Virus inactivator

Abstract

To provide an antibacterial or viral inactivating agent that has excellent antibacterial or viral inactivating effects against bacteria and viruses present in the environment. [Solution] (A) an antimicrobial or antiviral agent containing maleic acrylate copolymer or a salt thereof as an active ingredient, and (B) used in combination with a nonionic surfactant.

Description

[Technical Field]

[0001] The present invention relates to antibacterial or viral inactivating agents, and antibacterial or viral inactivating compositions. [Background technology]

[0002] Infections caused by bacteria and viruses, such as respiratory infections and infectious gastroenteritis, primarily spread when pathogens are brought into living spaces by infected individuals. The infection spreads directly from patients or through the environment, including people's hands, clothing, various utensils and materials, walls, air conditioners, and other equipment. Therefore, it is considered effective in preventing the spread of infection by cleaning and disinfecting hands, clothing, and various utensils and materials that may be contaminated with bacteria and viruses, eliminating viruses, and inactivating bacteria and viruses that have been dispersed into living spaces or are floating in the air as aerosols.

[0003] Traditionally, ethanol (high concentration and acidic or alkaline), chlorine-based disinfectants (such as sodium hypochlorite), iodine-based disinfectants, aldehyde-based disinfectants (such as glutaraldehyde), and peracetic acid preparations have been used for antibacterial or antiviral purposes. However, these common disinfectants are highly irritating to mucous membranes and skin, limiting their use due to safety concerns. Furthermore, they require a long time to exert antibacterial or antiviral effects, may not be sufficiently effective at room temperature, and their effectiveness may be reduced when used in combination with other bases, such as surfactants. Nonionic surfactants, in particular, tend to inhibit the effectiveness of bases that exhibit antibacterial and antiviral effects by hydrophobically acting on bacteria and viruses, as they incorporate these bases into their micelles, thus limiting the formulation design of these disinfectants.

[0004] Acrylates-maleic acid copolymer is a type of acrylic acid-based water-soluble polymer that exhibits excellent properties in terms of adsorption to inorganic salts by carboxyl groups and dispersion through ionization. Therefore, it is used in applications such as detergent builders, metal ion chelating agents, and scale inhibitors (for calcium carbonate, phosphates, etc.). On the other hand, Patent Document 1 reports that a disinfectant formulation containing an acidic polymer and an anionic surfactant, which include maleic acid as a constituent unit, possesses both antiviral and antibacterial activity. However, it is completely unknown that the combined use of maleic acrylate copolymer and nonionic surfactants has antibacterial and viral inactivation effects. [Prior art documents] [Patent Documents]

[0005] [Patent Document 1] Special Publication No. 2011-511764 [Overview of the project] [Problems that the invention aims to solve]

[0006] The present invention relates to providing an antibacterial or viral inactivating agent that has excellent antibacterial or viral inactivating effects against bacteria and viruses present in the environment. [Means for solving the problem]

[0007] The inventors have discovered that the acrylic acid-maleic acid copolymer has the effect of inactivating viruses, and that even when the polymer is used in combination with a nonionic surfactant, the effect of the polymer is not inhibited, and it exhibits excellent antibacterial and virus-inactivating properties.

[0008] In other words, the present invention relates to the following 1) to 3). 1) An antimicrobial or antiviral agent containing (A) maleic acrylate copolymer or a salt thereof as an active ingredient, and used in combination with (B) a nonionic surfactant. 2) An antimicrobial or antiviral composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant. 3) A method for antimicrobial or viral inactivation, comprising applying a composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant to an object suspected of bacterial or viral contamination. [Effects of the Invention]

[0009] The antibacterial or viral inactivating agent and antibacterial or viral inactivating composition of the present invention can kill or inactivate bacteria and viruses attached to hands, hard and soft surfaces in the living environment, and bacteria and viruses scattered in living spaces, thereby preventing or reducing the spread of infection caused by such bacteria or viruses. Furthermore, in the present invention, the effect of the acrylic acid maleic acid copolymer is not inhibited by nonionic surfactants, thus offering a high degree of freedom in formulation design. [Modes for carrying out the invention]

[0010] The present invention relates to an antimicrobial or antiviral agent used in combination with (A) maleic acrylate copolymer or a salt thereof as an active ingredient, and (B) a nonionic surfactant. Another embodiment of the present invention is an antimicrobial or antiviral composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant. (A) Acrylates-maleic acid copolymer is a copolymer containing constituent units derived from acrylic acid and constituent units derived from maleic acid. The polymerization mode of acrylics-maleic acid copolymer is not particularly limited and may be random copolymerization, block copolymerization, etc., but random copolymerization is preferred. In the acrylic acid-maleic acid copolymer of the present invention, acrylic acid and maleic acid are essential components as monomers. Maleic acid and acrylic acid may be partially or completely neutralized. Maleic anhydride may also be used as maleic acid. Furthermore, in the present invention, monomers other than acrylic acid (salt) and maleic acid (salt) may be included as monomer components, as long as they do not impair the effects of the present invention. Such monomers can be copolymerized with acrylic acid (salt) or maleic acid (salt), for example, unsaturated monocarboxylic acid monomers such as methacrylic acid and crotonic acid; neutralized products obtained by partially or completely neutralizing the above unsaturated monocarboxylic acid monomers with monovalent metals, divalent metals, ammonia, organic amines, etc.; unsaturated dicarboxylic acid monomers such as fumaric acid, itaconic acid, and citraconic acid; neutralized products obtained by partially or completely neutralizing the above unsaturated dicarboxylic acid monomers with monovalent metals, divalent metals, ammonia, organic amines, etc.; amide monomers such as (meth)acrylamide and t-butyl(meth)acrylamide; (meth) Hydrophobic monomers such as acrylic acid esters, styrene, 2-methylstyrene, and vinyl acetate; unsaturated sulfonic acid monomers such as vinyl sulfonic acid, allyl sulfonic acid, methallyl sulfonic acid, styrene sulfonic acid, 2-acrylamido-2-methylpropanesulfonic acid, 3-alyloxy-2-hydroxypropanesulfonic acid, sulfoethyl (meth)acrylate, sulfopropyl (meth)acrylate, 2-hydroxysulfopropyl (meth)acrylate, and sulfoethylmaleimide; neutralized products obtained by partially or completely neutralizing the above unsaturated sulfonic acid monomers with monovalent metals, divalent metals, ammonia, organic amines, etc.3-Methyl-2-buten-1-ol (prenol), 3-methyl-3-buten-1-ol (isoprenol), 2-methyl-3-buten-2-ol (isoprene alcohol), 2-hydroxyethyl (meth)acrylate, polyethylene glycol mono(meth)acrylate, polypropylene glycol mono(meth)acrylate, polyethylene glycol monoisoprenol ether, polypropylene glycol monoisoprenol ether, polyethylene glycol monoallyl ether, polypropylene glycol monoallyl ether, glycerol monoallyl ether Examples include, but are not limited to, hydroxyl-containing unsaturated monomers such as tel, α-hydroxyacrylic acid, N-methylol(meth)acrylamide, glycerol mono(meth)acrylate, and vinyl alcohol; cationic monomers such as dimethylaminoethyl(meth)acrylate and dimethylaminopropyl(meth)acrylamide; nitrile monomers such as (meth)acrylonitrile; and phosphorus-containing monomers such as (meth)acrylamidemethanephosphonic acid, (meth)acrylamidemethanephosphonic acid methyl ester, and 2-(meth)acrylamide-2-methylpropanephosphonic acid. These monomers may be used individually or in combination of two or more. Examples of salts of maleic acrylate copolymer include alkali metal salts, alkaline earth metal salts, ammonium salts, alkyl or alkenylammonium salts having 1 to 22 carbon atoms, alkyl or alkenyl-substituted pyridinium salts having 1 to 22 carbon atoms, alkanolammonium salts having 1 to 22 carbon atoms, and basic amino acid salts. Preferably, alkali metal salts are used, and more preferably, sodium salts.

