JP2023022689A - Antibacterial/antifouling material and antibacterial/antifouling surface treatment method - Google Patents

Abstract

To provide an antibacterial/antifouling material with antibacterial property and antifouling property.SOLUTION: An antibacterial/antifouling material has an antifouling layer and antibacterial peptide immobilized to the antifouling layer on a material surface. The antifouling layer includes: a monomer unit (A) having at least one of an amphoteric ionic structure and an ether structure in a side chain; a monomer unit (B) having a hydrophobic group containing a Si atom in a side chain; and a monomer unit (C) having a trialkoxysilyl group in a side chain. The monomer unit (A) contains a copolymer forming block segments.SELECTED DRAWING: Figure 1

Description

TECHNICAL FIELD The present disclosure relates to an antibacterial antifouling material and an antibacterial antifouling surface treatment method.

It is required to impart antifouling properties to materials for instruments and facilities used in a wide range of fields such as medical care, construction, environment and hygiene. For example, many ceramic materials are used in plumbing equipment, and since such plumbing equipment is exposed to various stains on a daily basis, it is necessary to impart antifouling properties to the surface of the materials used there. be.

In response to the above problems, a polymer-type surface modification that does not require pretreatment such as immersion in water, can easily hydrophilize the surface of materials such as ceramics, and can impart highly durable antifouling properties. Agents and surface treatment means have been disclosed (see, for example, Patent Document 1).

JP 2017-036419 A

By the way, in recent years, from the viewpoint of hygiene management such as measures to prevent the spread of infectious diseases, it is required to impart antibacterial properties in addition to antifouling properties to the surfaces of materials used in plumbing equipment and the like. However, in the conventional technology, when the material surface is simply imparted with antibacterial properties, it is possible to sterilize some of the viable bacteria, but there is a problem that the remaining viable bacteria cannot be washed off. Moreover, when simply imparting antifouling properties to the material surface, there is a problem that the desired antibacterial properties cannot be obtained. Therefore, the current situation is that materials with antibacterial and antifouling properties have not been sufficiently studied.

The present disclosure has been made in view of the above, and an object thereof is to provide an antibacterial and antifouling material imparted with antibacterial and antifouling properties.

The present disclosure has an antifouling layer and an antibacterial peptide immobilized on the antifouling layer on the surface of the material, and the antifouling layer has at least a zwitterionic structure and an ether structure in the side chain. a monomer unit (A) having either; a monomer unit (B) having a hydrophobic group containing an Si atom in its side chain; and a monomer unit (C) having a trialkoxysilyl group in its side chain, wherein the monomer unit (A) relates to an antibacterial and antifouling material containing a copolymer forming a block segment.

1 is a photograph showing the evaluation results of the antibacterial and antifouling results of the antibacterial and antifouling material of Example 1. FIG. 4 is a photograph showing the evaluation results of antibacterial and antifouling results of the antibacterial and antifouling material of Example 2. FIG. 3 is a photograph showing the evaluation results of antibacterial and antifouling results of the antibacterial and antifouling material of Example 3. FIG. 4 is a photograph showing the evaluation results of the antibacterial and antifouling results of the antibacterial and antifouling material of Example 4. FIG. 4 is a photograph showing the results of evaluating the antibacterial and antifouling results of the antibacterial and antifouling material of Comparative Example 1. FIG. 3 is a photograph showing the evaluation results of antibacterial and antifouling results of the antibacterial and antifouling material of Comparative Example 2. FIG. 3 is a photograph showing the evaluation results of antibacterial and antifouling results of the antibacterial and antifouling material of Comparative Example 3. FIG.

<Antibacterial antifouling material>
The antimicrobial antifouling material according to this embodiment has an antifouling layer and an antimicrobial peptide on the surface of the material, and the antimicrobial peptide is immobilized on the antifouling layer. The antifouling layer is formed by applying the surface modifier described below to the surface of the material.

(Surface modifier)
The surface modifier according to the present embodiment includes (A) a monomer unit having at least one of a zwitterionic structure and an ether structure in a side chain; (B) a monomer unit having a hydrophobic group containing a Si atom in a side chain. and (C) a copolymer having a monomer unit having a trialkoxysilyl group in its side chain, wherein the monomer unit (A) forms a block segment.

