JP7539772B2 - Human norovirus inactivation agent - Google Patents

Description

The present invention relates to an inactivating agent for human norovirus (HNV).

Norovirus is a non-enveloped (membrane-like structure) RNA virus classified in the Caliciviridae family and Norovirus genus. It is known to have strong resistance to acidity (gastric acid) and to be infectious in small amounts (approximately 10 to 100 viruses).
At present, there is no vaccine or treatment for norovirus, so it is important to prevent infection by removing or inactivating the virus by cleaning and disinfecting cooking utensils, clothing, hands, etc., on which the virus may be attached.

Norovirus has strong physicochemical resistance, so disinfectants containing ethanol or cationic surfactants, which are effective against many bacteria, may not be sufficiently effective against norovirus when used in the usual way. For this reason, chlorine-based disinfectants (sodium hypochlorite, etc.), iodine agents (povidone-iodine, etc.), aldehyde agents (glutaral, etc.), peracetic acid preparations, etc. are used to inactivate norovirus. However, chlorine-based disinfectants have the effect of bleaching the colors and patterns of textiles, which limits their use on textile products and clothing.

Sodium percarbonate is used as an oxygen bleaching agent because it dissociates into sodium carbonate and hydrogen peroxide in an aqueous solution and exerts its bleaching power. Sodium percarbonate has a mild bleaching power and is easy to handle, and has the advantage that it can be used on colored and patterned clothing, unlike chlorine bleaches.

Until now, since there has been no established laboratory culture system for norovirus that infects humans, feline calicivirus (FCV) and murine norovirus (MNV), which are also members of the Caliciviridae family, have been used as surrogate viruses to evaluate the inactivation effect of norovirus. However, FCV is a virus that causes respiratory infections rather than diarrheal infections, and is vulnerable to acids and alkalis, and many differences between FCV and HNV have been reported. It has also been reported that MNV is significantly more vulnerable to alcohol than other enveloped viruses, including HNV (Non-Patent Document 1). In fact, Non-Patent Document 2 reports that alcohol, which has been thought to be effective for some surrogate viruses, is insufficient to inactivate HNV in the recently established HNV culture system.

There have been reports in the past that aqueous solutions of compositions containing sodium percarbonate have an inactivating effect on FCV (Patent Document 1, Non-Patent Documents 3 and 4), but considering the above circumstances, it is not certain whether the aqueous solutions of compositions containing sodium percarbonate, whose effects on FCV have been reported in Patent Document 1 and Non-Patent Documents 3 and 4, can actually exert a sufficient effect on HNV.

JP 2015-78479 A

Cromeans Theresa et al., Appl. Environ. Microbiol. 80.18 (2014): 5743-5751. Costantini, Veronica, et al. "Human norovirus replication in human intestinal enteroids as model to evaluate virus inactivation." Emerging infectious diseases 24.8 (2018): 1453. Seiichi Tobe et al., J. Oleo Sci. 61, (4) 211-216 (2012) Hirotaka Takagi, Tomoyoshi Sugiyama, Medicine and Pharmacology, Vol. 57, No. 3, 311-312, March 2007

The present invention relates to providing an HNV inactivating agent that is effective in inactivating HNV, and a method for using the same.

In 2016, the inventors succeeded in culturing HNV using human small intestine organoids (hSIO), which are three-dimensional structures similar to intestinal epithelium (Science, 353(6306), 1387-1393, 2016 Sep. 23). Following this, the inventors three-dimensionally cultured hSIO to obtain 3D organoids, dispersed the 3D organoids to prepare single cells, and then cultured the single cells in a monolayer on an extracellular matrix and induced their differentiation to obtain intestinal organoids. They constructed an evaluation system in which the intestinal organoids were infected with HNV, and used this to evaluate oxygen bleaching agents. They found that when oxygen bleaching agents were used so that sodium percarbonate was at a certain concentration or higher, an inactivation effect against HNV was exhibited.

