventing and treating oral disease, wherein the extract is isolated from natural substances. The natural extract is characterized in that is shows growth inhibition activity of 50% or more and reactive oxygen species inhibition activity of 50% or more with respect to Streptococcus mutans.

Description

 Natural extracts for the prevention and treatment of oral diseases

The present invention relates to extracts having the activity for the prevention and treatment of oral diseases isolated from natural materials, more specifically, at least 50% growth inhibitory activity and at least 50% active oxygen against Streptococcus mutans. It is directed to a natural extract characterized by simultaneously exhibiting species inhibitory activity.

 Various kinds of bacteria form a colony in the mouth of a person. Saliva and secretions in the oral cavity contain a variety of proteins.For example, saliva containing an enzyme called Lysozyme is continuously secreted in the mouth to control microorganisms, but the nutrient-rich oral environment is optimal for microbial growth. It can be a condition, but it does not induce complete death. In particular, the oral tongue and dental plaque are used as a refuge for pathogenic microorganisms.

 About 600 to 800 species of microorganisms are believed to survive in the oral cavity of humans, and most of them are bacteria, but archae and fungi and yeast are also known. It is known to contain a lot.

 Most of them are harmless common bacteria or microorganisms that are symbiotic with them, but some of them are opportunistic pathogens, such as dental cavities and periodontal disease. Cause.

Dental caries is caused by microorganisms that form plaque (dental plaque), which undergo fermentation metabolism using nutrients in the mouth to produce various organic acids such as lactic acid and remove calcium from the tooth surface. The plaque is formed by a protein in the saliva forming a thin film on the teeth, the Streptococcus sobrinus ( Streptococcus sobrinus ), and Streptococcus mutans ( Streptococcus mutans ), a kind of Streptococcus It is made by forming a biofilm and is further thickened by Fuscobacterium.

 Once the tooth surface enamel layer is destroyed by organic acids, the tooth layer below it is also destroyed by the protease secreted by microorganisms, which in turn causes severe dental caries.

 Periodontal disease is a disease that causes inflammation and loss of structure of the periodontal tissue, also called the appearance. Specifically, periodontal disease is a disease in which tissues around the teeth holding the teeth are inflamed, and are clinically caused by gingival bleeding, swelling, periodontal pockets, and destruction of the alveolar bone. This results in loss of teeth.

 The main cause of periodontal disease is plaque. When the plaque is mechanically accumulated in the periodontal sac, it becomes a habitat for bacteria present in the periphery, and the habitat gradually migrates into aerobic, breathable and Gram-positive bacteria, and proliferates deep into the periodontal sac. Will be. At this time, toxins and all products of anaerobic gram-negative bacteria proliferated together directly destroy tissues or stimulate the immune system, causing inflammation along with periodontal tissue destruction by various actions from the stimulated immune system.

 According to a recent study, periodontal disease is an inflammatory disease that causes tissue damage as a result of complex interactions between the bacteria and the host's immune response. oxygen species (ROS) are also reported to be involved.

 Host cells and bacteria have an antioxidant defense system that eliminates damage caused by reactive oxygen species.

 The main extracellular antioxidants located in saliva, gingival crevicular fruid or plasma are ascorbic acid (vitamin C), uric acid, α-tocopherol (vitamin E), Carotenoids (vitamin A), bilirubin, bilirubin, reduced glutathione, transferrin, lactoferrin, haptoglobin, and the like.

 Recently, research has been actively conducted on herbal extracts that can be safely used without side effects while effectively removing oral bacteria in order to prevent dental caries. This is because the stability of is verified.

Herbal extracts exhibiting antimicrobial activity against S. mutans , the main cause of dental caries, include digging, grapefruit seeds, missing seeds, Angelica, cinnamon, propolis, seaweed, cocoa polyphenols, flavonoids, and palms. Cactus and the like are known. In addition, many polyphenols have been isolated from tea extracts such as green tea, black tea and oolong tea, which are known to partially inhibit the production of biofilm-forming substances emitted by caries.

 In addition, herbal extracts for preventing or treating periodontal disease include myrrha, raw white skin extract, lettuce extract, corn extract, thick extract, ginkgo leaf extract, jujube extract, milky white extract, rye extract, ginger extract, Toxic extracts are known.

 However, the conventional natural extracts exhibit only antibacterial activity or only antioxidant activity, so there is a problem that they can be used only for the prevention and treatment of some diseases of oral diseases.

