JP2014005241A - Antimicrobial material - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide an antibiotic material obtained by immobilizing silver nanoparticles on a solid material.SOLUTION: The present invention is a solid antimicrobial material comprising chitin derivatives with deacetylation degrees of 0% to 50% and silver nanoparticles bound to the chitin derivatives. In the present invention, the deacetylation degrees of chitin derivatives are preferably 5% to 40%. Silver nanoparticles have an average particle diameter of 1 nm to 30 nm. The weight fraction of the chitin derivatives to the antimicrobial material is preferably 1% to 30%. The molecular weights of the chitin derivatives are preferably 10,000 or more.

Description

  The present invention relates to an antimicrobial material, and more particularly to a solid antimicrobial material containing silver nanoparticles. The present invention also relates to an article containing an antimicrobial material and a method for producing the antimicrobial material.

  Unlike simple macro massive metals, metal nanoparticles are known to produce optical, electromagnetic and chemical properties depending on their size and shape. As a method for producing such metal nanoparticles, a method of producing a metal colloid solution containing nano-sized silver nanoparticles under mild conditions using a simple apparatus is known (for example, Patent Document 1). Non-patent documents 1 to 3). Patent Document 1 discloses a technique for controlling the particle size of metal nanoparticles by changing the type and concentration of reducing sugar. Further, as a method for producing a metal colloid, a technique is known in which an aqueous suspension containing a metal ion-containing glass powder and reducing sugar (glucose) is heated under pressure using a general autoclave apparatus or the like ( For example, Patent Document 2).

JP 2010-268694 A JP 2011-157623 A Y. Mori et al. Simple and environmentally friendly preparation and size control of silber nanoparticles using an inhomogeneous system with silver-containing glass powder.J Nanopart Res 13, 2799 (2011) T. Cunha et al. Biological and pharmacological activity of chitosan and derivatives.Chitosan-based systems for biopharmaceuticals: Delivery, targeting and polymer therapeutics, First edition.Edited by Bruno Sarmento and Jose das Neves. By John Wiley & Sons, Ltd. (2012) J.L.Elechiguerra et al. Interaction of silver nanoparticles with HIV-1. J Nanobiotechnol 3, 6 (2005)

  Silver nanoparticles have been reported to have some effects on living organisms, and in particular, biotoxicity to aquatic organisms is feared. It is also known to inhibit the growth of mouse sperm stem cells in mammals. In addition, since silver nanoparticles are colored by surface plasmon resonance, application to antimicrobial materials in a colloidal solution in which silver nanoparticles are dispersed in a solvent is not only harmful to living organisms but also stains such as clothing and household goods. It was difficult to expect.

  Therefore, the application of silver nanoparticles as an antimicrobial agent is limited to a state where the silver nanoparticles are immobilized in a substrate such as a polymer or ceramic. In this case, since the expression of antimicrobial properties is due to selective physical adsorption of the silver nanoparticle part to the microorganism, control of the three-dimensional structure of the silver nanoparticle and the substrate substance is particularly important, and simple and effective production is possible. The technology was unreachable. In particular, it was difficult to disperse in the substrate material while suppressing the coarsening of the silver nanoparticles.

  This invention is made | formed in view of the above background, Comprising: It aims at providing the antibiotic material which fix | immobilized silver nanoparticle to the solid substance. Another object is to provide an article containing an antibiotic material. Another object is to simplify (make easy) the manufacturing method.