[Copolymer]
The above copolymer, which is the main component of the surface modifier, is a polymer obtained by copolymerizing the above monomer units (A) to (C), that is, a so-called terpolymer. Furthermore, in the copolymer, the monomer unit (A) forms a block segment and corresponds to a so-called block polymer. On the other hand, the monomer units (B) and (C) do not necessarily have to form a block segment, and may be combined randomly or in a manner having some regularity or periodicity.

That is, the copolymer may be a copolymer in the form of "A-block-B-block-C", "A-block-(B-random-C)", "A-block -(B-stat-C)”, “A-block-(B-alt-C)”, etc. may be copolymers. Preferably, the monomer units (B) and (C) are randomly polymerized so that the copolymer has a structure of "A-block-(B-random-C)". As a result, on the surface of the material treated with the surface modifier of the present embodiment, the adhesive portions and the like due to the monomer unit (C) are evenly present without agglomeration, and the hydrophilic block segments due to the monomer unit (A) are present in the material. By being dispersed on the surface, a stable and uniform surface coating can be obtained.

The above copolymer may contain monomer units other than the above monomer units (A) to (C). The other monomer units are typically bonded randomly, but may have some regularity or periodicity. For example, a mode of forming a segment of a statistical polymer, an alternating polymer, a periodic polymer, a block polymer, and, in some cases, a mode of forming a segment of a graft polymer can be mentioned.

The monomer unit (A) has at least one of a zwitterionic structure and an ether structure in its side chain. The monomer unit (A) forms a hydrophilic block segment (hydrophilic portion) in the copolymer and imparts hydrophilicity to the material surface after surface modification. Examples of the amphoteric ion structure and ether structure that impart hydrophilicity include amphoteric ion structures such as phosphorylcholine groups (PC groups) and betaine groups, and ether structures such as polyethylene glycol chains. The monomer unit (A) may have both a zwitterionic structure and an ether structure.

The phosphorylcholine group (PC group) is a polar group having the same structure as the polar group of phospholipid (phosphatidylcholine), which is the main component of biological membranes. Therefore, by including a PC group in the side chain, the copolymer has hydrophilicity (wettability), specifically, extremely good biocompatibility possessed by the surface of a biological membrane, particularly non-adsorption of biomolecules. , and non-activating properties are imparted, and non-specific adsorption to various molecules such as proteins and blood cells can be effectively suppressed. This makes it possible to impart excellent antifouling properties to the surface of the material to be treated.

The monomer unit (B) has a hydrophobic group containing a Si atom in its side chain. Hydrophobic groups containing Si atoms mainly function to increase the film-forming properties of coatings during surface treatment, and also improve the stability of the coating surface by physically adsorbing to the material surface through hydrophobic interactions. contribute. The hydrophobic group containing Si atoms is, for example, a substituent having an alkylsilyl group or an alkylsilyloxysilyl group, preferably a tris(trialkylsilyloxy)silyl group, more preferably a tris(trimethylsilyloxy)silyl or a tris(triethylsilyloxy)silyl group.

The monomer unit (C) forms a covalent bond (--SiO-- bond) by dehydration condensation through a chemical reaction such as a silane coupling reaction with an OH group present on the material surface, and chemically adsorbs onto the material surface. It has a functional group on the side chain. The functional group is preferably a trialkoxysilyl group. The monomer units (B) and (C) improve the stability and uniformity of the surface coating film of the copolymer.

The skeletal portion of the monomer unit forming the main chain (skeleton) of the copolymer is not particularly limited as long as it can polymerize with each other to form a polymer. is preferably, for example, vinyl-based monomer residues, acetylene-based monomer residues, ester-based monomer residues, amide-based monomer residues, ether-based monomer residues, urethane-based monomer residues, etc. Among these, vinyl-based monomer residues Residues are more preferred. The vinyl-based monomer residue is not particularly limited. Among these, an acryloxy group and a methacryloxy group are particularly preferred. The skeletal moieties can be the same for each monomer unit, or can be independently different, but the embodiment in which they are all the same is preferred. Therefore, the main chain in the above copolymer preferably has a skeleton selected from polyalkyl acrylate or polyalkyl methacrylate.

