JP4515732B2 - Glucosyltransferase inhibitors, plaque formation inhibitors, antibacterial agents, oral preparations and foods for preventing touch - Google Patents

Description

  The present invention relates to a glucosyltransferase inhibitor utilizing a plant extract, a plaque formation inhibitor, an antibacterial agent against caries-causing bacteria and periodontal disease bacteria, an oral preparation, and food and drink for preventing touch.

  The occurrence of dental caries involves glucosyltransferase, which is an enzyme produced by microorganisms in the oral cavity, particularly Streptococcus mutans. That is, a part of sucrose in food and drink remaining in the oral cavity is changed into water-insoluble and highly adherent glucan by the action of glucosyltransferase, and it adheres to the tooth surface together with oral microorganisms and becomes plaque (dental Form). The dental caries metabolize sugars in food and drink to produce acid, and this acid decalcifies and erodes tooth enamel.

  Therefore, in order to prevent dental caries, not only the plaque adhered to the tooth surface is removed by brushing, but also the production of glucan is inhibited by inhibiting the growth of Streptococcus mutans and the action of glucosyltransferase in the oral cavity. It is most effective to prevent and thus avoid plaque.

  From such a point of view, in recent years, by containing a substance having a glucosyltransferase inhibitory action or a substance that suppresses plaque formation, oral preparations and foods and drinks having a caries preventive action have been provided ( Patent Documents 1 to 3).

On the other hand, as the action which No grape has, liver cirrhosis prevention action (patent document 4), liver disease treatment action (patent document 5), fibrosis inhibiting action (patent document 6), immunoregulatory action (patent document 7) and the like are known. ing.
JP-A-4-221308 JP 2001-114694 A JP 2003-81847 A Japanese Patent Publication No. 60-7974 Japanese Patent Laid-Open No. 2-48533 Japanese Patent Laid-Open No. 5-271667 JP-A-8-3052

  The present invention finds a plant extract that has not been known to have a glucosyltransferase inhibitory action, plaque formation-inhibiting action, or caries-causing bacteria / periodontal disease bacteria, and is highly safe glucosyl using the same. An object of the present invention is to provide a transferase inhibitor, a plaque formation inhibitor, an antibacterial agent, an oral preparation, and a food and drink for preventing touch.

  In order to achieve the above object, firstly, the present invention provides a glucosyltransferase inhibitor (Claim 1) and a plaque formation inhibitor (Claim 2) each containing an extract from Ampelopsis brevipedunculata Trautv as an active ingredient. And an antibacterial agent against caries-causing bacteria and periodontal disease bacteria (Claim 3).

  ADVANTAGE OF THE INVENTION According to this invention, the safe novel glucosyltransferase inhibitor using a plant extract, the plaque formation inhibitor, the antibacterial agent with respect to a caries causative microbe, and a periodontal disease microbe, an oral agent, and the food and drink for touch prevention are provided. be able to. According to the glucosyltransferase inhibitor, the plaque formation inhibitor, the antibacterial agent, the oral preparation or the food for preventing touch according to the present invention, the glucosyltransferase activity is inhibited, the plaque formation is inhibited, or the caries By suppressing the growth of causative bacteria or periodontal disease bacteria, caries and periodontal disease can be effectively prevented.

Hereinafter, the present invention will be described in detail.
The glucosyltransferase inhibitor, the plaque formation inhibitor and the antibacterial agent according to the present invention all contain an extract from no grape as an active ingredient.

  Here, the "extract" from grapevine is an extract obtained from grapevine by extraction treatment, a diluted or concentrated solution of the extract, a dried product obtained by drying the extract, or a crude product thereof. Both products and purified products are included.

  No Grapes is a vine family plant with the scientific name “Ampelopsis brevipedunculata Trautv”, also known as gabion, and is a deciduous vine plant that grows naturally in Hokkaido, Okinawa and the mountains of the Korean Peninsula. In China, the leaves and vines of wild grapes are said to be effective for diuresis and gastric fever, and have been decocted for a long time, and have antibacterial and hemostatic effects against E. coli and staphylococci. In the present invention, among each part of grapevine, one or both of a leaf part and a vine part containing a component exhibiting a glucosyltransferase inhibitory action, a plaque formation inhibitory action and an antibacterial action against caries-causing bacteria and periodontal bacteria are used. It is preferable to do this.