In order to solve the above problems, the present invention provides a solid antimicrobial material comprising a chitin derivative having a deacetylation degree of 0% to 50% and silver nanoparticles bonded to the chitin derivative. It is. Here, the chitin derivative is a polysaccharide having a molecular weight of several thousands to several hundred thousand, which includes a plurality of N-acetylglucosamine (C ₈ H ₁₃ NO ₅ ) and glucosamine (C ₆ H ₁₃ NO ₅ ) in a straight chain. is there. N-acetylglucosamine becomes glucosamine by deacetylating the acetyl group. Similar to chitin, chitosan is a polysaccharide having a molecular weight of several thousands to several hundred thousand including a plurality of N-acetylglucosamine and glucosamine in a straight chain, and is distinguished from chitin by the degree of deacetylation. The degree of deacetylation is defined as the ratio of the number of glucosamines to the total number of N-acetylglucosamines and glucosamines contained in the chitin derivative. A chitin derivative is defined as chitin having a degree of deacetylation of 0 to 50% and chitosan having a deacetylation degree of 51% to 100%. Moreover, the N-acetylglucosamine and glucosamine contained in the chitin derivative may have an additional functional group. Chitin derivatives include, for example, various compounds such as N-acetylglucosamine and a carboxy compound of a hydroxyl group of glucosamine, an alkoxy compound, an adduct of alkylene glycol, an acyloxy compound, a carbamate compound, and specifically, for example, carboxymethyl chitin (CMCT).

  According to this configuration, a chitin derivative having a degree of deacetylation of 0% to 50% becomes a solid at room temperature (20 ° C. ± 15 ° C.). Can be obtained. Since chitin has a positive charge, chlorides and oxides are formed on the surface and can bind to silver nanoparticles having a negative charge. In particular, since chitin can be combined with silver nanoparticles by adding to a colloidal solution containing silver nanoparticles, it is easy to form a chitin / silver nanoparticle composite. In addition, the chitin / silver nanoparticle composite is stable as a solid, so it can be formed into sheets, sponges (porous), etc., increasing the contact area between silver nanoparticles and microorganisms Can be made.

  In the said invention, it is preferable that the deacetylation degree of the said chitin derivative is 5%-40%.

  According to this configuration, the solid form of the chitin derivative is stabilized.

  In the above invention, the silver nanoparticles preferably have an average particle size of 1 nm to 30 nm and a weight fraction of 1% to 30% with respect to the antimicrobial material.

  According to this configuration, the antibiotic activity of the antibiotic material is increased.

  In the above invention, the chitin derivative preferably has a molecular weight of 10,000 or more.

  According to this configuration, the solid form of the chitin derivative is stabilized.

  You may comprise the article which has antimicrobial property from the resin containing the said antimicrobial material in the said invention. The resin may be a known resin such as polyethylene terephthalate, polyacetal, polycarbonate, polyamide, etc., and the article may be a known plastic molded part such as an automotive instrument panel, steering wheel, air conditioner filter or duct. It's okay.

  According to this configuration, antibiotic properties can be imparted to various known resin articles.

  Another aspect of the present invention is a method for producing an antimicrobial material, comprising heating a water-based suspension containing silver ion-containing glass powder and reducing sugar, and eluting the silver ion eluted into the water-based suspension. A first step of obtaining a silver nanoparticle colloidal solution by reducing the above, a chitin derivative having a deacetylation degree of 0 to 50% added to the silver nanoparticle colloidal solution, and the silver nanoparticle bound to the chitin derivative A second step of obtaining a silver nanoparticle composite, and a solid chitin derivative to which the silver nanoparticles are bound by separating the chitin / silver nanoparticle composite from the aqueous suspension and drying And a third step of obtaining a solid material.

  According to this configuration, by using the chitin derivative, a chitin / silver nanoparticle composite can be formed while suppressing the coarsening of the silver nanoparticles from the silver nanoparticle colloid solution. In particular, in a silver nanoparticle colloidal solution obtained by reducing silver ions using a reducing sugar, it is difficult to separate the silver nanoparticles from the reducing sugar, but the use of a chitin derivative facilitates the removal of the reducing sugar. Moreover, according to this manufacturing method, the particle size of the silver nanoparticles can be easily controlled.

  In the above invention, it is preferable that the reducing sugar is caramelized by heating in the first step.

  According to this configuration, in the silver nanoparticle colloid solution, the caramelized reducing sugar protects the silver nanoparticles, and the coarsening of the silver nanoparticles is suppressed.