 Therefore, there is a high need for a natural material having inhibitory activity against oral disease bacteria and periodontal disease-inducing reactive oxygen species.

 The present invention, in order to solve the problems of the prior art as described above and the technical problem that has been requested from the past, oral disease-inducing strain Streptococcus mutans(streptococcus mutans)of It is an object of the present invention to provide a natural extract having both growth inhibitory activity and active oxygen species inhibitory activity, which is a factor causing periodontal disease.

Therefore, the natural extract according to the present invention is an extract having the activity for preventing and treating oral diseases isolated from natural materials, and has at least 50% growth inhibitory activity and at least 50% active oxygen against Streptococcus mutans. It is characterized by simultaneously displaying species inhibitory activity.

 The 50% growth inhibitory activity and 50% reactive oxygen species inhibitory activity is the minimum concentration for the effectiveness of antimicrobial agents for the prevention and treatment of oral diseases, that is, the antimicrobial activity. When more than 50% bacterial growth inhibition is confirmed, it is considered to have antimicrobial activity.

 The natural extract may be isolated from natural materials selected from the group consisting of cypress, black samreung and persimmon.

The Thuja orientalis L. is an evergreen coniferous tree belonging to the genus Cypressaceae, reaching 25m in height and 1m in diameter, but belongs to shrubs, and small branches grow vertically. It is called. In oriental medicine, leaves and fruits are used as medicinal herbs, and the weakness of the leaves is mild and sour. Used. Fruit is weak and flat, and it has the effect of nourishing, calming, and ringing. It is a treatment for nervous breakdown, insomnia, palpitations, physical weakness, and constipation. Used.

In addition, the black samreung (Sparganium stoloniferum) is a perennial herb of the black samreung and grows in ponds and ditches, with stems extending laterally and stems coming from various places. Roots and stems are used as sojong, tong economy, and choije. Samneung (三稜), a Korean herb that has dried roots of black samreung, has no fragrance and a mild taste, but when chewed, it is slightly stinged and has an arine taste. Samneung protects qi to strengthen the flow of qi and eliminates blood, so it is used for postpartum blood cramps, dysmenorrhea, dysmenorrhea, dysmenorrhea, jinga, and harvesting. It also removes food that is difficult to digest food. . Pharmacological effects on ectopic pregnancy, effects on primary liver cancer, malignant tumors, gastric cancer, liver cancer, esophageal cancer, cervical cancer, mammary gland cancer, rectal cancer, lung cancer, etc. It is known that it has been shown to have an effect on improving cardiopathy and around the appendix of coronary artery disease.

 In addition, the persimmon tree is a deciduous tree belonging to the camphor tree growing in the sunny place of the foot of the mountain and is also called ganja or white camellia. According to the Korean Herb Book, "Sanchocho is the fruit of the camphor tree Lindera glauca Blume. The taste is spicy, warm in nature, and the effect is paralyzed, and it is used for cold-boiled cold. Suddenly speaking due to a stroke, you can put it in water and take it for months. Is written. Therefore, it is used in folk medicine for the treatment of tingling, paralysis, arthralgia and muscle pain due to customs, treatment of cardiac cold pain, traumatic bleeding, cold, headache, bleeding, and swelling.

 As such, natural medicinal materials such as leaves and berries of the cypress, black samreung and persimmon trees have been reported to have various pharmacological activities. At the same time, there are no reports and studies of pharmacological activity.

 The natural material may be a leaf, fruit or stem of the cypress tree, may be an outpost of the Black Samreung, may be a stem or leaf of Ecklonia chinensis.

The content of the leaf, fruit or stem extract of the cypress tree to have at least 50% growth inhibitory activity against Streptococcus mutans is at least 500 μg / ml.

In addition, the content of the black samreung outpost extract to have at least 50% growth inhibitory activity against Streptococcus mutans is at least 100 μg / ml.

In addition, the content of the stem or leaf extract of the persimmon tree to have at least 50% growth inhibitory activity against Streptococcus mutans is at least 100 μg / ml.