That is, the present invention relates to the following 1) and 2).
1) An HNV inactivator comprising a composition containing sodium percarbonate as an active ingredient, which is used by dissolving in water so that the sodium percarbonate concentration is 0.02 mass % or more.
2) A method for inactivating HNV, comprising the step of immersing an object contaminated with HNV in a treatment liquid containing a composition containing sodium percarbonate and water, wherein the sodium percarbonate concentration in the treatment liquid is 0.02% by mass or more.

According to the present invention, HNV attached to clothing, textiles, cooking utensils, etc. can be reliably inactivated, and the spread of HNV infection can be prevented or reduced.

<Composition>
The compositions of the present invention contain sodium percarbonate.
The HNV inactivating agent of the present invention can exert an HNV inactivating effect by sodium percarbonate. From this viewpoint, it is preferable to use sodium percarbonate having an effective oxygen concentration of 9 to 13 mass% as the sodium percarbonate. The effective oxygen concentration can be measured at 20°C using a standard aqueous potassium permanganate solution. Alternatively, the effective oxygen concentration can be measured by the method described in JIS L0889 Appendix A.

In terms of the HNV inactivation effect, the content of sodium percarbonate in the composition of the present invention is preferably 20% by mass or more, more preferably 25% by mass or more, more preferably 35% by mass or more, and preferably 90% by mass or less, more preferably 80% by mass or less, more preferably 70% by mass or less. Also, it is preferably 20 to 90% by mass, more preferably 25 to 80% by mass, more preferably 35 to 70% by mass. The concentration of sodium percarbonate in the composition can be determined from the mass ratio when the composition is prepared.

The composition of the present invention preferably contains an alkaline agent from the viewpoint of improving the virus inactivation effect.
The alkaline agent is preferably at least one selected from carbonates, hydrogen carbonates, dihydrogen phosphates, dihydrogen phosphates, and triphosphates, and is preferably at least one salt selected from sodium salts, potassium salts, and ammonium salts.
Of such alkaline agents, preferred are sodium carbonate, potassium carbonate, ammonium carbonate, sodium hydrogen carbonate, ammonium hydrogen carbonate, and tripotassium phosphate, and more preferred is sodium carbonate.
As the sodium carbonate, light ash, dense ash, etc. can be used. Among these, dense ash having an average particle size of 300±200 μm, more preferably 300±100 μm, is preferred.

The content of the alkaline agent in the composition is preferably 1% by mass or more, more preferably 2% by mass or more, more preferably 10% by mass or more, and is preferably 60% by mass or less, more preferably 55% by mass or less, more preferably 50% by mass or less. Also, it is preferably 1 to 60% by mass, more preferably 2 to 55% by mass, more preferably 10 to 50% by mass.

The compositions of the present invention may contain a bleach activator.
Here, the bleach activator means a compound that generates an organic peracid by reacting with an inorganic peroxide. As the bleach activator, a hydrophobic bleach activator is preferable. Specifically, a compound having an ester bond represented by the following general formula (1) can be mentioned.
R ¹ -C(=O)-LG (1)
[In the formula, ^R1 is a linear or branched alkyl or alkenyl group having 8 to 14 carbon atoms, an aryl group, or an alkyl group-substituted aryl group, and is preferably a linear or branched alkyl group having 10 to 14 carbon atoms. LG is a leaving group.]

Leaving groups LG include, for example,

[In the formula, ^R2 represents a divalent saturated hydrocarbon group, p represents 0 or 1, and M represents a hydrogen atom or an alkali metal.]
The carbon number of ^R3 is preferably 1 to 5.