  The grapes (leaves and / or vines) used as the raw material for extraction are preferably dried, cut into pieces, chopped or crushed immediately after collection. Drying may be performed in the sun or using a commonly used dryer. The grapes may be used as a raw material for extraction after pretreatment such as degreasing with a nonpolar solvent such as hexane or benzene. By performing a pretreatment such as degreasing, the extraction process with the polar solvent of No grape can be efficiently performed.

  In the extraction process, it is preferable to use a polar solvent as the extraction solvent. Components that exhibit glucosyltransferase inhibitory action, plaque formation inhibitory action, and caries-causing bacteria and periodontal disease bacteria contained in wild grapes can be easily extracted by extraction using a polar solvent as an extraction solvent.

  Specific examples of suitable extraction solvents include water, lower aliphatic alcohols, hydrous lower aliphatic alcohols, and the like, and these can be used alone or as a mixture of two or more thereof. Specific examples of suitable lower aliphatic alcohols include methanol, ethanol, propanol, isopropanol, 1,3-butylene glycol, ethylene glycol, glycerin, propylene glycol, isoprene glycol and the like. These extraction solvents are easy to handle and the extraction operation can be performed relatively easily.

  Water that can be used as the extraction solvent includes pure water, tap water, well water, mineral spring water, mineral water, hot spring water, spring water, fresh water, and the like, as well as those subjected to various treatments. Examples of the treatment applied to water include purification, heating, sterilization, sterilization, filtration, ion exchange, adjustment of osmotic pressure, buffering, and the like. Therefore, the water that can be used as the extraction solvent in the present invention includes purified water, hot water, ion-exchanged water, physiological saline, phosphate buffer, phosphate buffered saline, and the like.

  When using the liquid mixture of 2 or more types of polar solvents as an extraction solvent, the mixing ratio can be adjusted suitably. For example, when using the liquid mixture of water and a lower aliphatic alcohol, the mixing ratio of water and a lower aliphatic alcohol can be 7: 3 to 2: 8 (weight ratio).

  The extraction treatment is not particularly limited as long as the soluble component contained in wild grape can be eluted in the extraction solvent, and can be performed according to a conventional method. In the extraction process, it is not necessary to adopt a special extraction method, and any apparatus can be used at room temperature or under reflux heating.

  For example, the extraction raw material is put into a treatment tank filled with the extraction solvent, and the soluble components are eluted with gentle stirring. At this time, although the extraction conditions can be appropriately adjusted according to the extraction raw materials and the like, the amount of the extraction solvent is usually 5 to 15 times (weight ratio) of the dried extraction raw material, and the extraction time is usually 1 to 3 hours. The extraction temperature is usually from room temperature to about 80 ° C.

  After the soluble component is eluted by extraction treatment, it is subjected to solid-liquid separation by filtration or centrifugation, and further subjected to treatments such as concentration and drying as necessary to obtain the desired extract.

  The obtained grape extract has strong glucosyltransferase inhibitory activity, plaque formation inhibitory activity, and antibacterial activity against caries-causing bacteria and periodontal disease bacteria, so it can be used as it is as a glucosyltransferase inhibitor, plaque formation inhibitor or antibacterial agent. However, if desired, it may be purified to improve the activity, and may be formulated by a conventional method.

  Specifically, the grapevine extract can be purified by activated carbon treatment, adsorption resin treatment, ion exchange resin treatment, or the like. Moreover, when formulating a grape extract, any auxiliary, excipient, water or organic solvent for use as a solution can be appropriately contained.

  In addition, the grapevine extract usually exhibits any of the glucosyltransferase inhibitory action, the plaque formation inhibitory action, and the antibacterial action against caries-causing bacteria and periodontal disease bacteria, but depending on the case, either one or two of them may be used. In such a case, it may be used as a drug that is compatible with the action.

  Here, the glucosyltransferase inhibitor, plaque formation inhibitor and antibacterial agent of the present invention contain no grape extract as an active ingredient, but other than no grape extract, glucosyltransferase inhibitory action, plaque formation inhibitory action or caries cause It does not prevent the inclusion of substances having antibacterial action against bacteria and periodontal disease bacteria.

  The glucosyltransferase inhibitor, plaque formation inhibitor and antibacterial agent of the present invention are added to oral preparations or foods, respectively, to prevent glucan production in the oral cavity, suppress plaque formation, or caries-causing bacteria or periodontal disease bacteria. Can be used to control the breeding of That is, according to the oral preparation and food and drink containing the glucosyltransferase inhibitor, plaque formation inhibitor or antibacterial agent of the present invention, in the oral cavity, inhibiting glucosyltransferase activity to prevent the production of glucan, Caries and periodontal disease can be effectively prevented by suppressing the formation or by inhibiting the growth of caries-causing bacteria or periodontal disease bacteria.