 In addition, the polyphenol content of the leaves, berries, or stems of the cypress, which exhibited more than 50% of active oxygen species inhibition activity, was measured in mg per 100 g of dry sample using the calibration curve by gallic acid (mg GAE / 100g dw). ) Is 395.27 ± 24.39 mg GAE / 100g dw, and the flavonoid content is 250.14 ± 18.26 mg when calculated as mg content (mg RE / 100g dw) per 100 g of dry sample using the calibration curve by rutin. When rutin / 100g dw or more, the content of leaf, fruit or stem extract of the cypress, which exhibits 50% DPPH (2,2-diphenyl-2-picrylhydrazyl hydrate) radical scavenging activity, is 33.10 ± 3.67 ㎍ / ml or more. The content of the leaf, fruit or stem extract of the cypress, showing% ABTS (2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) radical scavenging activity is 62.96 ± 5.23 μg / ml or more.

 At this time, 1 mg / ㎖ of the cypress leaf, fruit and stem extract exhibits nitrite scavenging activity produced by the periodontal disease inflammation of 90.44 ± 1.18%.

 In addition, the polyphenol content of the black samreung outpost showing 50% or more active oxygen species inhibition activity was calculated as mg content (mg GAE / 100g dw) per 100g of dry sample using the calibration curve by gallic acid. ± 13.57mg GAE / 100g dw and flavonoid content is 61.01 ± 3.62 mg rutin / 100g dw or more calculated as mg content per 100g of dry sample (mg RE / 100g dw) using calibration curve by rutin , The content of Black Samreung Outpost extract showing 50% DPPH radical scavenging activity is 166.30 ± 5.94 μg / ml or more, and the content of Black Samneung Outpost extract showing 50% ABTS radical scavenging activity is 155.76 ± 9.05 μg / ml or more.

 At this time, 1 mg / ㎖ of black samreung outpost extract exhibits nitrite scavenging activity produced by inflammation of the periodontal disease of 60.29 ± 4.12%.

In addition, the polyphenol content of the stem or leaves of E. coli showing 50% or more active oxygen species inhibition activity was obtained in mg content (mg GAE / 100g dw) per 100g of dry sample using the calibration curve by gallic acid. When calculated, it is 189.70 ± 4.33 mg GAE / 100g, and the flavonoid content is 67.50 ± 1.45 mg RE / 100g dw when the mg content per 100g of dry sample (mg RE / 100g dw) is calculated using the calibration curve by rutin. In this case , the extract content of E. coli stem or leaf exhibiting 50% DPPH radical scavenging activity is 28.45 ± 3.96 μg / ml or more, and the extract content of stem or leaf of E. coli showing 50% ABTS radical scavenging activity 34.52 ± 0.34 μg / ml or more.

 At this time, 1 mg / ml of the extract of E. coli stem and leaf exhibits nitrite scavenging activity produced by inflammation of periodontal disease of 67.64 ± 7.06%.

 The natural extract according to the present invention can be prepared by heating the natural material described above in water or alcohol to obtain an extract and concentrating the extract.

 For example, 150 g of dried natural cypress material is put into an extraction bottle and poured distilled water and extracted three times at a temperature of 80 to 121 ° C. for 5 hours, or 150 g of dried natural material is 95 to 99% ethanol. Alternatively, the mixture is heated in methanol at a volume of 1 to 15 times and heated at 70 to 80 ° C. for 12 hours in an extractor equipped with a cooling capacitor. The extracted extract is filtered under reduced pressure, and the solids are removed and the filtrate is concentrated with a cooling capacitor. Preferred in the present invention may be prepared by concentrating under reduced pressure to 50 to 90 brix and then freeze drying.

 The natural extract is an active ingredient for preventing and treating oral diseases, and can be used as a medicine regardless of the dosage form, and non-limiting examples include liquid toothpaste, mouthwash, mouth spray, oral ointment, oral varnish, and the like. It can be used in the form.

1 is a graph showing 50% growth inhibitory activity concentration determination of Streptococcus mutans of the extracts of the cedar, black samreung, and persimmon tree extracted according to the embodiment of the present invention.

 Hereinafter, the present invention will be further described with reference to Examples and the like, but the scope of the present invention is not limited thereto.

<Example 1>

 150g of dried natural materials of dried cedar, black samreung and persimmon trees were placed in an extraction bottle, and distilled water was poured and extracted three times at a temperature of 90 ° C. The extract was again filtered under reduced pressure with a waterman paper filter paper, solids were removed, and the remaining solution was concentrated under reduced pressure in a condenser equipped with a cooling capacitor and freeze-dried (90 brix).