[0011] The weight-average molecular weight of the maleic acrylate copolymer or its salt is preferably 1,000 to 1,000,000, from the viewpoint of antibacterial effect or virus inactivation effect. The weight-average molecular weight can be measured, for example, by gel permeation chromatography (GPC).

[0012] The molar ratio (acrylic acid:maleic acid) of acrylic acid-derived constituent units to maleic acid-derived constituent units constituting the acrylic acid-maleic acid copolymer or its salt is preferably 0.20:0.80 to 0.80:0.20, from the viewpoint of antibacterial effect or virus inactivation effect.

[0013] The acrylic acid maleic acid copolymer or its salts can be commercially available. Furthermore, based on previously reported information, it is possible to produce the same substance as the commercially available product, or a composition containing it, through chemical synthesis.

[0014] (B) Examples of nonionic surfactants include polyoxyethylene alkyl ethers, polyoxyethylene alkenyl ethers, polyoxyethylene polyoxypropylene glycol, polyoxyethylene polyoxypropylene alkyl ethers, polyoxyethylene polyoxypropylene alkenyl ethers, polyoxyethylene fatty acid esters, polyoxyethylene resin acid esters, polyoxyethylene glycerin fatty acid esters, polyoxyethylene sorbitan fatty acid esters, polyoxyethylene sorbitol fatty acid esters, polyoxyethylene castor oil, polyoxyethylene hydrogenated castor oil, polyoxyethylene alkylamines, polyoxyethylene alkyl ethers (C12-14 secondary alcohols) and other substances having polyoxyethylene chains; polyglycerin fatty acid esters, glycerin fatty acid esters, ethylene glycol fatty acid esters, propylene glycol fatty acid esters, butylene glycol fatty acid esters, sorbitan fatty acid esters, sucrose fatty acid esters, alkyl glucosides, etc.

[0015] The number of moles of ethylene oxide added in the polyoxyethylene chain is shown as an average value, preferably 2 or more, more preferably 3 or more, still more preferably 4 or more, and also preferably 60 or less, more preferably 50 or less, still more preferably 40 or less, even more preferably 30 or less, even more preferably 25 or less, and even more preferably 20 or less. Further, the number of carbon atoms in the fatty acid moiety, alkyl moiety, and alkenyl moiety of the nonionic surfactant is preferably 6 or more, more preferably 8 or more, still more preferably 10 or more, and also preferably 24 or less, more preferably 22 or less, still more preferably 20 or less. Note that "having a polyoxyethylene chain" means a structure in which 1 mole or more of ethylene oxide is added to the molecule of the nonionic surfactant.

[0016] The nonionic surfactant may be one kind or a mixture of two or more kinds. Among them, from the viewpoint of antibacterial effect or virus inactivating effect, polyoxyethylene alkyl ether or alkyl glycoside is preferable. Commercially available nonionic surfactants can be used. Also, based on previously reported information, it is possible to produce a composition identical to or containing a commercially available product by chemical synthesis.

[0017] As shown in the examples described later, the combination of (A) acrylic acid - maleic acid copolymer or its salt and (B) nonionic surfactant exhibits excellent antibacterial and virus inactivating effects. While the acrylic acid - maleic acid copolymer or its salt has an effect of inactivating viruses, etc., the nonionic surfactant generally does not have antibacterial activity and virus inactivating activity by itself, and it is known that it takes in a base showing antibacterial and virus inactivating effects into the micelles and inhibits the effects. In contrast, even when the acrylic acid - maleic acid copolymer or its salt and the nonionic surfactant are used in combination, the effect of the acrylic acid - maleic acid copolymer or its salt is not inhibited by the nonionic surfactant, but rather the effect is improved by the combined use with the nonionic surfactant. Therefore, (A) an acrylic acid maleic acid copolymer or a salt thereof can be an antibacterial or virus inactivator used in combination with (B) a nonionic surfactant, and can also be used for producing an antibacterial or virus inactivator. Further, (A) an acrylic acid maleic acid copolymer or a salt thereof can be used in combination with (B) a nonionic surfactant to inactivate bacteria or viruses. Furthermore, the combination of (A) an acrylic acid maleic acid copolymer or a salt thereof and (B) a nonionic surfactant can be an antibacterial or virus inactivating composition exhibiting an antibacterial effect or a virus inactivating effect, and can also be used for producing an antibacterial or virus inactivating composition. Also, the combination of (A) an acrylic acid maleic acid copolymer or a salt thereof and (B) a nonionic surfactant can be used for antibacterial or virus inactivation. Applying a composition containing (A) an acrylic acid maleic acid copolymer or a salt thereof and (B) a nonionic surfactant to a subject suspected of being contaminated with bacteria or viruses enables antibacterial or virus inactivation.

[0018] Although various species of microorganisms can be the targets of the antibacterial or virus inactivating composition in the present invention, specifically, the following gram-positive bacteria, gram-negative bacteria, or viruses can be mentioned. Examples of Gram-positive bacteria include: Bacillus species such as Bacillus subtilis, Bacillus anthracis, and Bacillus cereus; Listeria species such as Listeria monocytogenes, Listeria ivanovii, and Listeria seeligeri; Alicyclobacillus species such as A. acidoterrestris (formerly B. acidoterrestris); Staphylococcus species such as S. aureus; and Streptococcus species such as S. pyogenes. Examples include Clostridium bacteria such as C. botulinum (C. botulinum), C. perfringens (C. perfringens), and C. sporogens; Clostridioides bacteria such as C. difficile; Leuconostoc bacteria such as L. mesenteroides; Desulfotomaculum bacteria such as D. nigrificans; Enterococcus bacteria such as E. faecalis, E. faecium, E. gallinarum, and E. casseriflavus; and Streptococcus bacteria such as Streptococcus pneumoniae. Examples of Gram-negative bacteria include Shigella bacteria such as S. dysenteriae (Shigella subgroup A), S. flexneri (Shigella subgroup B), S. boydii (Shigella subgroup C), and S. sonnei (Shigella subgroup D); Brucella bacteria; Escherichia coli such as E. coli O157; S. typhi (S. typhi), S. paratyphi A (S. paratyphi A), and S. paratyphi Examples include Salmonella bacteria such as Salmonella paratyphi B, Salmonella typhimurium, and Salmonella enteritidis; Vibrio bacteria such as Vibrio cholerae and Vibrio parahaemolyticus; Pseudomonas bacteria such as Pseudomonas aeruginosa; Acinetobacter bacteria such as A. baumannii; Klebsiella bacteria such as Klebsiella pneumoniae; Stenotrophomonas bacteria such as S. maltophilia; and Enterobacter bacteria such as E. cloacae.