The bleach activator is not limited to the compound represented by formula (1), and any bleach activator that has been conventionally used can be used.
Examples of the bleaching activator include tetraacetylethylenediamine, glucose pentaacetate, tetraacetylglycoluril, alkanoyl or alkenoyl (these groups have 8 to 14 carbon atoms) oxybenzene carboxylic acid or a salt thereof, and alkanoyl or alkenoyl (these groups have 8 to 14 carbon atoms) oxybenzene sulfonate. Of these, alkanoyl or alkenoyl (these groups have 8 to 14 carbon atoms, preferably 10 to 14 from the viewpoint of bleaching effect) oxybenzene carboxylic acid or a salt thereof, and alkanoyl or alkenoyl (these groups have 8 to 14 carbon atoms, preferably 10 to 14 from the viewpoint of HNV inactivation effect) oxybenzene sulfonate are preferred, and among these, decanoyloxybenzene carboxylic acid or a sodium salt thereof, and dodecanoyloxybenzene sulfonic acid or a sodium salt thereof are preferred. Any one of these bleaching activators or a combination of two or more thereof can be used.

The content of the bleaching activator in the composition of the present invention is preferably 0.1% by mass or more, more preferably 0.5% by mass or more, and is preferably 20% by mass or less, more preferably 15% by mass or less, more preferably 10% by mass or less. It is also preferably 0.1 to 20% by mass, more preferably 0.5 to 15% by mass, more preferably 0.5 to 15% by mass.

The composition of the present invention may contain a surfactant.
As the surfactant, any of nonionic surfactants, anionic surfactants, cationic surfactants and amphoteric surfactants can be used.

Here, examples of the nonionic surfactant include compounds obtained by adding an alkylene oxide to an aliphatic alcohol, and among these, nonionic surfactants represented by the following general formula (2) are preferred.
R ³ -O-[(EO) _a /(PO) _b ]-H (2)
[In the formula, ^R3 represents an alkyl group having 8 or more, preferably 10 or more, more preferably 12 or more, and 22 or less, preferably 18 or less, more preferably 14 or less carbon atoms, preferably a straight-chain alkyl group. EO represents an ethyleneoxy group, and PO represents a propyleneoxy group. a and b represent the average number of moles added, a represents a number of 0 or more and 20 or less, and b represents a number of 0 or more and 20 or less, and both cannot be 0 at the same time.]
In addition, in the general formula (2), the EO groups and the PO groups may be bonded either randomly or in blocks.

As the cationic surfactant, a quaternary ammonium salt represented by the following general formula (3) is preferably used.

[In the formula, at least one of R ⁴ , R ⁵ , R ⁶ and R ⁷ represents a linear or branched alkyl or alkenyl group having 6 to 20 carbon atoms, preferably 8 to 18 carbon atoms, and the remaining represent an alkyl or hydroxyalkyl group having 1 to 3 carbon atoms. In addition, L ^- represents an organic or inorganic anionic group.]

Specific examples of these quaternary ammonium salts include the following compounds:

(In the formula, h is a number of 6 or more and 20 or less, preferably 8 or more and 18 or less; g and d are the same or different and each independently is a number of 6 or more and 20 or less, preferably 8 or more and 18 or less; ^R8 is an alkyl group or _hydroxyalkyl group _having 1 ^to ₃ carbon atoms; and ^Q- is an organic or inorganic anionic group, preferably a halogen ion such as Cl- ^{or Br-} , an alkyl sulfate ion such as _CH3SO4- ^or _CH3CH2SO4- , or _a fatty acid ion such as _C11H23COO- ^or _CH3COO- . ⁾

Among these, cationic surfactants selected from the group consisting of dioctyldimethylammonium salt, didecyldimethylammonium salt, didodecyldimethylammonium salt, dodecyltrimethylammonium salt, tetradecyltrimethylammonium salt, tetradecyldimethylethylammonium salt, and hexadecyltrimethylammonium salt are preferred.

Examples of amphoteric surfactants include carbobetaine, sulfobetaine, and hydroxysulfobetaine.

Examples of anionic surfactants include alkyl sulfate ester salts, alkyl ether sulfate ester salts, alkylbenzenesulfonic acid or its salts, alkanesulfonic acid or its salts, saturated or unsaturated fatty acid salts, alkyl or alkenyl ether carboxylate salts, α-sulfofatty acid salts, and α-sulfofatty acid esters. Among these, alkylbenzenesulfonic acid or its salts are preferred.