  According to the above structure, the antibiotic material which fixed the silver nanoparticle to the solid substance can be provided. In addition, an article containing an antibiotic material can be provided. In addition, the manufacturing method can be simplified (facilitated).

TEM and SEM observation image of chitin / silver nanoparticle composite according to this embodiment UV-vis spectrum of the supernatant obtained by centrifugal washing of chitin / silver nanoparticle complex with PBS Graph showing the correlation between chitin weight concentration and adsorbed silver nanoparticles Graph showing antiviral activity evaluation results of chitin / silver nanoparticle composite The graph which shows the antibacterial activity evaluation result of chitin / silver nanoparticle composite or chitosan / silver nanoparticle composite

  The antimicrobial material comprising the chitin / silver nanoparticle composite according to this embodiment is one in which silver nanoparticles are bound to a chitin derivative having an acetylation degree of 0 to 50%. The chitin derivative is a polysaccharide containing a plurality of N-acetylglucosamine and glucosamine in a linear form, and N-acetylglucosamine and glucosamine may additionally have any functional group. Chitin derivatives include, for example, various compounds such as N-acetylglucosamine and a carboxy compound of a hydroxyl group of glucosamine, an alkoxy compound, an adduct of alkylene glycol, an acyloxy compound, a carbamate compound, and specifically, for example, carboxymethyl chitin (CMCT).

  Silver nanoparticles have negative charges due to adsorption of silver halides and oxides on their surfaces. On the other hand, chitin derivatives have a positive charge in an aqueous solution. Therefore, the silver nanoparticles and the chitin derivative are bonded to each other by electrostatic interaction.

  A chitin derivative having an acetylation degree of 0 to 50% is a solid at room temperature (20 ° C. ± 15 ° C.). Therefore, the chitin / silver nanoparticle composite is solid at room temperature and maintains a predetermined shape. A chitosan derivative having a degree of acetylation greater than 50% is partially liquefied (gelled) and cannot maintain a stable shape. A chitin / silver nanoparticle composite containing a chitin derivative having an acetylation degree of 0% to 50% can take various forms such as powder, sponge, sheet, film, and gel. In addition, the chitin / silver nanoparticle composite can be used alone or mixed with other resin materials as an injection molding material, and can be replaced with various known plastic articles. For example, the chitin / silver nanoparticle composite can be used for various well-known plastic products such as instrument panels and steering wheels of automobiles, casings of telephones and televisions, filter materials and ducts incorporated in air conditioners. it can. Moreover, the mixture of a carboxymethyl chitin / silver nanoparticle composite and polyvinyl alcohol can be used as a hydrogel.

  It has been confirmed that the bond between the chitin derivative and the silver nanoparticles tends to be weaker as the deacetylation of the chitin derivative is lower. Therefore, the chitin derivative is more preferable as the degree of deacetylation is higher in terms of the binding property with the silver nanoparticles. On the other hand, chitin is more stable in solid form as the degree of acetylation is lower, and can take a more stable form at 40% or less. From the above, the degree of acetylation of the chitin derivative is 0% to 50%, more preferably 5% to 40%, and still more preferably, from the viewpoint of the stability of the solid form and the binding property with the silver nanoparticles. It is good that it is 30 to 35%.

  Chitosan derivatives are known to have wound healing promoting activity as well as antifungal activity and antibacterial activity. This is thought to be because a chitin derivative having a positive charge strongly interacts with the cell membrane of a negatively charged fungus or fungus, causing cell membrane changes, inhibition of migration, and elution of cytoplasmic components (non-patented). Reference 2). The chitin derivatives have the above characteristics as well, but it is known that these antifungal properties and the like become higher as the degree of deacetylation increases. This is considered to result from the fact that the positive charge of the chitin derivative becomes stronger as the degree of deacetylation increases. In addition, chitosan derivatives and chitin derivatives alone have not been confirmed to have antiviral activity.