[0019] The term "virus" includes all types of viruses, regardless of the type of nucleic acid (RNA, DNA) and whether or not they have an envelope. Enveloped viruses include those with RNA as their nucleic acid, such as influenza virus; coronavirus (including SARS coronavirus and SARS coronavirus-2); respiratory syncytial virus (RSV); mumps virus; lassa virus; dengue virus; rubella virus; and human immunodeficiency virus; as well as those with DNA as their nucleic acid, such as human herpesvirus; vaccinia virus; and hepatitis B virus. Furthermore, examples of viruses that do not have an envelope include those with RNA as their nucleic acid, such as norovirus, poliovirus, echovirus, hepatitis A virus, hepatitis E virus, rhinovirus, astrovirus, rotavirus, coxsackievirus, enterovirus, and sapovirus, and those with DNA as their nucleic acid, such as adenovirus, B19 virus, papovavirus, and human papillomavirus. SARS-CoV-2 (Severe acute respiratory syndrome coronavirus 2) is a SARS-related coronavirus that causes acute respiratory illness (COVID-19).

[0020] Of these, Staphylococcus bacteria, influenza virus, coronavirus, norovirus, coxsackievirus, and adenovirus are preferred, with norovirus being more preferred. Norovirus includes HNV (human norovirus), FCV (feline calicivirus), MNV (mouse norovirus), and others. Noroviruses are divided into seven gene groups (genogroups, GI to GVII) based on the homology of their genome base sequences. The HNVs that infect humans are GI9 types (GI.1, GI.2, GI.3, GI.4, GI.5, GI.6, GI.7, GI.8, GI.9), GII19 types (GII.1, GII.2, GII.3, GII.4, GII.5, GII.6, GII.7, GII.8, GII.9, GII.10, GII.12, GII.13, GII.14, GII.15, GII.16, GII.17, GII.20, GII.21, GII.22), and GIV1 type (GIV.1). However, the HNV in this invention may be any of these genotypes.

[0021] In this invention, "antibacterial" is a term that includes all of the concepts of "sterilization" and "disinfection," which kill microorganisms, and "bacteriostatic" and "antimicrobial" actions, which suppress the occurrence, growth, and proliferation of microorganisms. Furthermore, "virus inactivation" refers to the process of reducing or eliminating the activity of a virus, thereby eliminating its ability to infect host cells. The virus inactivation effect can be confirmed, for example, by bringing the test product into contact with the virus, infecting host cells with the virus, and measuring the viral infectivity titer. Here, the host cell can be any cell in which the target virus can proliferate. For example, for feline calicivirus, cat kidney-derived cell lines (CRFK) can be used; for adenovirus, human laryngeal cancer-derived cells (HEp-2) can be used; for influenza virus, canine kidney cells (MDCK), African green monkey kidney epithelial cells (Vero), or duck embryonic stem cell-derived cell lines (EB66) can be used; for human coronavirus, human ileocecal adenocarcinoma cells (HCT-8), African green monkey kidney epithelial cells (VeroE6), or human liver cancer-derived cell lines (Huh7) can be used; and for coxsackievirus, African green monkey kidney epithelial cells (Vero) can be used. Furthermore, the human norovirus inactivation effect can be evaluated by a system in which human intestinal organoids are cultured in three dimensions on an extracellular matrix, and then differentiated to obtain intestinal epithelial cells, which are then infected with HNV (see Examples).

[0022] The virus inactivating agents and antibacterial or virus inactivating compositions (hereinafter also referred to as "virus inactivating agents, etc.") in the present invention include those used in a liquid or gaseous state, but from the viewpoint of ease of application to objects where bacterial or viral contamination is a concern, they are preferably used in a liquid state. Virus inactivators used in the liquid phase may essentially consist of (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant, but may also contain, in addition to (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant, a base, antimicrobial substances such as hypochlorous acid, hydrogen peroxide, and silver ion compounds, cationic antimicrobial agents (such as benzethonium chloride), bactericides (such as triclosan and isopropylmethylphenol), and surfactants other than component (B). Furthermore, they are prepared by appropriately blending additives such as chelating agents, humectants, lubricants, builders, buffers, abrasives, electrolytes, bleaches, fragrances, dyes, foaming control agents, corrosion inhibitors, essential oils, thickeners, pigments, gloss enhancers, enzymes, detergents, solvents, dispersants, polymers, silicones, and hydrophobic substances. Such virus inactivators may take the form of liquid, emulsion, cream, lotion, paste, gel, sheet (with base), oil, etc., but are not limited to these forms.

[0023] The virus inactivator can be used as is, or it can be appropriately incorporated into various cleaning agents (laundry detergents, household cleaning agents, dishwashing detergents, hair washes, hand washes, body washes, etc.), disinfectants, sanitary product compositions, etc. In particular, from the viewpoint of more easily enjoying the effects of the present invention, it is preferable to use it in cleaning agents that frequently use nonionic surfactants.

[0024] Examples of the above-mentioned sanitary product compositions include lotions, creams, shampoos, hair conditioners, hand soaps, body washes, facial cleansers, bath additives, foams, antiperspirants, deodorants, underarm odor preventatives, and oral hygiene products (mouthwash, toothpaste, mouth fresheners, gargles, etc.). These compositions can be prepared by conventional methods by appropriately combining carriers that are acceptable as cosmetics, etc. (for example, diluents, dispersants, buffers, pH adjusters, emulsifiers, surfactants, preservatives, stabilizers, antioxidants, colorants, humectants, thickeners, disinfectants, fragrances, etc.).

[0025] Virus inactivators used in the gas phase may essentially consist of (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant, but can also be prepared by blending (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant with a base and various additives (polyols (dipropylene glycol, propylene glycol, etc.), surfactants other than component (B), UV absorbers, antioxidants, preservatives, deodorants, natural extracts, silicones, thickeners, dyes, pigments, colorants, oils, fragrances, etc.). Such virus inactivators may take the form of a liquid or gel, but a liquid form is preferred. Furthermore, when preparing a gel-like formulation, it can be prepared by appropriately adding a natural or synthetic gelling agent, such as a water-soluble gelling agent like carrageenan or gellan gum, or an oil-soluble gelling agent like metal soap or aluminum octylate, according to conventionally known methods.

[0026] In this specification, examples of bases include conventionally known substances, whether oily or aqueous, such as water, methanol, ethanol, propanol, isopropanol, butanol, isobutanol, 3-methyl-3-methoxybutanol, propylene glycol, triethylene glycol, dimethyl ether, liquid propane, petrolatum, lanolin, castor oil, and paraffinic hydrocarbons (e.g., liquid paraffin). These bases can be used individually or in combination of two or more.

