Composition containing ginsenoside compound K or complex ginsenoside composition for reducing tooth surface bacterial film and inhibiting acidification of tooth surface bacterial film

Abstract

[Problem] To provide a composition derived from natural products that exhibits activity in reducing tooth surface bacterial membranes and inhibiting acidification of tooth surface bacterial membranes while having few or no side effects. [Solution] The present inventors have confirmed that ginsenoside compound K derived from red ginseng, etc., or a composite ginsenoside composition containing ginsenoside compound K, ginsenoside Rg3 (R+S), ginsenoside F2, ginsenoside Rh2 and ginsenoside Rb1, shows a remarkable effect of reducing bacterial membrane on tooth surface and inhibiting organic acid production on tooth surface. This composition can be used as oral compositions such as toothpaste and mouthwash, and can also be applied to food compositions such as gums, beverages and sweets.

Description

Technical Field

[0001] The present invention relates to a composition for reducing a dental surface bacterial film and inhibiting acidification of a dental surface bacterial film, which contains a composite ginsenoside composition extracted and / or processed from red ginseng or ginsenoside compound K.

[0002] The present invention reveals that it is the result of a project supported by the Ministry of SMEs and Startups of the Republic of Korea (Project ID: 1425147780, Project Name: Commercialization and efficacy verification of red ginseng specific saponin extract with anti-caries effect).

Background Art

[0003] Dental caries (caries) is one of the most prevalent oral diseases worldwide, and in recent years, its incidence has been increasing due to changes in dietary patterns, such as increased sugar intake. Dental caries (caries) is a multifactorial disease triggered by the interaction of bacteria in the plaque, food, and saliva. Among the bacteria in the plaque, Streptococcus mutans (S. mutans) and Streptococcus sobrinus (S. sobrinus) are the main causative agents. When these caries-causing bacteria attach to the tooth surface, they proliferate and produce acid, leading to demineralization of the inorganic matter in the tooth, destruction of the dentin, and loss of tooth tissue. Furthermore, for the organic acids produced by the caries-causing bacteria mentioned above to directly act on the tooth structure, the acid-producing bacteria must remain on the tooth surface, and for this to occur, insoluble glucans must be formed. The GTase (glucosyltransferase) secreted by these bacteria uses sucrose to form insoluble glucans, which are glucose polymers consisting of over 90% α-1,3-glucosidic linkages and the remainder being α-1,6-glucosidic linkages. Because these glucans are insoluble in water and can adhere to the tooth surface, they form a bacterial film on the tooth surface and create bacterial clumps. When these bacterial clumps cover the tooth surface, they form a dental plaque, and organic acids such as lactic acid accumulate within the dental plaque due to caries-causing bacteria, lowering the pH and inducing demineralization of the teeth. The bacterial film on the tooth surface hardens as calcium and phosphate from saliva adhere to it, forming tartar, and dental caries is accelerated in proportion to the amount of tartar (Guidelines for the Evaluation of Functional Health Foods, Ministry of Food and Drug Safety 2017.12, Bok et al., Kor.J. Food & Nutr. 13(2), 139(2000), Kim et al., J.Kor. Fish.Soc. 35(2), 191(2002)).

[0004] Various methods have been researched to develop treatments and preventive agents for dental caries. One promising method involves inhibiting the formation of biofilms on teeth to suppress tooth decay (Report on Research and Development of Health and Medical Technology, Biofilm Control Research for Preventive and Curative Agents for Dental Caries, 2013). Another frequently used method for suppressing biofilms is the development of GTase inhibitors that prevent biofilm formation (Ren. Z. et al., Antimicrob Agents Chemother. 2015). Furthermore, active research is being conducted on developing antimicrobial agents that suppress the growth of caries-causing bacteria for the treatment and prevention of dental caries (J. Periodont. Res., 21(16 suppl.)33(1986), Arch. Oral Biol., 17, 147(1972), J. Periodont. Res., 21(16 suppl.)74(1986)). In particular, there has been active development of antibacterial active preparations using natural extracts that are less toxic and have excellent stability. Tooth decay inhibitors using aloe extract (Kor.J.Food & Nutr.13(2), 139(2000)), kelp extract (J.Korean Fish.Soc.35(2), 191(2002)), or schisandra extract have been researched and developed (Korean Registered Patent No. 10-0296775). Since such natural extracts are made from edible plants, they have the advantage of being almost non-toxic and easy to ingest. However, because natural extracts are in the form of simple extracts that are mixtures of various substances, they contain trace amounts of specific pharmacologically active components. Moreover, it is difficult to mass-produce these components and adjust their content, resulting in poor reproducibility of pharmacological efficacy. Therefore, there is a limitation in that it is difficult to develop them into functional foods or pharmaceuticals.