From the viewpoint of ensuring virus removal properties, the content of the surfactant in the composition of the present invention is preferably 0.5% by mass or more, more preferably 0.75% by mass or more, more preferably 1% by mass or more, and also preferably 15% by mass or less, more preferably 10% by mass or less, more preferably 6% by mass or less, and even more preferably 3% by mass or less. Also, it is preferably 0.5 to 15% by mass, more preferably 0.75 to 10% by mass, more preferably 1 to 6% by mass, and even more preferably 1 to 3% by mass.

The composition of the present invention may further contain an acrylic acid polymer as a polymer.
The acrylic acid polymer may be a polymer having a monomer constituent unit derived from one or more monomers selected from acrylic acid, methacrylic acid, and salts thereof (hereinafter, referred to as acrylic acid monomers), and may be a homopolymer of the acrylic acid monomer or a copolymer containing the acrylic acid monomer as a monomer constituent unit.
The homopolymer of the acrylic acid monomer may be a homopolymer of a monomer selected from acrylic acid, methacrylic acid, and salts thereof. The copolymer of the acrylic acid monomer may be a copolymer of at least one selected from acrylic acid, methacrylic acid, and salts thereof, and a compound having an unsaturated bond copolymerizable with these compounds, more preferably a copolymer of a vinyl monomer or an allyl monomer. The copolymer may be one in which the monomer constituent unit derived from acrylic acid or methacrylic acid accounts for 30 mol% or more, preferably 50 mol% or more, more preferably 80 mol% or more of the total monomer constituent units forming the copolymer.
In these homopolymers and copolymers, the salt is preferably a salt with a cation selected from alkali metals, alkaline earth metals, and amines. That is, examples of the salt include alkali metal salts, alkaline earth metal salts, and amine salts, and sodium salts or potassium salts are preferred. The proportion of the monomer constituent units having the salt structure may be a part or the whole, and can be changed depending on the degree of neutralization of the polymer.

Examples of monomers copolymerizable with acrylic acid monomers include vinyl monomers. Examples of vinyl monomers include alkyl acrylate esters having an alkyl group with 8 or more, preferably 10 or more, and 40 or less, preferably 36 or less carbon atoms, alkyl methacrylate esters having an alkyl group with 8 or more, preferably 10 or more, and 40 or less, preferably 36 or less carbon atoms, maleic acid, maleic anhydride, hydroxyethyl acrylic acid, hydroxyethyl methacrylic acid, ethylene, propylene, n-butene, isobutene, pentene, isoprene, 2-methyl-1-butene, n-hexene, 2-methyl-1-pentene, 3-methyl-1-pentene, 4-methyl-1-pentene, 2-ethyl-1-butene, styrene, α-methylstyrene, vinyltoluene, vinylnaphthalene, indene, butadiene, cyclopentadiene, and dicyclopentadiene. Of these, water-soluble monomers such as maleic acid, maleic anhydride, hydroxyethyl acrylic acid, and hydroxyethyl methacrylic acid, and salts of these monomers are preferred.
Of these, homopolymers formed from monomers selected from acrylic acid, methacrylic acid and salts thereof are preferred, and the salts of the homopolymers are preferably potassium salts or sodium salts.

From the viewpoint of handling and dispersibility, the weight-average molecular weight of the acrylic acid polymer is preferably 3,000 to 70,000. The weight-average molecular weight can be measured by gel permeation chromatography using a mixed solution of acetonitrile and water (phosphate buffer) as the developing solvent and polyacrylic acid as the standard substance.

In addition to the above-mentioned components, the composition of the present invention may also contain known components that are usually added to powdered bleach compositions. For example, water; other inorganic salts such as Glauber's salt; fluorescent brighteners; sequestering agents; colorants such as dyes and pigments; and various other trace additives such as fragrances, silicones, bactericides, and ultraviolet absorbers may be blended as necessary.