[0027] Furthermore, surfactants other than component (B) include at least one selected from anionic surfactants, cationic surfactants, and amphoteric surfactants. (A) When maleic acrylate copolymer or a salt thereof is used in combination with these surfactants, the virus inactivation effect is further enhanced. Preferred anionic surfactants include sulfuric acid-based, sulfonic acid-based, and carboxylic acid-based surfactants. Examples include alkyl sulfates, alkylbenzene sulfonates, polyoxyalkylene alkyl ether sulfates, polyoxyalkylene alkenyl ether sulfates and polyoxyalkylene alkylaryl ether sulfates, fatty acid salts, pyrophosphates, lauryl phosphate, polycarboxylic acid-type polymers, polyoxyethylene alkylene alkyl acetates, aromatic sulfonate formalin condensates, polyoxyethylene distyrenated ether sulfates, alkyldiphenyl ether disulfonates, dialkyl sulfosuccinates, and alkylnaphthalene sulfonates. Examples of salts include alkali metal salts, alkaline earth metal salts, ammonium salts, organic ammonium salts, and basic amino acid salts.

[0028] As cationic surfactants, quaternary ammonium type surfactants are preferred, and examples include alkylbenzyldimethylammonium salts, dialkyldimethylammonium salts, alkyltrimethylammonium salts, and triethanolamine diester quaternary salts. From the viewpoint of surfactant activity, the number of carbon atoms in the alkyl portion is preferably 8 or more, more preferably 10 or more, and also preferably 22 or less, more preferably 20 or less, and even more preferably 18 or less.

[0029] As amphoteric surfactants, betaine-type and amine oxide-type surfactants are preferred. Examples include carbobetaines such as alkylcarbobetaine and alkyl(amidopropyl)carbobetaine, sulfobetaines such as alkylsulfobetaine, alkyl(amidopropyl)sulfobetaine and alkylhydroxysulfobetaine, and alkyldimethylamine oxide. From the viewpoint of surfactant activity, the number of carbon atoms in the alkyl portion is preferably 6 or more, more preferably 8 or more, even more preferably 10 or more, and also preferably 22 or less, more preferably 20 or less, and even more preferably 18 or less.

[0030] The content of the active ingredient in the antibacterial or virus-inactivating composition of the present invention can be set as appropriate. For example, the content of (A) maleic acrylate copolymer or its salt relative to the total amount of the composition is preferably 0.001% by mass or more, more preferably 0.01% by mass or more, and even more preferably 0.5% by mass or more. There is no particular upper limit, for example, preferably 99.999% by mass or less, and even more preferably 10% by mass or less. Furthermore, the content of (B) nonionic surfactant relative to the total amount of the composition is preferably 0.001% by mass or more.

[0031] In the present invention, the ratio of the combination of (A) maleic acrylate copolymer or a salt thereof and (B) nonionic surfactant is preferably 1000 parts by mass or less, more preferably 100 parts by mass or less, and even more preferably 10 parts by mass or less of (B) nonionic surfactant per 1 part by mass of (A) maleic acrylate copolymer or a salt thereof, from the viewpoint of antibacterial effect or virus inactivation effect.

[0032] The virus inactivators in this invention are active under acidic, neutral, and alkaline conditions, but from the viewpoint of safety and virus inactivation effect, the pH (at 20°C) is preferably 3 to 11, more preferably 3 to 5 or 10 to 11. The pH shall be measured using a pH meter after adjusting the temperature to 20°C.

[0033] The virus inactivating agent, etc., of the present invention makes it possible to perform antimicrobial action or virus inactivation on objects where bacterial or viral contamination is a concern. Examples of objects where bacterial or viral contamination is a concern include the skin or mucous membranes of animals to which bacteria or viruses adhere, hard or soft surfaces of inanimate objects, objects such as waste, and living spaces where bacteria or viruses are scattered or airborne. Examples of surfaces of inanimate objects include hard surfaces in homes and business facilities such as counters, sinks, restrooms, toilets, bathtubs, showers, floors, windows, doorknobs, walls, drains, pipes, and garbage collection areas; hard surfaces of special vehicles such as garbage trucks and sanitation vehicles such as garbage inlets, work surfaces, and switch surfaces; hard surfaces of transport vehicles such as railway cars and aircraft bodies such as floors, handrails, and door surfaces; hard surfaces of various appliances, tools, and miscellaneous goods such as kitchenware, furniture, telephones, and toys; and soft surfaces of textile products (carpets, area rugs, curtains, seats, fabric furniture, clothing, masks, etc.). Examples of waste include general waste (household waste such as food scraps, tissues, and masks) and industrial waste (sludge, human waste, medical waste, etc.). If the waste is contained within a bag, the bag surface may be targeted. Other examples of living spaces include the entrance hall, dining kitchen, bedroom, children's room, bathroom, toilet, etc. within ordinary homes; the interiors of facilities such as shops, restaurants, inns, hospitals, workshops, factories, garbage collection points, quarantine stations, livestock farms, and markets; the interiors of vehicles such as cars, trains, and airplanes; and semi-enclosed spaces (lockers, storage rooms, closets, etc.; storage boxes (for toys, karaoke microphones, dishes, condiments, writing instruments, stationery)).

[0034] In the present invention, the manner in which a composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant is applied to an object where bacterial or viral contamination is a concern is not particularly limited, and (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant may be brought into contact with or reacted with bacteria or viruses in a liquid phase or a gas phase. The method for contacting bacteria or viruses with (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant in a liquid phase may be any of the following: applying the composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant directly to the object to be treated; diffusing and sprinkling the composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant onto the object to be treated; or wiping the surface of the object with a sheet, gauze, towel, wet wipe, tissue, or the like impregnated with the composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant. Another method involves filling a composition containing (A) acrylic acid maleic acid copolymer or a salt thereof and (B) a nonionic surfactant into a known spray container, such as a trigger spray container (direct pressure or stored pressure type), a dispenser-type pump spray container, or an aerosol spray container equipped with a pressure-resistant vessel, and spraying it onto the target object while appropriately adjusting the spray volume. Another method involves filling an atomizing device such as a pressurized air atomizing spray device, a nebulizer, or a diffuser, or a diffusion device such as a washer nozzle or misting machine, and spraying it into a space where bacteria or viruses are present. By spraying it in a mist form into a space where bacteria or viruses are present, the volatilization rate can be increased, and an antibacterial effect or virus inactivation effect can be rapidly exerted.