Specific examples of the copolymer are not limited thereto, but preferred examples thereof include polymers having a structure represented by the following formula (I).

In formula (I), monomer unit (A) is MPC (2-methacryloyloxyethylphosphorylcholine), monomer unit (B) is MPTSSi (3-(methacryloyloxy)propyltris(trimethylsilyloxy)silane), and the monomer Unit (C) is MPTMSi (3-methacryloyloxytrimethoxylsilane). Each of n, m, and l independently represents an integer of 2 or more, and each is 2,000 or less, preferably 1,000 or less.

The monomer composition ratio in the copolymer is preferably in the range of (A):(B):(C) = 2:1:1 to 9:1:1 for the monomer units (A) to (C). is the molar ratio of More preferably, the molar ratio of (A):(B):(C)=4:1:1 to 8:1:1. An increase in the proportion of the monomer unit (A) is preferable in that the hysteresis of the dynamic contact angle of the material surface is reduced. It is considered that this is because the region of the copolymer that physically and chemically bonds to the surface of the material can be sufficiently covered with the hydrophilic portion derived from the monomer unit (A).

The weight average molecular weight (Mw) of the copolymer is not particularly limited, but is preferably 1.0×10 ⁴ to 5.0×10 ⁵ and more preferably 5.0×10 ⁴ to 1.0. ^x105 .

The ratio of the weight average molecular weight to the number average molecular weight (Mw/Mn) in the copolymer is not particularly limited, but is preferably in the range of 1.0 to 2.0, more preferably 1.0 to 1. .5.

The above copolymer may contain structural units derived from monomers other than the monomer units (A) to (C), if necessary. The ratio of the structural units derived from the other monomer is not particularly limited, but is preferably 30 mol% or less, more preferably 10 mol% or less, relative to the total structural units constituting the polymer.

Synthesis of the above copolymer can be carried out by a conventional method based on the technical level of those skilled in the art, but from the viewpoint that it is suitable for obtaining the block segment of the above monomer unit (A), the polymerization method is as follows. Preferably, a living radical polymerization method can be used, more preferably a reversible addition-fragmentation chain transfer (RAFT) polymerization method can be used. Here, the RAFT polymerization method is a radical polymerization reaction using a thiocarbonyl compound as a chain transfer agent (RAFT agent), and since a RAFT residue is present at the end of the polymerization at the end of the polymerization, a new monomer is added to polymerize. A block copolymer can be obtained by initiating Such a RAFT polymerization method does not require the use of a metal catalyst, is easy to operate, and has advantages such as a wide range of applicable monomer species.

Synthesis of the monomer compound used as the above-mentioned monomer unit can be carried out by a conventional method based on the technical level of those skilled in the art.

The surface modifier is preferably in the form of a solution and may contain a solvent. As the solvent, a polar solvent such as alcohol is preferable, but any solvent in which the copolymer can be dissolved can be appropriately changed depending on the application, material, and the like. Methanol, ethanol, or 1-propanol is preferred, and ethanol is more preferred. The concentration of the surface-modifying copolymer of the present invention contained as a main component is, for example, preferably 0.01 to 3.0 weight percent, more preferably 0.05 to 2.0 weight percent, still more preferably 0.05 to 2.0 weight percent. 1 to 1.0 weight percent.

The surface modifier may contain components other than those described above. For example, it may contain any other component that is generally used as a component of a substrate surface modifier.

[Antibacterial peptide]
The antimicrobial peptide according to this embodiment has antimicrobial properties. As used herein, the term "antibacterial" means killing microorganisms including bacteria, yeast, fungi, mycoplasma, viruses or virus-infected cells, cancerous cells and/or protozoa (hereinafter sometimes referred to as "bacteria, etc."). means the property of causing Types of antimicrobial peptides include cell penetrating peptides (CPP), defensin antimicrobial peptides, cathelicidin antimicrobial peptides and the like, and are not particularly limited, but membrane permeable peptides are preferred. Membrane-permeable peptides include, for example, the Tet peptide. Tet peptides include, for example, Tet-213 (amino acid sequence: KRWWKWWRRC). Membrane-permeable peptides used as antibacterial peptides exhibit antibacterial properties by destroying cell membranes of bacteria and the like. By immobilizing the membrane-permeable peptide on the antifouling layer, bacteria and the like can adhere to the antifouling layer and be sterilized.