  The chitin derivative is greatly enhanced in antifungal and antibacterial activities by combining silver nanoparticles into a chitin / silver nanoparticle composite. Further, since the silver nanoparticles have antiviral activity, the chitin / silver nanoparticle composite has antiviral activity.

  The size of the silver nanoparticles contained in the chitin / silver nanoparticle composite (diameter when approximated to a spherical shape) is preferably smaller than the micro order when obtaining materials for antibacterial applications. In the case of obtaining, it is preferably 50 nm or less, more preferably 1 to 30 nm, still more preferably 2 to 20 nm.

  The chitin derivative contained in the chitin / silver nanoparticle composite preferably has a molecular weight of 5,000 or more, more preferably 10,000 to 300,000.

  The chitin / silver nanoparticle composite according to the embodiment is applied to various uses such as medical use and industrial use. Silver nanoparticles exhibit optical, electromagnetic, and chemical properties that are different from bulk metals by reducing them to nano-size. For example, silver nanoparticles of 10 nm or less have been found to selectively adsorb to the HIV-1 virus envelope and inactivate the virus (Non-patent Document 3). The chitin / silver nanoparticle composite exhibits strong antiviral activity with good stability. Examples of the virus include A (H1N1) influenza virus.

  Below, the manufacturing method of a chitin / silver nanoparticle composite is demonstrated. The chitin / silver nanoparticle composite is formed by adding a -chitin derivative to the prepared silver nanoparticle colloid and purified to a solid state by removing the solution.

  The silver nanoparticle colloid is produced by pressurizing and heating an aqueous suspension containing silver ion-containing glass powder and reducing sugar (glucose) using a general autoclave apparatus or the like. Silver nanoparticles produced by this method have a constant particle size and good storage stability. In heating the suspension in this manner, the glucose is heated, decomposes and binds irregularly, resulting in a brown candy-like substance that is caramelized.

  The particle size of the silver ion-containing glass powder is preferably 2 to 20 μm. The smaller the particle size of the silver ion-containing glass powder, the faster the elution rate of silver ions and the easier it is to obtain the target metal colloid. The preferable lower limit of the particle size of the glass powder is based on the ease of handling and cost of the powder, and it is also preferable to consider that the outflow rate of silver ions does not become too fast. Moreover, the ratio of the silver ion contained in a silver ion containing glass powder does not have limitation in particular, Preferably it is 1-3 weight%. Within this range, metal ions are eluted at a preferred rate in the aqueous suspension. Further, it is typically contained in the glass component in the form of monovalent silver ions, and specifically, it may be contained in the glass powder in the form of silver nitrate or silver oxide.

  A reducing sugar is a sugar that forms an aldehyde group and a ketone group in a basic solution. Examples of reducing sugars include all reducing monosaccharides such as glucose, fructose and glyceraldehyde, and reducing disaccharides and oligosaccharides such as lactose, arabinose and maltose. A reducing sugar having a reducing power sufficient to reduce the eluted silver ions can be used without particular limitation, and glucose, which is a monosaccharide, may be used from the viewpoints of availability, water solubility, and the like. In the suspension autoclave, glucose as a reducing sugar decomposes and binds irregularly, resulting in a brown candy-like substance (caramel) that is caramelized. The produced caramel inhibits the reduced silver nanoparticles from growing into coarse particles and stabilizes them as fine silver nanoparticles.

  In this embodiment, the metal reduction reaction is performed in an aqueous solvent. The aqueous solvent is at least 50% water, preferably water alone. Silver ions are reduced by oxidation-reduction reaction between glucose and silver ions to form silver nanoparticles, and a silver nanoparticle colloidal solution is obtained. In this redox reaction, it is desirable that the glucose is in a large excess. Specifically, the concentration of reducing sugar in the aqueous solvent was 0.2 to 8 wt%. According to the present inventors' new knowledge, the size of the metal particles in the obtained metal colloid is approximately proportional to the square root of the amount of reducing sugar in the reaction system. Therefore, the amount of reducing sugar used can be a factor controlling the size of the metal particles in the metal colloid.