[0035] A method for contacting or reacting bacteria or viruses with (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant in the gas phase includes, for example, a form in which (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant are allowed to volatilize naturally or by forced volatilization. This can inactivate bacteria or viruses present in living spaces, and allows for easy removal of bacteria or viruses (deviral removal) from the space. When used for natural volatilization, conventionally known methods can be applied, such as impregnating a core rod, filter paper, etc., with a composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant and allowing it to volatilize, or using a permeable membrane for volatilization. Alternatively, a composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant can be kneaded into a resin for use. Examples of resins that can be kneaded include natural, petroleum-based, and synthetic waxes, rosin resins, ethylene-vinyl acetate copolymers, ethylene-vinyl alcohol copolymers, polyesters, polyolefins, and acrylic resins. The above kneaded material can be used as is, or it can be supported on a porous carrier, formed into a sheet, or used as a laminate of the sheet. Examples of porous carriers include natural polymers such as cellulose and chitosan, the above synthetic resins, and inorganic porous materials such as calcium silicate, all in any shape such as granules or sheets. The above-mentioned mixtures and laminates can be used, for example, by installing them in air conditioning equipment, toilets, bathrooms, living rooms, hospital rooms, waiting rooms, dustbins, etc., while gradually volatilizing (A) maleic acrylate copolymer or its salt and (B) nonionic surfactant. Alternatively, a composition containing (A) maleic acrylate copolymer or its salt and (B) nonionic surfactant can be supported on products made of paper, nonwoven fabric, etc. (such as air purifier filters) and used. When (A) an acrylate maleic acid copolymer or a salt thereof and (B) a nonionic surfactant are used by forced volatilization, such means include, for example, a method of volatilization using a fan, a heating volatilization method using a heater, or a method of volatilization using ultrasound. When performing airborne virus removal treatment, the amount of composition containing (A) maleic acrylate copolymer or its salt and (B) nonionic surfactant used can be appropriately adjusted depending on the treatment method, the ambient environment such as temperature and humidity, the vapor pressure of the compound, etc. It is also possible to use a concentration above the saturation concentration of the compound in the space. For example, the concentration of (A) maleic acrylate copolymer or its salt or (B) nonionic surfactant in the target space is preferably 0.1% by mass or more, preferably 1% by mass or more, more preferably 5% by mass or more, even more preferably 10% by mass or more, preferably 100% by mass or less, preferably 50% by mass or less, more preferably 25% by mass or less, and even more preferably 20% by mass or less, when it volatilizes. The concentration of (A) acrylic acid maleic acid copolymer or its salt, or (B) nonionic surfactant in the target space can be detected by measuring the concentration of the compound in a gas sample taken from the space. Methods for determining the concentration include using a volatile organic compound (VOC) concentration meter (VOC meter, odor sensor, etc.), or using gas chromatography or gas chromatography / mass spectrometry in combination with a gas collection tube, etc.

[0036] With regard to the embodiments described above, the present invention further discloses the following aspects.

[0037] <1> (A) an antimicrobial or antiviral agent containing maleic acrylate copolymer or a salt thereof as an active ingredient, and (B) used in combination with a nonionic surfactant. <2> An antimicrobial or virus-inactivating composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant. <3> A method for antimicrobial or viral inactivation, comprising applying a composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant to an object suspected of bacterial or viral contamination.

[0038] <4> The molar ratio (acrylic acid:maleic acid) of acrylic acid-derived constituent units to maleic acid-derived constituent units constituting the acrylic acid-maleic acid copolymer or its salt is preferably 0.20:0.80 to 0.80:0.20. <1> ~ <3> An antibacterial or viral inactivating agent, antibacterial or viral inactivating composition, or antibacterial or viral inactivating method as described in any of the above. <5> The nonionic surfactant is preferably at least one selected from polyoxyethylene alkyl ethers, polyoxyethylene alkenyl ethers, polyoxyethylene polyoxypropylene glycol, polyoxyethylene polyoxypropylene alkyl ethers, polyoxyethylene polyoxypropylene alkenyl ethers, polyoxyethylene fatty acid esters, polyoxyethylene resin acid esters, polyoxyethylene glycerin fatty acid esters, polyoxyethylene sorbitan fatty acid esters, polyoxyethylene sorbitol fatty acid esters, polyoxyethylene castor oil, polyoxyethylene hydrogenated castor oil, polyoxyethylene alkylamines, polyoxyethylene alkyl ethers (C12-14 secondary alcohols), polyglycerin fatty acid esters, glycerin fatty acid esters, ethylene glycol fatty acid esters, propylene glycol fatty acid esters, butylene glycol fatty acid esters, sorbitan fatty acid esters, sucrose fatty acid esters, and alkyl glucosides, and more preferably at least one selected from polyoxyethylene alkyl ethers and alkyl glycosides. <1> ~ <4> An antibacterial or viral inactivating agent, antibacterial or viral inactivating composition, or antibacterial or viral inactivating method as described in any of the above. <6> The bacteria are preferably Staphylococcus bacteria, and the virus is preferably an enveloped RNA virus, an unenveloped RNA virus, or an unenveloped DNA virus, more preferably an influenza virus, coronavirus, norovirus, coxsackievirus, or adenovirus, and even more preferably norovirus. <1> ~ <5> An antibacterial or viral inactivating agent, antibacterial or viral inactivating composition, or antibacterial or viral inactivating method as described in any of the above. <7> The pH of the composition is preferably 3 to 11, more preferably 3 to 5 or 10 to 11. <2> ~ <6> An antibacterial or viral inactivation composition or antibacterial or viral inactivation method as described in any of the above. [Examples]

[0039] Test Example 1 <Preparation of test solution> Preparations A to D, as described in Table 2, were prepared using acrylic acid / maleic acid copolymers (product names: Polymer A, Polymer B, Polymer C, see Table 1), ammonium lauryl sulfate (Emal AD-25, manufactured by Kao Corporation), sodium polyoxyethylene (EO=2) alkyl (R=10-16) ether sulfate (manufactured by Kao Corporation), polyoxyethylene (EO=10) alkyl (R=12-14) ether (product name: PANNOX710, manufactured by Pan Asia Chemical Corporation), benzoic acid, citric acid, and sterile deionized water. Preparations A to D were diluted with sterile ultrapure water to prepare the test solutions described in Table 3. After mixing the test solution with the virus solution or the test solution with the bacterial solution, the pH of each test solution was finely adjusted with monoethanolamine or 2M hydrochloric acid solution so that the pH was as described in Table 3. Sterile ultrapure water was used as a control. The units in the tables are "mass%". Table 3 shows the pH after mixing the test solution and virus solution, the pH after mixing the test solution and bacterial solution, and the log reduction [log(control pfu / mL after culture) - log(test solution pfu / mL after culture) or log(control cfu / mL after culture) - log(test solution cfu / mL after culture)]. The virus inactivation and bactericidal effects of the test solution were evaluated using the method described below.

[0040] [Table 1]

[0041] [Table 2]

[0042] <Virus inactivation evaluation method for Examples 1-5> (1) Human adenovirus inactivation evaluation method 3.75 mL of the test solution, prepared to the concentration described in Example 2, and 0.1 mL of adenovirus solution (Human adenovirus 5; Strain: Adenoid 75 ATCC VR-5) adjusted to a log pfu / mL of 7.5 were thoroughly mixed and reacted at 30°C for the time described in Table 3. 100 μL of the reaction mixture was diluted 10-fold by adding 900 μL of quenching solution (SCDLP solution) to stop the reaction. As a control, the same procedure was performed using sterile ultrapure water instead of the test solution. The solution after reaction cessation was 10 0 First, a 10-fold dilution series was prepared using EMEM (Eagle's Minimum Essential Medium). These dilutions were then used to infect HEp-2 cells (human laryngeal cancer-derived cells, ATCC CCL-23), and the cells were cultured at 37°C until plaque formation was observed. After that, the number of plaques in each dilution was counted.