  In addition to the use as an oral preparation or food and drink, the above-mentioned drug is used for all purposes that are meaningful to exhibit a glucosyltransferase inhibitory action, a plaque formation inhibitory action, or an antimicrobial action against caries-causing bacteria and periodontal disease bacteria. It is possible to use.

  The oral preparation and the food for preventing touch according to the present invention contain at least one drug selected from the group consisting of the glucosyltransferase inhibitor, the plaque formation inhibitor and the antibacterial agent. In this case, the glucosyltransferase inhibitor, the plaque formation inhibitor and the antibacterial agent may be composed of only grapevine extract.

  Examples of the form of the oral preparation include various toothpastes, mouthwashes, troches, oral pasta, gingival massage creams, gargles, mouth fresheners, and the like.

  The addition of the glucosyltransferase inhibitor, the plaque formation inhibitor or the antibacterial agent does not limit the other components of the oral preparation and the method for producing the oral preparation. For example, calcium hydrogen phosphate, calcium carbonate, Abrasives such as insoluble sodium metaphosphate, aluminosilicate, silicic acid anhydride, resin; surfactants such as sodium long-chain alkyl sulfate, sodium lauryl sulfoacetate, lauryl diethanolamide, sucrose fatty acid ester; CMC, hydroxyethyl cellulose, alginate , Carrageenan, gum arabic, polyvinyl alcohol, etc .; viscous agents such as polyethylene glycol, sorbitol, glycerin, propylene glycol; sweeteners such as saccharin, steviosides, glycyrrhizic acid, thaumatin; dehydro vinegar Preservatives such as sodium dehydroacetate; menthol, carvone, eugenol, anethole, peppermint oil, spearmint oil, peppermint oil, eucalyptus oil, ginger oil, anise oil, etc .; The raw material to be used can be arbitrarily selected according to the type and use of the product, and the oral preparation can be produced by a conventional method.

  In producing the oral preparation of the present invention, in addition to the glucosyltransferase inhibitor, the plaque formation inhibitor and the antibacterial agent, a glucosyltransferase inhibitor, a plaque formation inhibitor or an antibacterial agent against caries-causing bacteria and periodontal disease bacteria are used in combination. Further, by adding an optional anti-inflammatory agent, antibacterial agent, deodorant and the like, a more excellent oral preparation can be provided.

  Examples of anti-inflammatory agents that can be added include Acacia yak, licorice, walrus, Japanese cypress, Japanese cypress, psycho, hawthorn, perilla, peony, Sakuhaku, Kyonin, Taiso, Clove, Tounin, Nikuzuku, Buttonpi, Azulene, Allantoin, Ursolic acid, Examples include oleanolic acid, glycyrrhizic acid, glycyrrhetinic acid, tocopherol, and tranexamic acid.

  Examples of antibacterial agents that can be added include gobishi, saicin, salamander, ginger, dill, thyme, rosemary, ascorbic acid, mustatin, humic acid, linoleic acid, linolenic acid and the like.

  Examples of deodorants that can be used in combination include amacha, fennel, radish, ellagic acid, keihi, pepper, mace, sage, perilla, ginkgo, oyster leaf, green tea, oolong tea, red pepper, rosin, oyster astringent, actisol, chlorophyllin derivatives , Chlorhexidine, Maillard reactant and the like.

  On the other hand, examples of the types of food and drink include confectionery, bread, candy, chewing gum, gummi, jelly, chocolate, tablet confectionery, soft drinks, and pet food.

  The preferred blending ratio of the glucosyltransferase inhibitor, plaque formation inhibitor or antibacterial agent of the present invention in oral preparations or foods and drinks varies depending on the content of the active ingredient in the drug, the strength of activity, or the addition target, The grape extract is preferably about 0.001 to 5.0% by weight, particularly about 0.05 to 1.0% by weight.

  The glucosyltransferase inhibitor, plaque formation inhibitor, antibacterial agent, oral preparation and food and drink of the present invention described above are suitably applied to humans, but have the intended effects. As long as it is applicable to animals other than humans.

  Hereinafter, although a manufacture example, a test example, and a compounding example are shown and this invention is demonstrated concretely, this invention is not limited to each following example at all.