Experimental Example 1

 Using the natural extract prepared in Example 1 was carried out experiments to determine the growth inhibition activity of the caries disease-causing bacteria, growth inhibitory activity concentration for the caries disease-causing bacteria. Efficacy determination was determined as the minimum concentration concentration showing a growth inhibitory effect of more than 50%. This is the minimum growth inhibitory activity concentration of bacteria for use in prophylactic and therapeutic purposes.

 Streptococcus mutans, an oral disease-causing bacterium, as an experimental bacterium(Streptococcus mutans)Was sold as a frozen vial from the Korean Collection for Type Cultures (KCTC)Streptococcus mutans KCTC3065 was used.

Streptococcus mutans was inoculated in brain heart infusion medium (Sheep brain and heart extract 37%) and incubated for 20 to 24 hours at 37 ℃, aerobic conditions. To determine the 50% growth inhibitory activity concentration (µg / ml) of each natural oral disease-inducing bacterium, natural extracts were added to the liquid medium, and 0 to 500 µg / ml through the serial dilution method. Diluted to a range of concentrations and sterilized. In addition, each of the bacteria pre-cultured in the medium added by the concentration of each natural extract was inoculated to 2 × 10 ⁶ cells / ㎖ in the spectrophotometer OD _600nm and incubated under the growth conditions of each bacteria. After cultivation, the absorbance was measured at OD _{600 nm} at the time of inoculation and at the end of incubation using a spectrophotometer to measure 50% growth inhibitory activity concentration value (µg / ml). The results are shown in Table 1 below.

Table 1

As shown in Table 1, when the concentration of 100 ~ 500 ㎍ / ㎖ of each extract according to the present invention, it can be seen that the growth of Streptococcus mutans , oral disease-causing bacteria have. The sample used in the experiment showed low growth inhibitory activity as an extract that was not completely purified, but it would be expected to show growth inhibitory activity at lower concentrations when purified. This means that in the production of oral hygiene products, even at low concentrations, it suppresses the growth of oral disease-causing bacteria, and thus even if a small amount is added, it can effectively prevent or treat oral diseases.

 The experimental results verified the significance between sample groups after three repeated measurements. However, the extract of each natural material obtained as described above may vary in efficacy depending on the growth environment of the plant, such as when the plant is collected, the place of habitation.

Experimental Example 2 Measurement of Total Polyphenol Content and Total Flavonoid Content

 Total polyphenol content and total flavonoid content were measured using the natural extract prepared in Example 1.

Polyphenols have an antioxidant effect and are known as major substances applied to foods and medicine. 100 μl of 10% Na ₂ CO _{3 was} added to 100 μl of the natural extract 1 mg / ml according to the present invention and allowed to stand at room temperature for 5 minutes.

 50% Folin-Ciocalteu with the extract mixture 100 μl of the reactant (Folin-ciocalteau's phenol reagent) was mixed and reacted, and stored in a dark room at room temperature for 60 minutes, and then absorbance was measured at 725 nm using a spectrophotometer. The total phenol content of the sample extract was calculated from the standard calibration curve obtained by preparing gallic acid at a concentration of 0 to 1,000 mg / mL and analyzing the same method as the sample.

 According to Moreno method (Ref. Nieva Moreno 2000), 100 μl of 10% aluminum chloride was added to 100 μl of natural extract 1 mg / ml, and then allowed to stand at room temperature for 5 minutes. After reacting by mixing 100 μl M acetate (potassium acetate) and storing at room temperature for 40 minutes, the absorbance was measured at 420 nm using a spectrophotometer. The total flavonoid content of the extract was calculated from the standard calibration curve obtained at a concentration range of 0 to 1,000 mg / ml using Rutin as a standard.

 The content of the ingredient was expressed in mg content per 100g of dry sample using calibration curves prepared from gallic acid and rutin, which are standard samples according to the recommendations of the Food and Drug Administration. The results are shown in Table 2 below.

TABLE 2

 As shown in Table 2, the total polyphenol and total flavonoid content of the extract according to the present invention was confirmed to have the highest content in the cypress. As an antioxidant, it was found that the cypress extract is more preferable than the black samreung extract and the persimmon tree extract.

 The experimental results were expressed as mean ± standard deviation after three repeated measurements. The significance between the sample groups was verified and analyzed according to Duncan's multiple range test at p <0.05. However, the extract of each natural material obtained as described above may vary in efficacy depending on the growth environment of the plant, such as when the plant is collected, the place of habitation.