The density of the antimicrobial peptides is preferably 0.12/nm ² or more on the surface of the material. This provides the desired antimicrobial properties of the antimicrobial antifouling material.

An antimicrobial peptide is immobilized on the antifouling layer. Since the antimicrobial peptide itself does not have binding properties with the antifouling layer, the antimicrobial peptide is immobilized by, for example, chemical cross-linking with the antifouling layer treated with a linker. Antimicrobial peptides may contain cysteine residues.

[Linker (immobilized)]
The linker treatment for the antifouling layer includes, for example, a step of bonding an amino group-containing compound to a silanol group originating from the monomer unit (C) in the antifouling layer, and a step of binding a carbonyl group-containing compound to the amino group. and binding.

The amino group-containing compound is not particularly limited as long as it can be bonded to a silanol group by a condensation reaction or the like, and examples thereof include silane coupling agents containing an amino group. Specifically, ATPSi (3-aminopropyltrimethoxysilane), 3-aminopropyltriethoxysilane, N-2-(aminoethyl)-3-aminopropylmethyldimethoxysilane, N-2-(aminoethyl)- 3-aminopropyltrimethoxysilane, N-phenyl-3-aminopropyltrimethoxysilane, N,N'-bis[3-(trimethoxysilyl)propyl]ethylenediamine, and the like. The amino group-containing compound modifies the surface of the antifouling layer with amino groups. The amino group is preferably a primary amino group from the viewpoint of reactivity.

A carbonyl group-containing compound contains a carbonyl group and functions as a cross-linking agent (linker) having high reactivity with a nucleophilic amino group. Carbonyl group-containing compounds include, for example, polyfunctional aldehydes containing two or more aldehyde groups per molecule. Examples of polyfunctional aldehydes include glyoxal, malondialdehyde, succinaldehyde, glutaraldehyde, adipaldehyde, cyclohexane-1,3-dial, cyclohexane-1,4-dial, and dialdehydes derived from fatty acid dimers. , etc. As the carbonyl group-containing compound, diketones, dicarboxylic acids, imides, etc. may be used in addition to polyfunctional aldehydes. The carbonyl group-containing compound may have a thiol group-reactive chemical group in addition to the above. A thiol-reactive chemical group reacts with, for example, a reduced cysteine of an antimicrobial peptide to generate a thioether bond. Compounds having a thiol group-reactive chemical group include, for example, haloacetyl, maleimide, aziridine, acryloyl, arylating agents, vinyl sulfone, pyridyl disulfide, TNB-thiol, and the like. The carbonyl group-containing compound may further have a PEG chain introduced to improve hydrophilicity. Examples of such compounds include NHS-PEG-maleimide, which has an N-hydroxysuccinimide (NHS) group at one end of the PEG chain and a maleimide group at the other end. One carbonyl group of the carbonyl group-containing compound is chemically bonded to the amino group modified in the antifouling layer. Another carbonyl group- or thiol-reactive chemical group of the carbonyl-containing compound is chemically bonded to the terminal amino group or thiol group of the peptide chain of the antimicrobial peptide, thereby chemically cross-linking and immobilizing the antimicrobial peptide.

[material]
The antifouling layer and the antifouling antifouling material having the antifouling peptide immobilized on the antifouling layer may be either an inorganic material or an organic material, and is not particularly limited. Examples include silicon-containing polymers and glass. , polyethylene terephthalate, polyurethane, nylon, and polyurethane=nylon copolymer, and can also be applied to metals, alloys, metal oxides, ceramics, and the like. As applications of the antibacterial and antifouling material, it is preferable to apply it to members for wet areas such as houses, for example, members such as toilets, bathrooms, bathtubs, water tanks, pools, washrooms, and windows.

<Antibacterial antifouling surface treatment method>
The antibacterial antifouling treatment method according to the present embodiment includes a step of forming an antifouling layer on a material surface, and a step of immobilizing an antimicrobial peptide on the antifouling layer.