  In this embodiment, reducing sugar is dissolved in an aqueous solvent, and further, a metal-containing glass powder is dispersed to obtain an aqueous suspension (hereinafter also simply referred to as a suspension), and the suspension is heated. . A preferred example of heating is to continue heating the suspension in a closed system. According to this, since water vapor | steam produces | generates and a system internal pressure rises, the heating in a pressurized state is attained. Such pressure heating can be easily realized by using a general autoclave apparatus or the like.

  Glucose as a reducing sugar is decomposed by pressurization and heating of the suspension, and as a result of binding irregularly, it turns into a brown candy-like substance (caramelizes). The produced caramel inhibits the reduced and produced silver nanoparticles from growing into coarse particles and stabilizes them as fine silver nanoparticles. In this embodiment, unlike the conventional method, no special additive is required in the oxidation-reduction reaction. This is because caramel derived from reducing sugar is an alternative to the additive.

  The preferred conditions of the autoclave for producing the silver nanoparticle colloidal solution are a pressure of 180 to 230 kPa, a temperature of 110 to 130 ° C., and a pressure and heating time of 10 to 25 minutes. Within the above range, as a result of maintaining a suitable balance between the reaction rate and the rate of caramelization of glucose, the silver nanoparticle colloid has a constant particle size and further improved storage stability. In this embodiment, colloidal silver nanoparticles can be produced under relatively mild reaction conditions.

  After completion of the reaction by pressurizing and heating the suspension, the glass powder is removed by an appropriate technique such as centrifugation to obtain a silver nanoparticle colloidal solution. Silver nanoparticles contained in the colloidal solution of silver nanoparticles tend to be a spherical shape with a constant shape because the produced caramel acts as a protective agent, and the size (diameter approximated to a spherical shape) is 2 to 30 nm. is there. The smaller the size of the silver nanoparticles, the smaller the size variation and the higher the stability. In this embodiment, unlike the conventional method, it is not necessary to use a stabilizer such as poly (N-vinyl-2-pyrrolidone) when storing the metal colloid solution.

  Next, a chitin derivative having an acetylation degree of 0% to 50% is added to the silver nanoparticle colloid solution and stirred. The chitin derivative preferably has a molecular weight of 10,000 or more. As a result, the silver nanoparticles are bound to the chitin derivative by electrostatic interaction to form a chitin / silver nanoparticle composite and separated from the caramel. The interaction between the silver nanoparticles and the chitin derivative depends on the concentration and the degree of deacetylation. In other embodiments, a chitin derivative may be added before removing the glass powder from the silver nanoparticle colloid solution.

  The suspension containing the chitin / silver nanoparticle complex is subjected to centrifugation, dialysis, filtration and the like to remove caramel. Thereafter, the solvent is removed by freeze-drying the suspension to obtain a chitin / silver nanoparticle composite. In addition, when purifying a chitin / silver nanoparticle complex by filtration and washing with water instead of freeze-drying, 15 to 80% of silver nanoparticles depending on the degree of deacetylation of the chitin derivative by washing with water May leak.

  According to the above production method, caramel can be easily removed from the silver nanoparticles protected by caramel by adding the chitin derivative to the suspension. Further, the chitin derivative has an effect of suppressing the coarsening of the silver nanoparticles, and the generated silver nanoparticles can be stably stored.

  Examples according to the present invention will be described below. However, the present invention is not limited to the embodiments described in these examples.