Experimental Example 3 Measurement of Total Antioxidant Capacity Using DPPH Radical Scavenging Capacity and ABTS Radical

 Using the natural extract prepared in Example 1, DPPH antioxidant activity test and ABTS antioxidant activity test were performed to determine the free radical scavenging activity to determine the inhibitory effect of active oxygen species.

 DPPH radical scavenging activity was measured by mixing 60 μl of 0.2 mM DPPH (2,2-diphenyl-2-picrylhydrazyl hydrate) and 60 μl of the sample prepared for each concentration, and reacting in a dark room at room temperature for 15 to 30 minutes using a spectrophotometer. Absorbance was measured at.

 Total antioxidant activity using ABTS radicals was determined by the ABTS (2,2`-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) free radicals produced by reaction with potassium persulfate. Removed to decolorize the radical cyan.

Equivalently mix 7.4 mM ABTS and 2.6 mM potassium persulfate and react for 15 to 24 hours in the dark. Immediately before use, dilute with distilled water to absorbance 0.7 (± 0.03) at 734 nm. 50 μl of the sample was added to 950 μl of the diluent, mixed, and then reacted at room temperature for 10 minutes, and the absorbance was measured at 734 nm. The degree of antioxidant activity in the DPPH test is degree of antioxidant activity of a display and, ABTS test the 50% DPPH radical scavenging activity concentration of SC ₅₀ values are expressed as 50% ABTS radical scavenging activity concentration of ₅₀ SC value. The DPPH SC ₅₀ value of gallic acid used as a control was measured to be 9.07 ± 0.2 μg / ml and the ABTS SC ₅₀ value was 4.96 ± 0.28 μg / ml.

 In addition, the nitrite scavenging ability test of the natural extract showing the possibility of use as an antioxidant material to confirm the treatment effect of periodontal disease inflammation was performed.

Nitrite scavenging activity was performed by Kato et al. (Ref. Kato H, Lee IE 1987). 40 μl of 1 mM NaNO ₂ is added to 40 μl natural extract. After calibrating to pH 1.2 using 0.1 N HCl, the reaction was carried out at 37 ° C. for 1 hour. After adding 1 ml of 2% acetic acid and 100 μl of Griess reagent, the reaction was performed at room temperature for 15 minutes and the absorbance was measured at 520 nm. The results are shown in Table 3 below. 1 mg / ml of vitamin C (ascorbic acid) was used as a control of nitrite scavenging activity produced by periodontal disease inflammation, and the scavenging activity of the control group, vitamin C, was 96.64 ± 0.26%.

TABLE 3

 As shown in Table 3, the antioxidant efficacy measurement experiment using the DPPH and ABTS according to the present invention was able to confirm the excellent antioxidant effect in the cypress and Ecklonia cava, the nitrite scavenging activity was excellent in the cypress.

As shown in Tables 2 and 3, the extract used in the group consisting of the natural product of the natural material according to the present invention, the cedar, black samreung and persimmon tree is 50% growth inhibitory activity against Streptococcus mutans (Streptococcus mutans) At the same time, it exhibited 50% of active oxygen species inhibitory activity, in particular, the cypress had a desirable antioxidant effect.

 The experimental results were expressed as mean ± standard deviation after three repeated measurements. The significance between the sample groups was verified and analyzed according to Duncan's multiple range test at p <0.05.

 Those skilled in the art to which the present invention pertains will be able to perform various applications and modifications within the scope of the present invention based on the above contents.

As described above, since the natural extract according to the present invention simultaneously exhibits at least 50% growth inhibitory activity and at least 50% active oxygen species inhibitory activity against Streptococcus mutans , an oral disease causing strain at a predetermined concentration. In addition, medicines containing this as an active ingredient have the advantage of preventing and treating dental caries and periodontal disease at the same time.

In particular, when the natural extract according to the present invention is used as a toothpaste, oral cleaning agent, etc., the growth of Streptococcus mutans is suppressed by 50% or more, which has the advantage of efficiently removing plaque that is a bacterial precipitate. have.

 In addition, the drug and the like because it is used as an active ingredient natural extract has been proven safe, there are very few side effects.