[0005] Therefore, the inventors decided to search for an effective natural product-derived composition or a method for preventing tooth decay using the same that suppresses the formation of a bacterial film on the tooth surface and the acidification of that bacterial film, which are the key mechanisms of tooth caries development. [Overview of the Initiative] [Problems that the invention aims to solve]

[0006] The present invention aims to provide a composition derived from natural substances that exhibits a reduction effect on the bacterial film on the tooth surface and / or an inhibitory effect on the acidification of the bacterial film on the tooth surface, with few or no side effects. [Means for solving the problem]

[0007] The inventors have confirmed that a complex ginsenoside composition obtained by extracting a ginsenoside compound K extracted from red ginseng or ginseng, or by extracting a complex ginsenoside from red ginseng or ginseng using a solvent, adsorbing it onto an adsorption resin, then desorbing it with a solvent, partially purifying it, and mixing it with a red ginseng concentrate, exhibits the function of reducing bacterial film on the tooth surface and inhibiting acidification of bacterial film on the tooth surface. Based on this, they have invented toothpaste compositions, oral rinse compositions, food compositions, and pharmaceutical compositions containing these compositions. [Effects of the Invention]

[0008] The ginsenoside K or complex ginsenoside composition of the present invention demonstrated the effect of reducing bacterial film on the tooth surface and suppressing organic acid production on the tooth surface.

[0009] Therefore, the ginsenoside K or complex ginsenoside composition of the present invention can be used as oral compositions including toothpaste and mouthwash, and can also be applied to food compositions such as gum, beverages, and confectionery. [Brief explanation of the drawing]

[0010] [Figure 1] This shows the experimental results for the reduction of the bacterial film on the tooth surface, and photographs of caries bacteria attached to the glass tube after the experiment (*p<0.05, **p<0.01, ***p<0.001, ****p<0.0001 are compared to the control group).

[0011] [Figure 2] This shows the results of HPLC analysis of organic acid formation.

[0012] [Figure 3] This shows the results of HPLC analysis of a standard lactic acid sample and the standard curves for different concentrations.

[0013] [Figure 4] This shows the results of quantitative statistical processing by HPLC analysis of organic acid production (****p < 0.0001 compared with the control group; **p < 0.01 compared with compound K and BTEX-K).

Mode for Carrying Out the Invention

[0014] The present invention relates to

[0015] (a) ginsenoside compound K; or

[0016] (b) a composite ginsenoside composition containing 7 parts by weight or more of ginsenoside Rg3 (R + S), ginsenoside F2, ginsenoside Rh2, and ginsenoside Rb1, respectively, based on 100 parts by weight of ginsenoside compound K; as an active ingredient, and relates to a composition for reducing dental surface bacterial biofilms and inhibiting acidification of dental surface bacterial biofilms.

[0017] Further, the present invention relates to a composition for reducing dental surface bacterial biofilms and inhibiting acidification of dental surface bacterial biofilms, wherein the composite ginsenoside composition of (b) is obtained by

[0018] 1) extracting and concentrating red ginseng to obtain a red ginseng concentrate;

[0019] 2) dissolving the red ginseng concentrate, passing it through a synthetic adsorption resin column for adsorption, and then desorbing it with ethanol to obtain a composite ginsenoside containing ginsenoside Rb1, Rb2, Rc, and Rd;

[0020] 3) treating the composite ginsenoside with alpha-galactosidase to obtain a fermented composite ginsenoside; and

[0021] 4) adding the fermented composite ginsenoside obtained in step 3) to the red ginseng concentrate obtained in step 1) to obtain a composite ginsenoside composition.