[0043] (2)Human coxackievirus inactivation evaluation method 3.75 mL of the test solution, prepared to the concentration described in Example 3, and 0.1 mL of coxsackievirus solution (Human coxsackievirus B6 ATCC VR-155) adjusted to a log pfu / mL of 7.9 were thoroughly mixed and reacted at 30°C for the time described in Table 3. 100 μL of the reaction mixture was diluted 10-fold by adding 900 μL of quenching solution (SCDLP solution) to stop the reaction. As a control, the same procedure was performed using sterile ultrapure water instead of the test solution. The solution after reaction cessation was 10 0 First, a 10-fold dilution series was prepared using EMEM (Eagle's Minimum Essential Medium). These dilutions were then used to infect Vero cells (ATCC CCL-81), and the cells were cultured at 37°C until plaque formation was observed. After that, the number of plaques in each dilution was counted.

[0044] (3) Influenza A virus (H3N2) inactivation evaluation method 0.8 mL of influenza A virus solution (H3N2,A / Hong Kong / 8 / 68;TC adapted ATCC VR-1679) adjusted to a log pfu / mL of 7.6 was thoroughly mixed with 30 mL of test solution prepared to the concentration described in Example 1, and the mixture was reacted at 30°C for the time described in Table 3. Similarly, 0.5 mL of influenza A virus solution adjusted to a log pfu / mL of 7.8 was thoroughly mixed with 30 mL of test solution prepared to the concentration described in Example 4, and the mixture was reacted at 30°C for the time described in Table 3. After the reaction, 900 μL of quenching solution (SCDLP solution) was added to 100 μL of each mixture to dilute it 10-fold and stop the reaction. As a control, the same procedure was performed using sterile ultrapure water instead of the test solution. The solution after reaction cessation was 10 0 First, a 10-fold dilution series was prepared using EMEM (Eagle's Minimum Essential Medium). These dilutions were then used to infect MDCK cells (canine kidney-derived cells, ATCC CCL-34), and the cells were cultured at 34°C until plaque formation was observed. After that, the number of plaques in each dilution was counted.

[0045] (4) SARS-CoV-2 inactivation evaluation method 0.1 mL of the SARS-CoV-2 mutant strain "(Omicron strain, BA.5); hCoV-19 / Japan / TY41-702 / 2022 NIID isolate; JPN / TY / WK-521 (provided by the National Institute of Infectious Diseases)" with a log pfu / mL adjusted to 8.2 was thoroughly mixed with 0.9 mL of the test solution prepared to the concentration described in Example 5, and the mixture was reacted at 25°C for the time described in Table 3. 100 μL of the reaction mixture was diluted 10-fold with 900 μL of quench solution (SCDLP solution diluted 10-fold with 2% FBS-containing DMEM *FBS: fetal bovine serum, DMEM: Dulbecco's modified Eagle medium) to stop the reaction. As a control, the same procedure was performed using sterile ultrapure water instead of the test solution. The solution after reaction cessation was 10 0 A 10-fold dilution series was prepared using 2% FBS-containing DMEM. These dilutions were then used to infect VeroE6 / TMPRSS2 (JCRB1819) cells, respectively. After incubation at 37°C until plaque formation was confirmed, the number of plaques in each dilution was counted.

[0046] <Sterilization test of Examples 6-7 / Comparative Example 1> A solution of Staphylococcus aureus (ATCC6538) stored at low temperature was collected using a platinum loop, inoculated onto SCDLP agar, and incubated at 37°C for 18 hours. After incubation, Staphylococcus aureus colonies were taken from the SCDLP agar and added to a 0.85% sodium chloride aqueous solution. The turbidity (OD600) was 0.5 (10 8 A bacterial suspension (equivalent to cfu / mL) was prepared. 19.8 mL of sterile ultrapure water, or 19.8 mL of each test solution prepared to the concentrations described in Examples 6-7 and Comparative Example 1, was thoroughly mixed with 0.2 mL of the prepared bacterial suspension and reacted at room temperature for the time indicated in Table 3. After the reaction, 0.1 mL each of the mixture and dilutions obtained by diluting the mixture 10-fold, 100-fold, 1000-fold, and 10000-fold (all diluted with 0.85% sodium chloride solution) were inoculated onto SCDLP agar plates. After incubation at 37°C for 18 hours, the colonies formed on the medium were counted and the log reduction was calculated.

[0047] [Table 3]

[0048] Test Example 2 <Preparation of test solution> Acrylic acid / maleic acid copolymer (product name: Polymer A), polyoxyethylene (EO=10) alkyl (R=12-14) ether (product name: PANNOX710, manufactured by Pan Asia Chemical Corporation), ammonium lauryl sulfate (manufactured by Kao Corporation), polyoxyethylene (EO=2) alkyl (R=10-16) ether sodium sulfate (manufactured by Kao Corporation), benzoic acid, and citric acid were suspended in sterile deionized water to the concentrations shown in Tables 4-5. 2M hydrochloric acid solution and monoethanolamine were used for pH adjustment. Sterile deionized water adjusted to the pH shown in the table using the aforementioned acid / alkali agents was used as a control. These controls did not contain polymers or activators. Units in the table are "mass%". The table shows the pH, feline calicivirus amount (pfu / mL), and HNV genome copy number [copies / μL] after mixing the test solution / virus solution. The virus inactivation effect of the test solution was evaluated using the method described below.

[0049] <Virus Inactivation Evaluation Method> (1) FCV inactivation evaluation method The inactivation evaluation of the virus was performed based on the "FY2015 Survey Report on Norovirus Inactivation Conditions" (National Institute of Health Sciences) and ASTM-E1052, which are generally known as standard methods for evaluating viral inactivation. In a 1.5 mL sample tube, 90 μL of each test solution shown in Table 4 and 10 μL of feline calicivirus solution (F-9 strain, ATCC VR-782) adjusted to a log pfu / mL of approximately 6.3 were added and thoroughly mixed. After reacting at 30°C for the time indicated in Table 4, the reaction was stopped by diluting 10-fold with 900 μL of quench solution (SCDLP solution). The solution after reaction stoppage was serially diluted 10-fold, 100-fold, and 1000-fold in DMEM medium. 0.5 mL of each of these dilutions was then used to infect CRFK cells (feline kidney-derived cell line) (ATCC CCL-94) washed with PBS (phosphate-buffered saline), and after 1 hour, the medium was replaced with methylcellulose-containing medium. After culturing at 37°C for two days, the cells were stained with crystal violet and the number of plaques was counted.