Claims (13)

1.  Streptococcus mutans with extracts for the prevention and treatment of oral diseases isolated from natural products(Streptococcus mutans)50% or more growth inhibition and 50% activity against Natural extract, characterized in that simultaneously showing oxygen species inhibitory activity.
2.  According to claim 1, The natural extract is a natural extract, characterized in that separated from the natural material selected from the group consisting of cypress, black samreung and persimmon.
3.  The natural extract of claim 2, wherein the natural extract is prepared by heating a natural product into water or alcohol to obtain an extract, and concentrating the extract.
4. The method of claim 1, wherein the natural material is a leaf, fruit or stem of the cypress, the extract content of 500 ㎍ / ㎖ or more for 50% growth inhibition activity against Streptococcus mutans (Streptococcus mutans) Natural extract.
5. The method according to claim 1, wherein the natural material is the stem or leaves of the black samreung outpost or the Ego tree, the extract content of more than 100 ㎍ / ㎖ for growth inhibition activity of 50% or more against Streptococcus mutans (Streptococcus mutans) Characterized in natural extracts.
6. The method of claim 1, wherein the natural material is a polyphenol content of the leaves, berries or stems of the cypress, mg content (mg GAE / 100g dw) per 100g dry sample using a calibration curve by gallic acid (gallic acid) When calculated, it is 395.27 ± 24.39 mg GAE / 100g dw and the flavonoid content is 250.14 ± 18.26 mg rutin / 100g when calculated as mg content per 100g dry sample (mg RE / 100g dw) using the calibration curve by rutin. In the case of dw , the extract showing 50% DPPH (2,2-diphenyl-2-picrylhydrazyl hydrate) radical scavenging activity was 33.10 ± 3.67 ㎍ / ml or more, and 50% ABTS (2,2`-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) A natural extract, characterized in that the content of the leaf, fruit or stem extract of the cypress showing radical scavenging activity is 62.96 ± 5.23 μg / ml or more.
7.  The method of claim 1, wherein the natural material is 151.62 ± 13.57 when the polyphenol content of the black samreung outpost is calculated by the mg content per 100g dry sample (mg GAE / 100g dw) using a calibration curve by gallic acid (gallic acid) mg GAE / 100g dw and flavonoid content is calculated as mg content per 100g of dry sample (mg RE / 100g dw) using calibration curve by rutin, 50 or more 61.01 ± 3.62 mg rutin / 100g dw The content of the extract showing DPPH (2,2-diphenyl-2-picrylhydrazyl hydrate) radical scavenging activity of% was 166.30 ± 5.94 ㎍ / ml or more, and the content of the black tomb extract was 155.76 ± 9.05 ㎍ / Natural extract, characterized in that more than ml.
8.  The method of claim 1, wherein the natural material is the polyphenol content of the stem or leaves of Ecklonia cava, calculated as mg content (mg GAE / 100g dw) per 100g dry sample using a calibration curve by gallic acid (gallic acid) , 189.70 ± 4.33mg GAE / 100g, and the flavonoid content is 67.50 ± 1.45mg RE / 100g dw when calculated as mg content (mg RE / 100g dw) per 100g of dry sample using the calibration curve by rutin. An extract having a 50% DPPH (2,2-diphenyl-2-picrylhydrazyl hydrate) radical scavenging activity of 28.45 ± 3.96 μg / ml or more and having a 50% ABTS radical scavenging activity. Natural extract, characterized in that the content of the extract is 34.52 ± 0.34 ㎍ / ㎖ or more.
9. The method of claim 1, wherein the natural material is a leaf, fruit or stem of the cypress, the content of the extract for simultaneously having at least 50% growth inhibitory activity and 50% antioxidant activity against Streptococcus mutans (Steptococcus mutans) Natural extract, characterized in that more than 500 ㎍ / ㎖.
10. The method of claim 1, wherein the natural material is the stem or leaves of the black samreung outpost, 160 ㎍ content of the extract to have at least 50% growth inhibitory activity and 50% antioxidant activity against Streptococcus mutans (Streptococcus mutans) Natural extract, characterized in that more than / ml.
11. The method of claim 1, wherein the natural material is the stem or leaves of Ecklonia chinensis, the content of the extract for having at least 50% growth inhibitory activity and 50% antioxidant activity against Streptococcus mutans at the same time Natural extract, characterized in that more than / ml.
12.  A pharmaceutical formulation comprising a natural extract according to any one of claims 1 to 11.
13.  13. The pharmaceutical formulation of claim 12, wherein the pharmaceutical formulation is a liquid toothpaste, a mouthwash, an oral spray, an oral ointment or an oral varnish.

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