The step of forming the antifouling layer on the surface of the material includes a method of applying the surface modifier to the surface of the material by, for example, immersing the material to be treated in the surface modifier. The coating method is not limited to immersion, and known methods such as spray coating and brush coating can be used.

In general, in the case of surface modification using MPC (2-methacryloyloxyethylphosphorylcholine) polymer copolymerized with a monomer unit containing a hydrophobic moiety, the surface energy is more hydrophobic than the PC group in air (dry state). It is known that the nucleus is preferentially oriented on the surface of the material. Therefore, in order to reorient the PC groups on the surface and obtain a hydrophilic surface, a pretreatment step such as immersion in water is required before use. On the other hand, in the surface treatment using the surface modifier of the present invention, the copolymer contains three monomer units, and the hydrophilic part (monomer unit (A)) forms a block segment, so the use environment Hydrophilic groups such as PC groups are always oriented on the surface of the material regardless of the orientation. Therefore, a hydrophilic surface can be obtained without performing the above pretreatment step.

The step of immobilizing the antimicrobial peptide on the antifouling layer is performed by the above-described linker treatment method. Specific methods for linker treatment are not particularly limited, and methods known in the art can be used.

EXAMPLES The present disclosure will be described in more detail below based on examples, but the present disclosure is not limited by these examples.

<Example 1>
(Synthesis of copolymer)
As monomer units, monomer unit (A): 2-methacryloyloxyethylphosphorylcholine (MPC), monomer unit (B): 3-(methacryloyloxy)propyltris(trimethylsilyloxy)silane (MPTSSi), and monomer unit (C): 3-methacryloxypropyltrimethoxysilane (MPTMSi) was used. These were commercially available.

These monomer units were polymerized by two-step RAFT polymerization to synthesize a copolymer poly (MPC-block-(MPTSSi-random-MPTMSi)) in which MPC forms a block segment.

In the first-stage reaction, a polymer (PMPC) containing only MPC monomers was synthesized. MPC was dissolved in 1-propanol at a concentration of 0.66 M, followed by the RAFT agent 4-cyano-4-(thiobenzoylthio)pentanoic acid (CPD) and the initiator 2,2′-azobisisobutyro. After adding nitrile (AIBN) and bubbling nitrogen gas for 30 minutes, polymerization reaction was carried out in an oil bath at 65° C. for 24 hours. The concentration of the initiator was about one-fifth that of the RAFT agent. After the polymerization, the second-stage polymerization was performed without reprecipitation.

In the second-stage reaction, MPTSSi was adjusted to 0.073M and MPTMSi to 0.075M with 1-propanol, and these were added to the PMPC polymerization solution obtained in the first-stage reaction. After bubbling nitrogen gas through the solution for 30 minutes, a polymerization reaction was carried out at 65° C. for 24 hours. Then, it was reprecipitated with hexane:diethyl ether=7:3 (v/v), recovered as a solid, and stored as a coating liquid.

Regarding the obtained copolymer, the chemical structure of the obtained polymer was identified by ¹ H-NMR, and the molecular weight was measured by gel permeation chromatography (JASCO). The monomer ratio of the copolymer was 160:11:11 in MPC/MPTSSi/MPTMSi ratio.

(Immobilization of antimicrobial peptide)
A 1 vol % acetic acid aqueous solution was added to an ethanol solution (0.2 wt %) of the above copolymer at a ratio of 10/1 (v/v). A glass substrate (φ15) was washed with hexane, ethanol, and acetone for 5 minutes each, and then treated with oxygen plasma for 10 minutes. This glass substrate was immersed in the copolymer solution and allowed to stand for 30 minutes. Next, the glass substrate was vacuum-dried for 30 minutes in a desiccator. The glass substrate was kept at 70° C. in an oven and heated for 3 hours, and the surface was lightly washed with methanol. Next, a 1 vol % acetic acid aqueous solution was added to a methanol solution (0.1 wt %) of 3-aminopropyltrimethoxysilane at a ratio of 10/1 (v/v), and the glass substrate was immersed and allowed to stand for 30 minutes. , and vacuum dried for 30 minutes in a desiccator. Next, it was kept at 70° C. in an oven and heated for 3 hours, and the surface was lightly washed with methanol. Using Sylgard (registered trademark) 184, the glass substrate was fixed to a TCPS (Tissue Cultured Polystyrene) petri dish, and heated in an oven at 50° C. for 6 hours. Next, it was immersed in an aqueous NHS-PEG-maleimide solution (10 μM) at room temperature for 1 hour, and then washed with pure water. Next, it was immersed in Cys-LL-37 (PBS solution, 10 μM) as an antibacterial peptide at room temperature for 1 hour and washed with PBS to obtain an antibacterial antifouling material of Example 1.