[0050] (2) HuNoV inactivation evaluation method (i) Culturing of human intestinal epithelial organoids (hIEOs) hIEOs were embedded in Matrigel (Corning) on ​​24-well plates and then cultured in three dimensions under conditions of 37°C and 5% CO2. IntestiCult Organoid Growth Medium (Human) (STEMCELL Technologies) was used as the culture medium. The entire volume of medium (500 μL / 2 days) was changed every 2-3 days, and subculturing was performed every 5-7 days. After 3D culture, the hIEOs were suspended in TrypLE Express (Thermo Fisher Scientific) along with the Matrigel. The hIEOs were dispersed by enzymatic treatment at 37°C for 10 minutes, and to stop the reaction, a basal medium containing Advanced DMEM / F-12 (Gibco) supplemented with 100 U / mL Penicillin-Streptomycin (Thermo Fisher Scientific), 10 mM HEPES (Thermo Fisher Scientific), and 1×GlutaMAX (Thermo Fisher Scientific) was added. After settling the cells by centrifugation (3 minutes, 300×g, 4°C), the above medium was removed. Cells were suspended in differentiation medium supplemented with 1×B27 supplement (Thermo Fisher Scientific), 10nM gastrin I (Sigma-Aldrich), 1mM N-acetylcysteine ​​(Fujifilm Wako), 50ng / mL mouse EGF recombinant protein (Thermo Fisher Scientific), 100ng / mL mouse Noggin (Peprotech), 500μg / mL human R-spondin-1 (R&D Systems), 500nM A83-01 (Tocris), and 10μM Y-27632 (Fujifilm Wako) in basal medium. Furthermore, for monolayer culture, 96-well flat-bottom plates coated with Cell Matrix® type IC (Nitta Gelatin) were prepared. To prepare the plates, 100 μL / well of Cell Matrix® type IC (Nitta Gelatin), diluted 10-fold with 0.1 M acetic acid (Nacalai tesque), was added, followed by incubation at 37°C for 90 minutes. After that, the solution was removed and the plates were washed three times with 200 μL of D-PBS(-) (Nacalai tesque). 5 × 10⁶ cells were then added to the 96-well flat-bottom plates prepared as described above. 4 Cells were seeded at a concentration of 100 μL / well. Differentiation induction was carried out for a total of 6 days, with the entire differentiation medium being replaced every 2 days, to obtain monolayered hIEOs.

[0051] (ii) Preparation of a 10% emulsion of feces containing HNV (HuNoV) A 10% fecal emulsion was prepared from the feces of patients with HNV type GII.4. One tablet of the protease inhibitor cOmplete protease inhibitor cocktail tablets (Sigma-Aldrich, 11697498001) was suspended in 50 mL of D-PBS(-). 1 g of feces was suspended in 10 mL of cOmplete-containing D-PBS(-) and thoroughly mixed using a test tube mixer. After standing at 4°C for 20 minutes, the mixture was centrifuged at 2,000 × g for 10 minutes at 4°C. The supernatant was collected in a new tube and stored at -80°C until use in infection experiments.

[0052] (iii) Inactivation treatment of HNV and infection of differentiated hIEOs A 10% fecal emulsion containing HNV was diluted 10-fold with differentiation medium and filtered using a 1 mL syringe and a Millex HV Filter unit (Millipore, SLHVR04NL). 5 μL of the filtered fecal solution (2.8 × 10⁶) was collected in a PA microcentrifuge tube (Beckman coulter). 6A 45 μL test solution containing HNV genome copy equivalent and polymer at the specified concentrations shown in Table 5 was mixed and reacted at the specified temperature and time shown in Table 5. Next, 0.95 mL of Fetal Bovine Serum (Capricorn Scientific) solution was added to this drug-treated fecal solution. The centrifuge tube was set on a fixed-angle rotor TLA-55 (Beckman coulter) and ultracentrifuged for 1.5 hours at Rmax with a centrifugal force of 186047 × g (equivalent to 55,000 rpm) using an Optima MAX-TL (Beckman coulter), and the supernatant was removed. The pellet was suspended in 100 μL of differentiation medium to prepare the infection solution for hIEOs. After removing the existing medium from the wells (96-well plate), the infection solution prepared in the manner described above was applied to the hIEOs after 6 days of differentiation induction. Incubation was performed at 37°C for 1 hour. After washing three times with 300 μL of basal medium, 250 μL of differentiation medium was added, and the cells were cultured at 37°C under 5% CO2 conditions until sampling. 10 μL of supernatant was collected immediately after the start of culture and again after 6 days of culture. The collected supernatant was centrifuged at 15,000 rpm for 5 minutes to remove cell debris, and then frozen and stored at -80°C until HuNoV genome RNA quantification was performed.

[0053] (iv) RT-qPCR The number of HNV genome copies in the recovered supernatant was quantified using the Norovirus Detection Kit G1 / G2 (Toyobo, FIK-273). The procedure followed the protocol. PCR amplification and data measurement were performed using LightCycler480II (Roche).

[0054] [Table 4]

[0055] [Table 5]

[0056] Test Example 3 <Preparation of test solution> Acrylic acid / maleic acid copolymers (product names: Polymer A, Polymer B), polyoxyethylene alkyl ethers (product name: Fatty Alcohol 10 Ethoxylate, manufactured by PT. POLYCHEM INDONESIA Tbk), polyoxyethylene (EO=10) alkyl (R=12-14) ethers (product name: PANNOX710, manufactured by Pan Asia Chemical Corporation), alkyl (R=9-11) glucosides (product name: Mydol 10, manufactured by Kao Corporation), polyoxyethylene (EO=9) lauryl ethers (product name: Emulgen 109P, manufactured by Kao Corporation), and ammonium lauryl sulfate (Emal AD-25, manufactured by Kao Corporation) were suspended in sterile deionized water or various pH standard solutions (Fujifilm Wako Pure Chemical Industries) to achieve the concentrations and pH levels listed in Tables 6-9. Sterile deionized water was used to prepare the pH 5 test solution, neutral phosphate pH standard solution (Fujifilm Wako Pure Chemicals) was used to prepare the pH 7 test solution, and carbonate pH standard solution (Fujifilm Wako Pure Chemicals) was used to prepare the pH 10 test solution. The pH of each test solution was also finely adjusted using 2M hydrochloric acid solution and 1M potassium hydroxide solution. Aqueous solutions adjusted to the pH shown in the table using the aforementioned acid, alkali, pH standard solution, and sterile deionized water were used as controls. These controls did not contain polymers or activators. Units in the table are "mass%". The table shows the pH, feline calicivirus amount (pfu / mL), and HNV genome copy number [copies / μL] after mixing the test solution and virus solution. The virus inactivation effect of the test solutions was evaluated using the method described below.

