JP2004256557A - Antimicrobial agent containing chitosan oligosaccharide and method for suppressing growth of cariogenic bacterium - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a highly effective antimicrobial agent containing a chitosan oligosaccharide and causing no decalcification to dentin, to provide a method for suppressing the growth of cariogenetic bacteria. <P>SOLUTION: The agent contains the chitosan oligosaccharide as an active ingredient. The oligomer of the oligosaccharide preferably has 2-8 polymerization degree and 80-100 degree of deacetylation. More preferably, the agent has pH6.5 or more and the concentration of the oligomer is ≥0.5 wt.%. <P>COPYRIGHT: (C)2004,JPO&NCIPI

Description

  The present invention relates to a chitosan oligosaccharide-containing antibacterial agent containing chitosan oligosaccharide as an active ingredient, and a method for suppressing the development of cariogenic bacteria, and more particularly to self-care against dental caries at home as well as clinical application of dentistry. The present invention relates to a chitosan oligosaccharide-containing antibacterial agent that can also be used in care and a method for suppressing the growth of cariogenic bacteria.

With regard to the prevention and control of caries, so-called caries, fluorine preparations are generally used worldwide in the present and the past, but in recent years, their application to dentistry using the antibacterial action of chitosan has attracted attention.
Chitosan is obtained by chemical treatment of chitin, which is a component contained in the outer shell of crabs and shrimps, and its structure is a form in which D-glucosamine is linked by β-1,4 bonds. It is a polysaccharide having a degree of conversion. While chitosan is used as artificial skin by utilizing its high affinity for living organisms, in recent years, antibacterial action against Streptococcus mutans that contributes to caries has been reported (for example, see Non-patent Document 1). In addition, a dentifrice containing chitosan has also been reported (see, for example, Patent Document 1).

Kim et al., “Carbohydrate Research” 2001, 44, p. 71-76 JP-A-9-249541 (Claim 1)

However, the former is testing the ability to suppress dental caries at pH 6, and at this pH value, demineralization of the tooth occurs, which is rather counterproductive for caries prevention / suppression. On the other hand, the latter has tested chitosan only in polymers with a high degree of polymerization, and the effectiveness of chitosan with a low degree of polymerization is unclear.
Therefore, an object of the present invention is to develop a chitosan oligosaccharide-containing antibacterial agent that has chitosan as an active ingredient and exhibits a high effect at a pH value at which tooth decalcification does not occur.

In order to achieve the above object, the present inventors have paid attention to the antibacterial action of chitosan and conducted intensive studies, thereby completing the present invention.
The chitosan oligosaccharide-containing antibacterial agent of the present invention comprises chitosan oligosaccharide as an active ingredient. Furthermore, it is preferable that the degree of polymerization of chitosan oligosaccharide is 2 to 8, the degree of deacetylation is 80 to 100%, and / or the concentration is 0.5% by weight or more, and the pH value of the chitosan oligosaccharide-containing antibacterial agent is 6 More preferably, it is 5 or more.

Since the chitosan oligosaccharide-containing antibacterial drug of the present invention contains chitosan oligosaccharide, it has a high caries bacterial growth inhibitory effect, and therefore can effectively suppress the production of lactic acid that invade the tooth structure, and is excellent. Antibacterial effect of cariogenic bacteria is obtained.
In addition, by setting the pH value to 6.5 or more at which the dentin does not demineralize, the gingiva is retracted and the root surface is exposed, or the enamel is damaged and there is a minute gap reaching the dentin, The chitosan oligosaccharide-containing antibacterial agent of the present invention can be used effectively.

The present invention will be described in more detail.
The reason why the active ingredient of the chitosan oligosaccharide-containing antibacterial agent is an oligomer is that, in order to investigate the influence of the degree of polymerization of chitosan, the monomer, oligomer, and polymer were tested. It is because the growth inhibitory effect with respect to Streptococcus was seen.
In the present invention, the degree of polymerization of the oligosaccharide is 2 to 8, and this range is preferred. The chitosan oligomer having a degree of deacetylation of 80% or more only needs to have a degree of polymerization in the above-mentioned range. The reason for limiting the degree of polymerization is that if the degree of polymerization is higher than this, the antibacterial action becomes insufficient.

The degree of deacetylation of chitosan oligosaccharide is preferably 80 to 100%. This is because if the degree of deacetylation is less than 80%, the antibacterial action is insufficient and inconvenient.
Furthermore, the pH range of the chitosan oligosaccharide-containing antibacterial agent is preferably 6.5 or more in order to avoid dentin decalcification. Of the dentine, the enamel that covers the surface is stronger in acid resistance and harder to demineralize than the dentin, but the root surface where the dentin is exposed due to the withdrawal of the gingiva and the damaged enamel When the minute gaps reach the dentin, the effect of caries prevention / suppression is particularly required. In such a situation, the chitosan oligosaccharide-containing antibacterial agent has a pH that does not cause demineralization of the dentin. This is because it is desirable to set the value.

  In the chitosan oligosaccharide-containing antibacterial agent of the present invention, the concentration of chitosan oligosaccharide as an active ingredient is preferably 0.5% by weight or more, but if it is less than 0.5% by weight, the caries prevention / suppression effect is low. Because. Further, the upper limit of the concentration of chitosan oligosaccharide is not particularly limited, but when the concentration is 5% by weight or more, the viscosity is remarkably increased and complete dissolution becomes difficult, which is inconvenient in use.