[Example 1]
Silver-containing glass powder having a composition of SiO ₂ (about 25%)-B ₂ O ₃ (about 60%)-Na ₂ O (about 15%)-AgNO ₃ (1.6%) (Environmental Science, model number BSP21, D-glucose (> 98%, Wako Pure Chemical Industries Ltd.) at a predetermined concentration (0.25 wt%, 1.0 wt%, 4.0 wt%) in a 100 mL glass vial with 0.50 g of an average particle size of 10 μm) Manufactured) was dispersed in 50 mL of an aqueous solution. The obtained suspension was subjected to pressure heat treatment at 121 ° C. and 200 kPa (about 20 minutes) in an IMC-30L autoclave device manufactured by Ikemoto. Thereafter, the suspension was gradually cooled to room temperature and subjected to centrifugation at 3000 rpm for 10 minutes. At this time, the solvent portion (supernatant) containing glucose was yellowish brown. The supernatant containing the silver nanoparticle colloid was collected as a silver nanoparticle colloid solution and stored in the dark at room temperature. A predetermined weight of chitin (Yaizu Suisan Chemical Co., Ltd.) having a deacetylation degree of 30% or 5% or less was added to the silver nanoparticle colloid solution, and the mixture was allowed to react at room temperature for 1 hour with stirring. The chitin used here is composed of N-acetylglucosamine and glucosamine, and has no additional functional group. After the reaction, the solution is centrifuged twice (3,000 rpm, 10 min) using PBS (phosphate buffered saline) to remove the produced caramel and wash the produced chitin / silver nanoparticle composite. did. The washed chitin / silver nanoparticle composite was freeze-dried to obtain a solid chitin / silver nanoparticle composite having a stable shape. The obtained solid chitin / silver nanoparticle composite is taken as Example 1.

  In the above examples, the amount of D-glucose corresponds to 25 times or more of the equivalent of reducing silver ions in the case of 0.25% by weight, and corresponds to 100 times or more in the case of 1.0% by weight. The case of 4.0% by weight corresponds to 400 times or more.

[Comparative Example 1]
In the above Example 1, chitin added in the production process was changed to chitosan (Yaizu Suisan Chemical Co., Ltd.) having an acetylation degree of 84%, and the produced chitosan / silver nanoparticle composite was used as Comparative Example 1. .

(Appearance observation)
It was visually observed that the chitin / silver nanoparticle composite according to Example 1 was a stable solid at room temperature (20 ° C. ± 15 ° C.). On the other hand, the chitosan / silver nanoparticle composite according to Comparative Example 1 was visually observed to be a liquid (gel) at room temperature (20 ° C. ± 15 ° C.).

(Microscopic observation)
After freeze-drying the chitin / silver nanoparticle composite obtained as Example 1, TEM and SEM images of the chitin / silver nanoparticle composite stored at room temperature for 1 month were observed (JEOL JEM-1010, 80 kV) (FIG. 1). As shown in the TEM image of FIG. 1A, the silver nanoparticles in the composite were spherical and uniformly dispersed. The scale bar in the TEM observation image is 100 nm, and the scale bar in the SEM observation image is 1 μm. The particle size of the silver nanoparticles was measured from a TEM observation image photograph using particle size analysis software. As a result, when the concentration of D-glucose was 0.25 wt% in the production process, the average particle diameter (diameter) of the silver nanoparticles was 3.48 ± 1.83 nm, and the concentration of D-glucose was 1.00 wt. %, The silver nanoparticles have a particle size of 6.53 ± 1.78 nm, and those having a D-glucose concentration of 4.00% by weight have a silver nanoparticle size of 12.9 ± 2. It was 50 nm.

(UV-vis spectrum)
The results obtained by measuring the UV-vis spectrum (Hitachi, U-3300) of the supernatant obtained during centrifugal washing of the chitin / silver nanoparticle composite colloidal solution of Example 1 are shown in FIGS. C). In FIG. 2, the amount of chitin added with a deacetylation degree of 30% was changed for samples having a D-glucose concentration of 0.25 wt%, 1.0 wt%, and 4.0 wt%. Results are shown. The UV-vis spectrum shown in FIG. 2 is obtained by measuring the supernatant immediately after washing by centrifugation. In each figure of FIG. 2, the peak around the wavelength of 400 nm is attributed to the silver nanoparticles. It was confirmed that the peak due to the silver nanoparticles increases as the concentration of D-glucose increases, that is, as the particle size of the silver nanoparticles increases. Moreover, in each figure, it was confirmed that a peak reduces, so that the addition amount of chitin increases. This indicates that as the amount of chitin added increases, the silver nanoparticles that combine with chitin to form a chitin / silver nanoparticle composite increase. Since the chitin / silver nanoparticle complex is removed from the supernatant by centrifugation, the concentration of silver nanoparticles in the supernatant decreases.