[0022] The synthetic adsorption resin column is not particularly limited, but a porous adsorption resin column is preferred, and more preferably, it is one or more of HP2MG or HP-20.

[0023] In addition, in the present invention, the composite ginsenoside composition contains ginsenoside compound K at 100 mg / g or more, and ginsenoside Rg3 (R + S), ginsenoside F2, ginsenoside Rh2, and ginsenoside Rb1 each at 8 mg / g or more, preferably 8 mg / g or more and 30 mg / g or less, and more preferably 10 mg / g or more and 30 mg / g or less, and relates to a composition for reducing dental surface bacterial biofilm and suppressing acidification of dental surface bacterial biofilm.

[0024] In addition, in the present invention, the red ginseng concentrate in the above steps 1), 2), and 4) is in powder form or liquid form, and relates to a composition for reducing dental surface bacterial biofilm and suppressing acidification of dental surface bacterial biofilm.

[0025] In addition, in the present invention, the ginsenoside K or the composite ginsenoside composition is in liquid form, cream form, paste form, or solid form, and relates to a composition for reducing dental surface bacterial biofilm and suppressing acidification of dental surface bacterial biofilm.

[0026] In addition, the present invention relates to a pharmaceutical composition for preventing or treating dental caries, which contains the composition for reducing dental surface bacterial biofilm and suppressing acidification of dental surface bacterial biofilm.

[0027] Pharmaceutical compositions for the prevention or treatment of tooth decay, containing the ginsenoside compound K and / or the complex ginsenoside composition BTEX-K as an active ingredient, can be formulated with a carrier generally accepted in the pharmaceutical field and manufactured into dosage forms such as injection, oral administration, or topical application by conventional methods. For example, they can be formulated into liquids, syrups, capsules, granules, powders, ointments, emulsions, gels, creams, etc., which can be administered via various routes. Among the various dosage forms, for example, injection compositions are preferably isotonic aqueous solutions or suspensions. The compositions mentioned are sterilized and / or contain auxiliary agents (e.g., preservatives, stabilizers, wetting agents or emulsifiers, solution promoters, salts / or buffers for osmotic pressure adjustment). They may also contain other therapeutically useful substances. Pharmaceutically acceptable carriers include lactose, glucose, sucrose, sorbitol, mannitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate, and mineral oil. The composition may further contain lubricants, wetting agents, flavoring agents, emulsifiers, and preservatives. The composition of the present invention can be formulated into oral formulations such as granules, powders, liquids, tablets, capsules, or dried syrups, or parenteral formulations such as injections, but is not limited to these formulations.

[0028] The pharmaceutical preparations manufactured in this manner can be administered subcutaneously, intramuscularly, orally, intravenously, orally, orally, orally, orally, depending on the purpose. The daily dose can range from 0.001 μg / kg to 100 mg / kg and can be administered in a single dose or in multiple divided doses. The dosage level for a specific patient may vary depending on the patient's weight, age, sex, health status, administration time, administration method, and disease severity.

[0029] Furthermore, the pharmaceutical composition for the prevention or treatment of tooth decay containing the ginsenoside compound K and / or the complex ginsenoside composition BTEX-K as an active ingredient is particularly characterized as a topical preparation for application to the gums or oral cavity. The topical preparation containing the composition for the prevention or treatment of tooth decay of the present invention as an active ingredient can be easily manufactured in any form by conventional manufacturing methods. For example, when manufacturing a cream-type topical preparation, the anti-inflammatory composition of the present invention is incorporated into a general oil-in-water (O / W) or water-in-oil (W / O) cream base, and fragrances, chelating agents, dyes, antioxidants, preservatives, etc., are used as needed, while synthetic or natural materials such as proteins, minerals, and vitamins may be used in combination for the purpose of improving physical properties.