[Production Example 1]
100 g of dried pulverized leaves and vines were immersed in 1 liter of 30% ethanol (weight ratio of water to ethanol 7: 3) and heated under reflux for 2 hours. The resulting extract was filtered and the residue was again treated with 1 liter of 30% ethanol in the same manner. The extracts obtained by these two extraction processes were mixed and concentrated under reduced pressure to obtain 14 g of 30% ethanol-extracted no grape extract (no grape extract).

[Production Example 2]
The extraction treatment was performed in the same manner as in Example 1 except that 50% ethanol (water: ethanol weight ratio 1: 1) was used as the extraction solvent. As a result, 9.9 g of 50% ethanol-extracted no grape extract (no grape extract) was obtained.

[Production Example 3]
100 g of dried pulverized leaves and vines were immersed in 1 liter of hot water and extracted for 2 hours. The obtained extract was filtered, and the residue was treated in the same manner with 1 liter of hot water again. The extracts obtained by these two extraction treatments were mixed and concentrated under reduced pressure to obtain 10 g of hot water extraction no grape extract (no grape extract).

[Test Example 1] Test for production of glucosyltransferase inhibition The test for production of glucosyltransferase inhibition was carried out by the following test method for the grape extract (hereinafter referred to as "sample") obtained in Production Examples 1 to 3.

  50 μL of sample solution, 1000 μL of 2% sucrose aqueous solution containing 0.1% sodium azide, 50 μL of crude glucosyltransferase solution (prepared from Streptococcus mutans) and 900 μL of distilled water were added to a test tube and mixed.

  After the mixture was reacted at 37 ° C. for 5 hours, the produced glucan was dispersed in water with a stirrer, and the absorbance at 550 nm was measured as an index of turbidity. Separately, as a control, the same operation and measurement were performed on a sample solution added with a solvent instead of the sample solution. Further, in each case, the same operation and measurement were performed without adding the crude glucosyltransferase solution.

Based on the measurement results, the inhibition rate of glucosyltransferase activity was calculated by the following formula.
Inhibition rate (%) = [1− (A−B) / (C−D)] × 100
However, A: Absorbance of the sample solution after the enzyme reaction B: Absorbance of the sample solution when the enzyme is not added C: Absorbance after the enzyme reaction of the control D: Absorbance of the control solution when the enzyme is not added

The above measurement was carried out by gradually reducing the concentration of the sample solution, and the concentration IC ₅₀ (ppm) of the sample solution at which the inhibition rate was 50% was determined by interpolation. The results are shown in Table 1.

[Table 1] Glucosyltransferase inhibition test
Sample IC ₅₀ (ppm)
30% ethanol extract 560
50% ethanol extract 710
Hot water extract 690

  From the results shown in Table 1, it was confirmed that the grapevine extract has an action of inhibiting glucosyltransferase. In addition, since the strength of the glucosyltransferase inhibitory effect of the grapevine extract changes depending on the concentration of the grapevine extract, the strength of the inhibitory effect of the glucosyltransferase activity is adjusted by adjusting the concentration of the grapevine extract. It was confirmed that it was possible.

[Test Example 2] Plaque Formation Inhibition Test The grape formation extract (hereinafter referred to as “sample”) obtained in Production Examples 1 to 3 was subjected to a plaque formation inhibition test by the following test method.

  5.35 mL of brain heart infusion broth (manufactured by Nissui Pharmaceutical Co., Ltd.) containing 2% sucrose was added to a pre-weighed test tube and heat sterilized. Into the test tube, 0.15 mL of a sample solution (solvent: hot water extract is water, hydrous ethanol extract is 50% ethanol) and 0.5 mL of a culture solution of Streptococcus mutans 6715 are added, and the mixture is incubated at 37 ° C. for 19 hours. Culture was performed. After completion of the culture, the supernatant was gently removed, and the plaque-like deposits on the test tube wall were washed three times with distilled water as they were and then dried at 105 ° C. for 5 hours. Finally, each test tube was weighed to determine the dry weight w of the plaque-like deposit in the tube.

Separately, as a control, the same operation as described above was performed using the solvent of the sample solution instead of the sample solution, and the dry weight W of the plaque-like deposit was obtained. From the measured dry weights w and W of the plaque-like deposit, the plaque formation inhibition rate was calculated by the following formula.
Plaque formation inhibition rate (%) = (1−w / W) × 100

The above-mentioned measurement was performed by gradually reducing the concentration of the sample solution, and the concentration IC ₅₀ (ppm) of the sample solution at which the inhibition rate was 50% was determined by interpolation. The results are shown in Table 2.