The chitosan oligosaccharide-containing antibacterial agent of the present invention is liquid, pasty, tablet, etc., and is effective when used in the form of a coating agent, but in the case of liquid, it may be used as a mouthwash. In particular, since caries develops due to resident bacteria in the oral cavity, it can be easily used by gargle regardless of the method of sterilization / disinfection that is a response to general bacterial infections.
However, it is more effective to apply this paste directly to the pits and fissures of the occlusal surface, the adjacent surface, the root surface exposed from the exit of the gingiva, the cervical region or the adjacent surface. . In this case, it is desirable to refrain from contact with saliva by, for example, natural drying for at least 15 minutes after application. This is to maintain a concentration necessary for obtaining a caries bacterial growth inhibitory effect. For the application operation, a cotton swab, cotton, a dental tool, or the like can be used, and a toothpick or the like may be used at home.
Note that the mouthwash is highly available because it can easily prevent and control root caries that are increasing in Japan, which has become a super-aging society.
Furthermore, chitosan also has an immunostimulatory effect, so it is not only useful for swallowing after application or after gargle, but is also beneficial for health promotion. Since the shell is used as a raw material, it is advantageous in terms of cost, and versatility can be expected.

  Furthermore, in order to make a paste, calcium glycerophosphate that is not harmful to the living body (for example, a standard special product manufactured by Nacalai Co., Ltd.) is added to the chitosan oligosaccharide-containing antibacterial agent of the present invention at a concentration of, for example, 5 mM or more. Phosphorus deposits on the tooth and is preferable because the effect of supplying the constituents of the tooth is obtained.

The present invention will be specifically described.
(1) Effect of degree of polymerization and pH value The following chitosan (monomer, oligomer, polymer) powders were dissolved in 0.4% acetic acid aqueous solution, respectively, and then sodium hydroxide was used to adjust the pH to 6.0, pH 6.5. , PH is adjusted to 7.4, and then sterilized by filtration to obtain a chitosan solution having a concentration of 4% by weight.
As a control, an acetic acid solution adjusted to each pH value with sodium hydroxide was used.
For each chitosan solution and control thus obtained, S. Mutans was added at a concentration of 5 × 10 ³ CFU / ml and allowed to react at 37 ° C. for 1 hour. Then, 0.1 ml of this reaction solution was inoculated on Todd-Hewitt agar medium and cultured at 37 ° C. for 48 hours. Thereafter, the number of colonies on the agar medium was counted, and the ratio with the control was calculated by the following formula. Each experiment was performed three times, and the average value of each experiment is shown in Table 1.
Ratio = number of colonies (chitosan solution) / number of colonies (control)

Details of the bacteria, chitosan (manufactured by Koyo Chemical Co., Ltd.), and the medium used in the experiment are as follows.
Cariesogenic bacteria: Streptococcus mutans (GS-5 strain)
Chitosan:
Monomer: Koyoglucosamine (chitosan monomer hydrochloride)
Oligomer: Oligoglucosamine (chitosan oligomer hydrochloride, mixture of 2-8 sugars, degree of deacetylation 100%)
Polymer: SK-2 (chitosan lactate, molecular weight about 10,000, deacetylation degree 80% or more)
Medium: Todd-Hewitt medium

（注）
＊印はコントロールに対して危険率１％で有意差が認められることを示す。
平均値約０．２以下を効果ありと判定した。平均値が０．２であるということは、８０％の細菌発育抑制効果が得られたことを示す。

(note)
An asterisk (*) indicates that a significant difference is observed at a risk rate of 1% relative to the control.
An average value of about 0.2 or less was determined to be effective. An average value of 0.2 indicates that 80% bacterial growth inhibitory effect was obtained.

  As shown in Table 1 and FIG. 1 where this is graphed, at pH 7.4, no monomer, oligomer, or polymer is effective compared to the control, but at pH 6.5 and pH 6.0, It turns out that it is effective in the case of an oligomer.

(2) Influence of degree of polymerization According to the above method, the pH value was set to 6.5, chitosan oligomer was used to prepare chitosan solutions of each concentration, and the number of colonies was measured in the same manner as described above. The ratio was calculated.

  As can be seen from Table 2 and FIG. 2 which is a graph of this, 75% or more caries bacterial growth inhibitory effect is shown at a concentration of 0.5% by weight or more. In particular, at the concentration of 2% by weight, the highest bacterial growth inhibitory effect is shown (minimum growth inhibitory concentration of carious bacteria).

  In the same manner as described above, the influence of the concentration of the monomer and polymer was examined at pH 6.5, and the results are shown in FIGS. 3 and 4, respectively. In either case, it can be seen that there is not much difference compared to the control.

It is a graph which shows the influence of the polymerization degree of chitosan and pH of chitosan oligosaccharide containing antibacterial agent with respect to a bacterial growth inhibitory effect. It is a graph which shows the influence of the density | concentration of a chitosan oligomer with respect to a bacterial growth inhibitory effect. It is a graph which shows the influence of the density | concentration of a chitosan monomer with respect to a bacterial growth inhibitory effect. It is a graph which shows the influence of the density | concentration of a chitosan polymer with respect to a bacterial growth inhibitory effect.

Claims (6)

A chitosan oligosaccharide having a degree of deacetylation of 80 to 100% and a degree of polymerization of 2 to 8 is used as an active ingredient, has a pH value of 6.5 or more, and has an inhibitory effect on the growth of cariogenic bacteria. Antibiotics containing chitosan oligosaccharides. The antibacterial agent according to claim 1, wherein the concentration of the chitosan oligosaccharide is 0.5% by weight or more. The antibacterial agent according to claim 1, further comprising calcium glycerophosphate. The antibacterial agent according to claim 1, wherein the cariogenic bacterium is a mutans bacterium. A method for inhibiting the development of cariogenic bacteria by applying the antibacterial agent according to claim 1. 6. The method according to claim 5, wherein the cariogenic bacterium is a mutans bacterium.

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