[0030] Furthermore, the present invention relates to a toothpaste composition comprising a composition for reducing the bacterial film on the tooth surface and suppressing the acidification of the bacterial film on the tooth surface.

[0031] Furthermore, the present invention relates to an oral cleansing composition comprising a composition for reducing the bacterial film on the tooth surface and suppressing the acidification of the bacterial film on the tooth surface.

[0032] Furthermore, the present invention relates to a food composition comprising a composition for reducing the bacterial film on the tooth surface and suppressing the acidification of the bacterial film on the tooth surface.

[0033] As used in this invention, the term "food" means a natural product or processed product containing one or more nutrients, preferably one that has undergone some processing steps and is ready to be eaten directly, and may include, in its ordinary sense, any of the following: food, food additive, functional food, health functional food, health supplement, and beverage. The food composition of this invention may be manufactured in the form of various beverages, gum, tea, confectionery, vitamin complex, health supplement, etc., as non-limiting examples. The preferred intake amount of the food composition of this invention will vary depending on the condition of the person consuming it, their weight, the severity of their symptoms, the food form, and the duration of intake, and may be appropriately selected. For optimal effect, it is preferable that the food composition of this invention be ingested at a rate of 0.2 mg / kg to 200 mg / kg of the active ingredient per day.

[0034] The present invention also relates to a toothpaste composition containing the ginsenoside compound K or a complex ginsenoside composition as an active ingredient. It can be formulated by adding mastic oil, propolis extract, sodium pyrophosphate, sodium fluoride, tocopherol acetate, hydroxyapatite, sweeteners, thickeners, abrasives, wetting agents, or surfactants as needed, and by a conventional method for manufacturing toothpaste compositions.

[0035] Furthermore, the present invention relates to an oral rinse composition containing the ginsenoside compound K or a complex ginsenoside composition as an active ingredient. It can be manufactured in dosage form by conventional methods for producing oral rinse compositions, with the addition of solubilizers, wetting agents, surfactants, lubricants, fragrances, sweeteners, preservatives, or pharmacokinetic agents as needed. It can be produced in dosage forms selected from the group consisting of gum, oral rinses, oral cleaners, oral sprays, oral ointments, oral patches, or combinations thereof. [Examples]

[0036] The specific methods of the present invention will be described in more detail below with reference to examples. However, it will be obvious to those with ordinary skill in the art to which the present invention belongs that the scope of the present invention is not limited by the description of these examples.

[0037]

[0038] Example 1: Production of complex ginsenosides

[0039] Dried red ginseng (4-year-old roots) from Korea was purchased and used as the experimental raw material (Chuncheonnam Deokmusan, Deokwon Ginseng). The complex ginsenoside was extracted from the dried red ginseng. 10 kg of dried red ginseng was repeatedly extracted and concentrated with 50% ethanol at 60°C for 12 hours three times to obtain a red ginseng concentrate.

[0040] The total volume of red ginseng concentrate was 5.6 kg (63 Brix). 5.5 kg of this concentrate was dissolved in water, then passed through a column packed with synthetic adsorption resin (HP2MG, Mitsubishi Chemical, or HP-20, Diaion) for adsorption. After desorption with 95% ethanol, 460 g of complex ginsenosides containing ginsenosides Rb1, Rb2, Rc, and Rd was obtained.

[0041]

[0042] Example 2: Production of Red Ginseng Concentrate Powder

[0043] Of the 5.6 kg of red ginseng concentrate produced in Example 1, 100 g was taken separately and concentrated under reduced pressure to obtain 71 g of red ginseng concentrate powder.

[0044]

[0045] Example 3: Fermentation of complex ginsenosides

[0046] Using 100 g of the complex ginsenoside obtained in Example 1 above as a substrate, it was reacted with alpha-galactosidase (0.75 g) derived from the Aspergillus genus in a 50 mM sodium acetate buffer solution (pH 4.5) in a 60°C constant temperature water bath for more than 84 hours to produce a fermented complex ginsenoside (41 g).