  FIG. 3 shows that the amount of chitin added with a degree of deacetylation of 30% was changed for samples having a D-glucose concentration of 0.25 wt%, 1.0 wt%, and 4.0 wt%. The absorbance at a wavelength of 400 nm is shown. From this change in absorbance, the weight of silver nanoparticles bound (adsorbed) to 1 mg / mL chitin (degree of deacetylation: 30%) was calculated. The results are shown in Table 1 below. Similarly, the weight of silver nanoparticles bound to 1 mg / 1 mL of chitosan (acetylation degree: 84%) and 1 mg / 1 mL of chitin (acetylation degree: 5% or less) was calculated. Show. As can be seen from Table 1, the higher the degree of deacetylation, the greater the weight of the bound silver nanoparticles.

[Example 2]
Each silver obtained by dissolving 10 mg of chitin having a degree of deacetylation (Yaizu Suisan Chemical Co., Ltd., average molecular weight: 54000) in 1 mL of water under acidic conditions (pH 2) and following the procedure of Example 1 Nanoparticle colloidal solutions (average particle size: 12.9 ± 2.50 nm, 6.53 ± 1.78 nm and 3.48 ± 1.83 nm) were added and stirred well. Then, it neutralized by addition of NaOH and the chitin / silver nanoparticle composite was prepared as a precipitate. This complex was washed several times with saline.

(Antiviral activity evaluation)
Influenza fluid of A / PR / 8/34 strain (H1N1 type) is added to the chitin / silver nanoparticle complex, the resulting suspension is stirred for 1 hour at room temperature, and the virus is adsorbed by centrifugation. The supernatant of the supernatant was added to a 96-well plate containing infected cells (MDCK) by serial dilution and cultured for 7 days. The number of wells in which plaque was formed was counted and evaluated based on the TCID50 method represented by the following formula 1.

In Equation 1, c is the concentration of the first stage virus, p is the number of plaque-formed wells, w is the number of wells per concentration, and v is the volume of virus solution per well.

  FIG. 4 shows the evaluation results of antiviral activity. The vertical axis in FIG. 4 represents the degree of influenza infection. The degree of influenza infection in the case of chitin without silver nanoparticles was normalized as 100% on the vertical axis. Here, the anti-influenza activity could not be observed without containing silver nanoparticles in 30% deacetylated chitin. Therefore, if the value on the vertical axis is smaller than 100%, the anti-influenza activity is higher than that of chitin alone. Moreover, it can be said that anti-influenza activity is so high that the value of a vertical axis | shaft is small. The horizontal axis shows the content of silver particles with respect to 1 mg of chitin. As plotted in FIG. 4, the complexes containing silver nanoparticles, without exception, improved anti-influenza activity over chitin alone. It was found that the smaller the average particle size of the silver nanoparticles, the higher the anti-influenza activity.

[Example 3 and Comparative Example 3]
The silver nanoparticle colloidal solution produced according to the process of Example 1 (three kinds of average particle diameters of 12.9 ± 2.50 nm, 6.53 ± 1.78 nm and 3.48 ± 1.83 nm) was deacetylated. A chitin sponge fragment (diameter 5 mm × thickness 1 mm) having a degree of 30% or 5% or less was added and stirred. Each sponge fragment was washed three times by centrifugation using PBS and then lyophilized. These are referred to as Example 3. As a comparative object, Comparative Example 3 was prepared by using a sponge fragment (diameter 5 mm × thickness 1 mm) made of photocured chitosan having a deacetylation degree of 84% instead of the sponge fragment of Example 3.