[Table 2] Plaque formation inhibition test
Sample IC ₅₀ (ppm)
30% ethanol extract 135
50% ethanol extract 62.5
Hot water extract 275

  From the results shown in Table 2, it was confirmed that the grape extract has an action of suppressing the formation of plaque. In addition, since the strength of the inhibitory effect on the plaque formation of the grapevine extract changes depending on the concentration of the grapevine extract, the strength of the inhibitory effect on the plaque formation can be adjusted by adjusting the concentration of the grapevine extract. confirmed.

[Test Example 3] Antibacterial action test against caries-causing bacteria and periodontal disease-causing bacteria The grapevine extracts obtained in Production Examples 1 to 3 have the following antibacterial activities against microorganisms A to F, which are the minimum by the agar medium dilution method. The growth inhibitory concentration (MIC) was evaluated. For Streptococcus mutans (A), use BHI medium, culture temperature is 37 ° C., culture time is 48 hours, and for other microorganisms (BF), use modified GAM agar medium. The temperature was 37 ° C., the culture time was 72 hours, and the presence or absence of growth of each microorganism was judged with the naked eye. The results are shown in Table 3. In addition, the numerical value in a table | surface is MIC (microgram / ml), and this numerical value becomes small, so that antibacterial activity is strong.

[Test microorganism]
A: Streptococcus mutans
B: Porphyromonas gigivalis (periodontal disease-causing bacteria)
C: Propionibacterium acnes (causing periodontal disease)
D: Actinomyces viscosus (periodontal disease-causing bacteria)
E: Prevotella intermedia
F: Prevotella melaninogenica (periodontal disease-causing bacteria)

[Table 3] Minimum inhibitory concentration [MIC (μg / ml)]
Microbe 30% ethanol extract 50% ethanol extract Hot water extract A 2000 2000 2000
B 2000 2000 2000
C 1000 2000 2000
D 1000 2000 2000
E 1000 2000 2000
F 1000 2000 2000

  From the results shown in Table 3, it was confirmed that the grapevine extract has an antibacterial action against caries-causing bacteria and periodontal disease-causing bacteria.

[Formulation Example 1]
The following raw materials were processed by a conventional method of toothpaste production to produce a toothpaste that is a kind of oral preparation.
No Grape Hot Water Extract (Production Example 3) 0.1g
Calcium carbonate 35.0g
Sorbitol 20.0g
Carboxymethylcellulose 1.0g
Sodium lauryl sulfate 1.0g
Saccharin 0.1g
Fragrance 1.0g
Chlorhexidine hydrochloride 0.01g
Allantoin 0.01g
0.01g dipotassium glycyrrhizinate
The remainder of purified water (the total amount is 100 g)

[Formulation Example 2]
The following raw materials were processed by a conventional method for producing a mouth freshener to produce a mouth freshener which is a kind of oral preparation.
No Grape 30% Ethanol Extract (Production Example 1) 0.5g
l-Menthol 0.1g
Peppermint oil 0.1g
4g polyglycerin fatty acid ester
Ethanol 45g
Saccharin sodium 0.01g
pH adjuster Appropriate amount of purified water The remainder (the total amount is 100 g)

[Composition Example 3]
Candy was produced according to a conventional method using the following raw materials.
Grape 30% ethanol extract (Production Example 1) 1g
Turmeric extract 0.2g
45g of granulated sugar
Enzyme saccharified starch syrup 43g
Cyclodextrin 0.6g
Fragrance 0.1g
Azulene 0.1g
The remainder of water (the total amount is 100 g)

[Formulation Example 4]
A chewing gum was produced according to a conventional method using the following raw materials.
No Grape 50% Ethanol Extract (Production Example 2) 3g
Silymarin 2g
Chewing gum base 20g
50g of granulated sugar
Minamata 15g
Softener 5g
Chlorophyll 0.1g
Fragrance 0.1g

The glucosyltransferase inhibitor, plaque formation inhibitor, antibacterial agent, oral preparation or food and drink for preventing touch according to the present invention can greatly contribute to the prevention of dental caries or periodontal disease.

Claims (3)

  A glucosyltransferase inhibitor characterized by containing an extract from grapevine (Ampelopsis brevipedunculata Trautv) as an active ingredient.   A plaque formation inhibitor characterized by containing an extract from vine grape (Ampelopsis brevipedunculata Trautv) as an active ingredient.   An antibacterial agent against caries-causing bacteria and periodontal disease bacteria characterized by containing an extract from grapevine (Ampelopsis brevipedunculata Trautv) as an active ingredient.