[0047] The fermented ginsenoside complex produced in this way contains ginsenoside compound K, ginsenoside F2, ginsenoside Rh2, and others.

[0048]

[0049] Example 4: Preparation of a complex ginsenoside composition (hereinafter mixed with "BTEX-K")

[0050] 100 g of the fermented complex ginsenoside produced in Example 3 was mixed with 40 g of the red ginseng concentrate powder prepared in Example 2 to produce the complex ginsenoside composition BTEX-K.

[0051] The BTEX-K complex ginsenoside composition contains various ginsenosides, including ginsenoside compound K=100mg / g, as well as ginsenoside F2 and ginsenoside Rh2.

[0052] [Table 1] TIFF0007899320000001.tif15104

[0053] Experimental Example 1: Analysis of reduction in bacterial film on tooth surface

[0054] 1) 5 mL of BHI (brain-heart infusion) liquid medium (broth) containing 1% sucrose was placed in a 50 mL glass tube, and 50 μl of the sample extracts at the concentrations shown in Table 2 below was added. For the control group, sterile water was added instead of the sample extract.

[0055] [Table 2] TIFF0007899320000002.tif21140

[0056] 2) S. mutans is 10 5 The inoculation was performed in this manner. 3) The glass tube was tilted 30° and incubated statically in a 37°C incubator for 24 hours.

[0057] 4) After culturing, 5 mL of 0.5 M NaOH solution was added to suspend the bacterial cells attached to the wall surface.

[0058] 5) Take 1 mL of the suspension and measure the absorbance at 550 nm.

[0059]

[0060] Experimental Example 2: Amount of organic acid produced by bacterial film on tooth surface

[0061] 1) 20 µl of sample extracts at the concentrations shown in Table 3 were added to 2 mL of BHI (brain-heart infusion) liquid medium. For the control group, sterile water was added instead of the sample extracts.

[0062] 2) Number of bacteria is 10 5 ~106 S. mutans was administered to achieve this result.

[0063] 3) The cells were incubated in a 37°C incubator for 48 hours.

[0064] 4) The amount of acid secreted into the culture medium was measured using HPLC.

[0065] [Table 3] TIFF0007899320000003.tif21140

[0066] **HPLC: Analysis requested from the Joint Instrumental Laboratory of the Industry-Academia Cooperation Group, Danguk University (Citric, pyruvic, succinic, lactic, formic, acetic, propionic, isobutyric, butyric, isovaleric acid)**

[0067] Result 1: Reduction in bacterial film on tooth surface

[0068] After inoculation with the S. mutans strain, the tubes were left to stand for 24 hours at a 30° angle. After standing culture, the culture medium was carefully poured out and discarded, and it was confirmed that bacteria were adhering to the wall of the glass tube (Figure 1, red arrow). Subsequently, the solution was suspended in 0.5M NaOH, and the absorbance of 1 ml of the suspended solution was measured at 550 nm. The results showed that a decrease in the bacterial film on the tooth surface began at a BTEX-K concentration of 50 ug / ml. In particular, a significant decrease was observed at a BTEX-K concentration of 100 ug / ml (p<0.0001).

[0069]

[0070] Result 2: Amount of organic acid produced by bacterial film on tooth surface

[0071] In 2 mL of BHI (brain-heart infusion) liquid medium, add 1 × 10⁶ S. mutans strain. 5The sample was inoculated at a CFU / ml concentration, and after inoculating a fixed amount of the sample, it was incubated for 48 hours. The culture medium was centrifuged at 13,000 rpm for 5 minutes, the supernatant was transferred to a new tube, filtered through a 0.45 μm syringe filter, and then analyzed by HPLC. The HPLC analysis conditions were as follows: Mobile phase (Isocratic elution) 10 mM Potassium phosphate monobasic (pH 3, phosphate) / MeOH = (98 / 2); Column oven temperature 30°C; Flow rate 1 mL / min; Wavelength 220 nm; Injection volume 10 μL

[0072] As confirmed by the tooth surface bacterial film reduction experiment, the growth of S. mutans strains was inhibited at concentrations of 100 ug / ml ginsenoside compound K and 100 ug / ml BTEX-K.