<Example 2>
Instead of the NHS-PEG-maleimide aqueous solution, it was immersed in glutaraldehyde (PBS solution, 2.5 wt%) for 1 hour at 4 ° C. and washed with PBS, and Cys-LL-37 was replaced with Tet as the antimicrobial peptide. An antibacterial antifouling material of Example 2 was obtained in the same manner as in Example 1, except that a -213 PBS solution (100 μM) was used and immersed at 4° C. for 20 hours.

<Example 3>
In the same manner as in Example 1, except that Sylgard (registered trademark) 184 was not immobilized on a TCPS petri dish, and instead it was placed in a 24-well plate after the preparation was completed and immersed in PBS, the antibacterial antibacterial agent of Example 3 was used. Got dirty material.

<Example 4>
In the same manner as in Example 2, except that Sylgard (registered trademark) 184 was not immobilized on a TCPS petri dish, and instead it was placed in a 24-well plate after preparation and immersed in PBS, the antibacterial antibacterial agent of Example 4 was used. Got dirty material.

<Comparative Example 1>
(Synthesis of copolymer)
To 20 mL of methanol, a total amount of 0.5 M of monomer and 2.5 mM of initiator AIBN were added so that MPC/MPTMSi/MPTSSi=6/2/2 (molar ratio). After bubbling Ar gas for 15 minutes, a polymerization reaction was carried out in an oil bath at 65° C. for 24 hours. Then, it was reprecipitated with acetone:ethanol=19:1 (v/v) and recovered as a solid. This was not vacuum-dried, but was dissolved in methanol and stored as a coating liquid. The chemical structure of the obtained polymer was identified by ¹ H-NMR, and the molecular weight was measured by gel permeation chromatography (JASCO). The resulting copolymer had a monomer ratio of MPC/MPTSSi/MPTMSi of 68:17:15.

(Immobilization of antimicrobial peptide)
A 1 vol % acetic acid aqueous solution was added to a methanol solution (0.1 wt %) of the copolymer and 3-aminopropyltrimethoxysilane at a ratio of 10/1 (v/v). A glass substrate (φ15) was washed with hexane, ethanol, and acetone for 5 minutes each, and then treated with oxygen plasma for 10 minutes. This glass substrate was immersed in the copolymer solution and allowed to stand for 30 minutes. Next, the glass substrate was vacuum-dried for 30 minutes in a desiccator. The glass substrate was kept at 70° C. in an oven and heated for 3 hours, and the surface was lightly washed with methanol. The glass substrate was fixed to a TCPS petri dish using Sylgard (registered trademark) 184, and heated in an oven at 50° C. for 6 hours. Next, it was immersed in an aqueous NHS-PEG-maleimide solution (10 μM) at room temperature for 1 hour, and then washed with pure water. Next, it was immersed in Cys-LL-37 (PBS solution, 10 μM) as an antibacterial peptide at room temperature for 1 hour, washed with PBS, and an antibacterial antifouling material of Comparative Example 1 was obtained.

<Comparative Example 2>
The same copolymer as in Comparative Example 1 was used, the modification method of the copolymer was the same as in Example 1, and the antimicrobial peptide immobilization method was the same as in Example 2. An antibacterial antifouling material was obtained.

<Comparative Example 3>
Except that the antimicrobial peptide was not immobilized, and that Sylgard (registered trademark) 184 was not immobilized on a TCPS petri dish, instead, it was placed in a 24-well plate after preparation and immersed in PBS. An antibacterial antifouling material of Comparative Example 3 was obtained in the same manner as in Example 1.