(Antifungal activity test)
50 mL of an E. coli (strain B, ATCC no. 11303) suspension (3 × ^{10 10} colony forming units (CFU) / mL in PBS) was added dropwise to each sponge fragment of Example 3 and Comparative Example 3 for 15 minutes at room temperature. I left it alone. Each sponge fragment to which the E. coli was dropped was cultured in 1 mL of a cell culture medium (NB: Nutrient Broth) for 24 hours at 37 ° C. under vibration. The bacterial cell culture solution containing each sponge fragment was stirred for 1 minute by a vortex mixer, and then the absorbance (OD650) at a wavelength of 650 nm of the bacterial cell culture solution from which each sponge fragment was removed was measured. The results are shown in FIG.

  In each figure of FIG. 5, the rightmost part is the absorbance of the bacterial cell culture solution only (background, no bacteria), and the left side does not include the sponges of Example 2 and Comparative Example 2 and is cultured under the same conditions. Absorbance of bacterial culture containing E. coli (bacteria only), bacterial culture containing E. coli when using sponges according to Example 2 and Comparative Example 2 that do not contain silver nanoparticles on the left side (1 mg chitin or chitosan). From FIG. 5, the sponge made of chitin with a deacetylation degree of 5% or less and 30% and the sponge made of chitosan with a deacetylation degree of 84% are both antifungal, even if they do not contain silver nanoparticles. It turns out that it has activity. And it was confirmed that the antifungal activity of the sponges according to Example 2 and Comparative Example 2 in which silver nanoparticles were bonded to these sponges was further increased. It can be seen that the sponges according to Example 2 and Comparative Example 2 have higher antifungal activity as the degree of deacetylation of chitin or chitosan constituting the sponge is higher. From FIG. 5, it was confirmed that the silver nanoparticles had higher antifungal activity as the average particle size was smaller, and higher antifungal activity as the concentration was higher.

  From the results of FIG. 5, it was confirmed that the chitin sponge had a lower antifungal activity than the chitosan sponge, but the antifungal activity was increased by binding silver nanoparticles. A chitin sponge can be said to be more versatile than chitosan which requires a photo-curing treatment because it can stably maintain its form at room temperature.

  Although the description of the specific embodiment is finished as described above, the present invention is not limited to the above embodiment and can be widely modified.

Claims (7)

A chitin derivative having a degree of deacetylation of 0% to 50%,
A solid antimicrobial material comprising silver nanoparticles bonded to the chitin derivative.   The antimicrobial material according to claim 1, wherein the chitin derivative has a degree of deacetylation of 5% to 40%.   3. The antimicrobial material according to claim 1, wherein the silver nanoparticles have an average particle diameter of 1 nm to 30 nm and a weight fraction with respect to the antimicrobial material of 1% to 30%. .   The antimicrobial material according to any one of claims 1 to 3, wherein the chitin derivative has a molecular weight of 10,000 to 300,000.   An article having antimicrobial properties composed of a resin containing the antimicrobial material according to claim 1. A first step of heating a water-based suspension containing silver ion-containing glass powder and reducing sugar to reduce the silver ions eluted in the water-based suspension to obtain a silver nanoparticle colloidal solution;
A second step of adding a chitin derivative having a deacetylation degree of 0 to 50% to the silver nanoparticle colloid solution to obtain a chitin / silver nanoparticle composite in which the silver nanoparticles are bound to the chitin derivative;
Separating the chitin / silver nanoparticle composite from the aqueous suspension and drying to obtain a solid material containing a solid chitin derivative to which the silver nanoparticles are bound. A method for producing an antimicrobial material.   In the said 1st process, the said reducing sugar is caramelized by heating, The manufacturing method of the antimicrobial material of Claim 6 characterized by the above-mentioned.