The composition of the present invention is in the form of a powder. Here, the powder includes granular and particulate forms.
A powder composition can be obtained by mixing only powder components, or by mixing a powder component and a liquid component.
For example, sodium percarbonate and the bleaching activator may be blended in advance as particles. Examples of sodium percarbonate particles include those obtained by granulating sodium percarbonate powder with water, hydrogen peroxide, or the like, and those coated with a nonionic surfactant, an acrylic acid-based polymer, or the like.
The average particle size of the composition is preferably 250 μm or more, more preferably 300 μm or more, and is preferably 1000 μm or less, more preferably 800 μm or less. Here, the average particle size is the 50% particle size value based on the particle size distribution by the dry sieving method described in Japanese Industrial Standards JIS K0069 (1992).
The bulk density is preferably 600 g/cm ³ or more, more preferably 700 g/cm ³ or more, and is preferably 1100 g/cm ³ or less, more preferably 1000 g/cm ³ or less.
Furthermore, the pH of the aqueous solution during dissolution is preferably 9.0 or more, more preferably 9.5 or more, and is preferably 11.5 or less, more preferably 11.0 or less. The pH is measured at 20° C. in accordance with JIS K 3362; 2008, item 8.3.

<HNV inactivation>
The composition of the present invention is diluted with water and used to inactivate HNV in objects contaminated with the virus. The water can be tap water or leftover bath water.
That is, the method for inactivating HNV using the composition of the present invention comprises a step of immersing or contacting an object contaminated with HNV (object to be treated) with a treatment solution (a solution obtained by diluting an oxygen-based bleach composition with water) containing the composition of the present invention and water.

Noroviruses are divided into seven genogroups (GI to GVII) based on the homology of their genome sequences. Among them, the HNV that infects humans are nine GI types (GI.1, GI.2, GI.3, GI.4, GI.5, GI.6, GI.7, GI.8, GI.9), 19 GII 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 one GIV type (GIV.1). The HNV in the present invention may be of any of these genotypes.

Examples of objects contaminated with HNV include textile products such as clothing, bedding, carpets, curtains, bags, and shoes, cooking utensils, washing machines, bathtubs, toilets, etc. The objects to be treated are preferably immersed in or contacted with a solution (treatment liquid) obtained by diluting the composition with water as described above.
The concentration of the composition in the treatment liquid is preferably 0.1% by mass or more, more preferably 0.3% by mass or more, and is preferably 1.5% by mass or less, more preferably 1% by mass or less.

The concentration of sodium percarbonate in the treatment solution is 0.02% by mass or more, preferably 0.1% by mass or more, and more preferably 0.2% by mass or more, from the viewpoint of inactivation of HNV. From the viewpoint of solubility, it is preferably 14% by mass or less, and more preferably 1% by mass or less. It is also preferably 0.02 to 14% by mass, more preferably 0.1 to 1% by mass, and more preferably 0.2 to 1% by mass. It is preferable to mix the composition of the present invention with water to achieve this concentration.

The concentration of the alkaline agent in the treatment solution is preferably 0.02% by mass or more, more preferably 0.1% by mass or more. From the viewpoint of solubility, it is preferably 22% by mass or less, more preferably 1% by mass or less. It is also preferably 0.02 to 22% by mass, more preferably 0.1 to 22% by mass, more preferably 0.1 to 1% by mass. It is preferable to mix the composition of the present invention with water to achieve this concentration.

The bath ratio of the treatment liquid to the object to be treated (mass ratio of treatment liquid/object to be treated) is preferably 10 or more, more preferably 20 or more, and preferably 200 or less, more preferably 100 or less. For objects to be treated that are difficult to immerse, the treatment liquid can be applied, and further, to enhance the treatment effect, the object to be treated can be covered with paper, wrap, cloth, etc. soaked in the treatment liquid.