[Antibacterial and detergency evaluation]
(Example 1-2, Comparative Example 1-2)
Antibacterial evaluation was performed by the following procedure. The results are shown in Figures 1-2 and 5-6. 1. The substrate fixed in the petri dish was previously immersed in PBS for 1 hour or longer. 2. S. aureus was cultured in TSB medium until OD600≈0.5. 3. After centrifuging the suspension and removing the solution, PBS was added. 4.3. operation was performed again. 5. The PBS suspension was diluted to OD600=0.02. 6. The seeds were seeded in a petri dish and allowed to stand at 37°C for 2 hours. 7. The suspension was removed and PBS was added and gently rocked. 8. It was immersed in a LIVE/DEAD BacLight staining solution (SYTO9: 4 nM, Propidium iodide: 24 nM) and allowed to stand at room temperature in the dark for 10 minutes. 9. Staining solution was removed and PBS was added. 10. Observed with a fluorescence microscope. 11. The entire petri dish was moved back and forth, left and right, and in a circular motion and shaken. 12. Observation was made with a fluorescence microscope (Olympus CKX53).

(Example 3-4, Comparative Example 3)
One glass substrate was placed for each well in the 24-well plate, except that surface washing with a pipette removed the bacterial suspension with an irregular and strong force compared to washing on the TCPS petri dish. Antibacterial evaluation was performed in the same manner as in Example 1-2 and Comparative Example 1-2. The results are shown in Figures 3-4 and 7.

In FIGS. 1 to 7, "LIVE" indicates a micrograph of live bacteria colored with a staining solution, and "DEAD" indicates a micrograph of dead bacteria colored with a staining solution. In the micrographs of FIGS. 1 to 7, white spots indicate locations where positive bacteria and dead bacteria are present, respectively. In FIGS. 1-2 and 5-6, "before shaking" indicates the micrograph of the state before the shaking operation, and "after shaking" indicates the micrograph after the shaking operation. In FIGS. 3-4 and 7, "without washing" indicates micrographs without pipette washing, and "with washing" indicates micrographs after pipette washing. In each example and comparative example, the test was performed three times, and micrographs were taken under each condition.

From the results of Examples and Comparative Examples, on the surface of the material according to Comparative Example 2 in which the monomer unit (A) does not form a block segment, removal of both live and dead bacteria was not confirmed even by shaking, and Comparative Example 1 The results confirmed the killing of the bacteria, but not the removal by shaking. On the other hand, on the surface of the material according to Example 2, it was confirmed that bacteria were killed and dead bacteria were removed. On the surface of the material according to Example 1, there were few viable bacteria and almost no dead bacteria were observed. In Examples 3 and 4 where bacteria were removed from the surface with a pipette, it was confirmed that most of the bacteria were removed from the material surface, similar to the surface of Comparative Example 3 without peptide. It is considered that no antibacterial peptide was immobilized on the surface of Comparative Example 3, and the bacteria were washed in a viable state.

Claims (6)

Having an antifouling layer and an antimicrobial peptide immobilized on the antifouling layer on the surface of the material,
The antifouling layer includes a monomer unit (A) having at least one of an amphoteric ion structure and an ether structure in a side chain; a monomer unit (B) having a hydrophobic group containing an Si atom in a side chain; An antibacterial antifouling material comprising a copolymer having a monomer unit (C) having a trialkoxysilyl group, wherein the monomer unit (A) forms a block segment. The antibacterial antifouling material according to claim 1, wherein the antibacterial peptide is a membrane-permeable peptide. 3. The antibacterial antifouling material according to claim 1, wherein said copolymer is a block polymer having a hydrophilic polymer and a silane coupling group. The antibacterial and antifouling material according to any one of claims 1 to 3, wherein the material is ceramics. The antibacterial and antifouling material according to any one of claims 1 to 4, which is used for a member for plumbing. An antibacterial antifouling surface treatment method comprising a step of forming an antifouling layer on a material surface and a step of immobilizing an antibacterial peptide on the antifouling layer,
The antifouling layer includes a monomer unit (A) having at least one of an amphoteric ion structure and an ether structure in a side chain; a monomer unit (B) having a hydrophobic group containing an Si atom in a side chain; A method for antibacterial and antifouling surface treatment, comprising a copolymer having a monomer unit (C) having a trialkoxysilyl group, wherein the monomer unit (A) forms a block segment.

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