[0073] HPLC analysis revealed a decrease in the peak observed at 4.6 minutes compared to the control group at these concentrations. When compared with organic acid standard samples, lactic acid was detected at the same retention time.

[0074] After creating a graph of the lactate standard sample concentration, the difference in lactate content between the control group and the experimental group was analyzed and statistically processed. The results showed that ginsenoside compound K had approximately twice the lactate content compared to the control group (p<0.0001), and BTEX-K had approximately 3.2 times the lactate content compared to the control group (p<0.0001).

[0075] Furthermore, HPLC results from the control group showed a decrease in the 6-6.5 frequency band for ginsenoside compound K and BTEX-K. However, comparison with organic acid standard samples revealed no matching organic acids.

[0076] Furthermore, in the HPLC peaks of ginsenoside compound K and BTEX-K, the peak observed at the 13.5-minute retention band was not observed in the control group.

[0077] A reduction in the bacterial film on the tooth surface was observed with ginsenoside compound K and BTEX-K at concentrations of 20 ug / ml and 100 ug / ml, respectively.

[0078]

[0079] Furthermore, the inhibition of organic acid production on the tooth surface was analyzed by HPLC. Using 10 organic acid standard samples, ginsenoside compound K and BTEX-K were added to liquid culture media at different concentrations. A certain amount of S. mutans was then inoculated, and the culture was incubated at 37°C for 48 hours. The changes in organic acids produced in the culture medium were then observed in comparison with the control group. The results showed differences in the HPLC results for liquid culture media with high concentrations (100 ug / ml) of red ginseng extract, ginsenoside compound K, and BTEX-K compared to the control group.

[0080] The peak detected at approximately 4.6 minutes of HPLC retention time was confirmed to decrease in culture media supplemented with ginsenoside compound K and BTEX-K. Comparison with a standard sample confirmed that the peak detected at the 4.6-minute retention time was lactic acid. [Industrial applicability]

[0081] The ginsenoside K or complex ginsenoside composition of the present invention can be used as oral compositions, including toothpaste and mouthwash, and can also be applied to food compositions such as gum, beverages, and confectionery.

Claims

1. A composition for reducing the bacterial film on the tooth surface and inhibiting the acidification of the bacterial film on the tooth surface, comprising as an active ingredient a composite ginsenoside composition containing 8 to 30 parts by weight each of ginsenoside Rg3 (R+S), ginsenoside F2, ginsenoside Rh2, and ginsenoside Rb1 per 100 parts by weight of ginsenoside compound K;

2. The composite ginsenoside composition is characterized by containing 100 mg / g or more of ginsenoside compound K, and 8 mg / g or more and 30 mg / g or less each of ginsenoside Rg3 (R+S), ginsenoside F2, ginsenoside Rh2, and ginsenoside Rb1, as described in claim 1, for reducing the bacterial film on the tooth surface and inhibiting the acidification of the bacterial film on the tooth surface.

3. The composite ginsenoside composition is characterized in that it is liquid, cream, paste, or solid, and is a composition for reducing the bacterial film on the tooth surface and inhibiting the acidification of the bacterial film on the tooth surface according to claim 1 or 2.

4. A toothpaste composition comprising a composition for reducing the bacterial film on the tooth surface and inhibiting the acidification of the bacterial film on the tooth surface, which contains the complex ginsenoside composition according to claim 1 or 2.

5. An oral cleansing composition comprising a composition for reducing the bacterial film on the tooth surface and suppressing the acidification of the bacterial film on the tooth surface, which contains the complex ginsenoside composition according to claim 1 or 2.

6. A food composition comprising a composition for reducing the bacterial film on the tooth surface and suppressing the acidification of the bacterial film on the tooth surface, which contains the complex ginsenoside composition according to claim 1 or 2.

7. A pharmaceutical composition comprising the composition for reducing the bacterial film on the tooth surface and inhibiting the acidification of the bacterial film on the tooth surface according to claim 1 or 2.

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