[0057] <Virus Inactivation Evaluation Method> (1) FCV inactivation evaluation method The tests were conducted with reference to the "FY2015 Survey Report on Norovirus Inactivation Conditions" (National Institute of Health Sciences) and ASTM-E1052, which are generally known as standard methods for evaluating virus inactivation. First, feline calicivirus solutions (F-9 strain, ATCC VR-782) with log pfu / mL values ​​of approximately 6 and 6.3, respectively, were prepared. The virus solution with log pfu / mL of approximately 6 was used in the tests shown in Table 6, and the virus solution with log pfu / mL of approximately 6.3 was used in the tests shown in Tables 7 and 8. In a 1.5 mL sample tube, 90 μL of each test solution shown in the table and 10 μL of the above-mentioned feline calicivirus solution (F-9 strain, ATCC VR-782) were added and mixed thoroughly. After reacting at 30°C for the time indicated in the table, the reaction was stopped by diluting 10-fold with 900 μL of quench solution (SCDLP solution). After stopping the reaction, the solution was serially diluted 10-fold, 100-fold, and 1000-fold in DMEM medium. 0.5 mL of each of these dilutions was then used to infect CRFK cells (feline kidney-derived cell line) (ATCC CCL-94) that had been washed with PBS (phosphate-buffered saline). After 1 hour, the medium was replaced with methylcellulose-containing medium. After culturing at 37°C for 2 days, the cells were stained with crystal violet and the number of plaques was counted.

[0058] (2) HuNoV inactivation evaluation method (i) Culturing of human intestinal epithelial organoids (hIEOs) hIEOs were embedded in Matrigel (Corning) on ​​24-well plates and then cultured in three dimensions under conditions of 37°C and 5% CO2. IntestiCult Organoid Growth Medium (Human) (STEMCELL Technologies) was used as the culture medium. The entire volume of medium (500 μL / 2 days) was changed every 2-3 days, and subculturing was performed every 5-7 days. After 3D culture, the hIEOs were suspended in TrypLE Express (Thermo Fisher Scientific) along with the Matrigel. The hIEOs were dispersed by enzymatic treatment at 37°C for 10 minutes, and to stop the reaction, a basal medium containing Advanced DMEM / F-12 (Gibco) supplemented with 100 U / mL Penicillin-Streptomycin (Thermo Fisher Scientific), 10 mM HEPES (Thermo Fisher Scientific), and 1×GlutaMAX (Thermo Fisher Scientific) was added. After settling the cells by centrifugation (3 minutes, 300×g, 4°C), the above medium was removed. Cells were suspended in differentiation medium supplemented with 1×B27 supplement (Thermo Fisher Scientific), 10 nM gastrin I (Sigma-Aldrich), 1 mM N-acetylcysteine ​​(Fujifilm Wako), 50 ng / mL mouse EGF recombinant protein (Thermo Fisher Scientific), 100 ng / mL mouse Noggin (Peprotech), 500 μg / mL human R-spondin-1 (R&D Systems), 500 nM A83-01 (Tocris), and 10 μM Y-27632 (Fujifilm Wako) in basal medium. Furthermore, for monolayer culture, 96-well flat-bottom plates coated with cell matrix type IC (Nitta Gelatin) were prepared. To prepare the plates, 100 μL / well of cell matrix type IC (Nitta Gelatin), diluted 10-fold with 0.1 M acetic acid (Nacalai tesque), was added, followed by incubation at 37°C for 90 minutes. After that, the solution was removed and the plates were washed three times with 200 μL of D-PBS(-) (Nacalai tesque). 5 × 10⁶ cells were then added to the 96-well flat-bottom plates prepared as described above. 4 Cells were seeded at a concentration of 100 μL / well. Differentiation induction was carried out for a total of 6 days, with the entire differentiation medium being replaced every 2 days, to obtain monolayered hIEOs.

[0059] (ii) Preparation of a 10% emulsion of feces containing HNV (HuNoV) The 10% fecal emulsion was prepared from the feces of GII.4 type HNV-infected patients. One tablet of cOmplete protease inhibitor cocktail tablets (Sigma-Aldrich, 11697498001), a protease inhibitor, was suspended in 50 mL of D-PBS(-). For 1 g of feces, it was suspended in 10 mL of D-PBS(-) containing cOmplete and mixed well with a test tube mixer. After standing at 4°C for 20 minutes, it was centrifuged at 2,000×g at 4°C for 10 minutes. The supernatant was collected in a new tube and stored at -80°C until used for the infection experiment.

[0060] (iii) Inactivation treatment of HNV and infection of differentiated hIEOs The 10% fecal emulsion containing HNV was diluted 10-fold with the differentiation medium and filtered using a 1 mL syringe and a Millex HV Filter unit (Millipore, SLHVR04NL). 5 μL of the filtered fecal solution (2.8×10 6A 45 μL test solution containing HNV genome copy equivalent and polymer at a predetermined concentration as shown in Table 9 was mixed and reacted at the predetermined temperature and time shown in Table 9. Next, 0.95 mL of Fetal Bovine Serum (Capricorn Scientific) solution was added to this drug-treated fecal solution. The centrifuge tube was set on a fixed-angle rotor TLA-55 (Beckman coulter) and ultracentrifuged for 1.5 hours at Rmax with a centrifugal force of 186047 × g (equivalent to 55,000 rpm) using an Optima MAX-TL (Beckman coulter), and the supernatant was removed. The pellet was suspended in 100 μL of differentiation medium to prepare the infection solution for hIEOs. After removing the existing medium from the wells (96-well plate), the infection solution prepared in the manner described above was applied to the hIEOs after differentiation induction for 6 days. Incubation was performed at 37°C for 1 hour. After washing three times with 300 μL of basal medium, 250 μL of differentiation medium was added, and the cells were cultured at 37°C under 5% CO2 conditions until sampling. 10 μL of supernatant was collected immediately after the start of culture and again after 6 days of culture. The collected supernatant was centrifuged at 15,000 rpm for 5 minutes to remove cell debris, and then frozen and stored at -80°C until HuNoV genome RNA quantification was performed.

[0061] (iv) RT-qPCR The number of HNV genome copies in the recovered supernatant was quantified using the Norovirus Detection Kit G1 / G2 (Toyobo, FIK-273). The procedure followed the protocol. PCR amplification and data measurement were performed using LightCycler480II (Roche).

[0062] [Table 6]

[0063] [Table 7]

[0064] [Table 8]

[0065] Table 9

Claims

1. (A) an antimicrobial or antiviral agent containing maleic acrylate copolymer or a salt thereof as an active ingredient, and (B) used in combination with a nonionic surfactant.

2. An antimicrobial or antiviral composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant.

3. The antibacterial or viral inactivating agent according to claim 1, or the antibacterial or viral inactivating composition according to claim 2, wherein the bacterium is a bacterium of the genus Staphylococcus, and the virus is an enveloped RNA virus, an envelopeless RNA virus, or an envelopeless DNA virus.

4. The antibacterial or viral inactivating agent according to claim 1, or the antibacterial or viral inactivating composition according to claim 2, wherein the virus is an influenza virus, coronavirus, norovirus, coxsackievirus, or adenovirus.

5. A method for antimicrobial or viral inactivation, comprising applying a composition containing (A) maleic acrylate copolymer or a salt thereof and (B) a nonionic surfactant to an object suspected of bacterial or viral contamination.

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