The temperature of the treatment liquid is preferably 20° C. or higher, more preferably 25° C. or higher, and preferably 50° C. or lower, more preferably 40° C. or lower. In general, immersion is preferably carried out in a treatment liquid prepared by dissolving the composition in water (20 to 25° C.) or lukewarm water (30 to 40° C.).
The treatment time is preferably 5 minutes or more, more preferably 10 minutes or more, more preferably 15 minutes or more, and is preferably 120 minutes or less, more preferably 60 minutes or less, more preferably 30 minutes or less, and is preferably 5 to 120 minutes, more preferably 5 to 60 minutes, more preferably 10 to 60 minutes.
In particular, it is preferable to immerse the material in lukewarm water (30 to 40° C.) for 5 to 30 minutes, and in cold water (20 to 25° C.) for 30 to 60 minutes.

The treatment can be carried out in a suitable container. When the object to be treated is a textile product such as clothing, the treatment can be carried out in the washing tub of a washing machine, and the product may be optionally stirred during immersion.
When the object to be treated is a textile product, after immersion, the garment can be taken out and the excess treatment liquid can be removed by squeezing it out, etc., and then the garment can be washed. In addition, depending on the degree of contamination released into the treatment liquid or from the viewpoint of hygiene, the object after immersion can be rinsed with water as necessary.

Examples 1 to 10, Comparative Example 1
Powdered compositions A to K containing sodium percarbonate were suspended in water (autoclaved tap water) in a usual manner to give the dissolution concentrations shown in Table 1. The compositions of the main components in the treatment solutions are shown in Table 1. The HNV (HuNoV) inactivation effect of the treatment solutions was evaluated by the method described below. The results are also shown in Table 1.
As a result, the proliferation of HNV could not be suppressed in the comparative example in which the sodium percarbonate concentration in the treatment solution was 0.02% or less. On the other hand, it was found that the proliferation of HNV was suppressed in the examples in which the sodium percarbonate concentration in the treatment solution was 0.02% or more. In particular, the proliferation of HNV was suppressed to below the lower limit of quantification of the gene number (50 copies/μL) by treatment with lukewarm water, and HNV was completely inactivated.

<Evaluation method>
(1) Culture of human small intestine organoid (hSIO) hSIO was embedded in Matrigel (Corning, 356231) on a 48-well plate and cultured three-dimensionally. The medium used was IntestiCult Organoid Growth Medium (Human) (STEMCELL Technologies, ST-06010). The procedures for medium replacement, subculture, and monolayer formation using a 96-well plate were performed according to the user manual. For two days after trypsin treatment, CultureSure Y-27632 (Fujifilm Wako Pure Chemical Industries, 036-24023), a ROCK (Rho-associated coiled-coil forming kinase) inhibitor, was added to the medium at a final concentration of 10 μM to inhibit anoikis. A basal medium was prepared by adding 5 mL of GlutaMAX I (100x) (Gibco, 35050-061), 5 mL of 1 M HEPES (Gibco, 15630080), and 5 mL of Penicillin-Streptomycin (Gibco, 15140122) to 500 mL of Advanced DMEM/F12 (Gibco, 12634010). A differentiation medium was prepared by mixing equal amounts of the basal medium and component A of IntestiCult Organoid Growth Medium. Differentiation was induced for a total of 6 days by replacing the differentiation medium at 200 μL per well at 2-day intervals with cells monolayered in a 96-well plate.

(2) Preparation of 10% emulsion of HNV (HuNoV)-containing feces A 10% emulsion of feces was prepared from feces of a patient with GII.4 type HNV. One tablet of cOmplete protease inhibitor cocktail tablets (Sigma-Aldrich, 11697498001), a protease inhibitor, was suspended in 50 mL of D-PBS(-). 1 g of feces was suspended in 10 mL of cOmplete-containing D-PBS(-) and mixed well with 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 used in an infection experiment.

(3) Inactivation of HNV and Infection of Differentiated hSIO 10% fecal emulsion containing HNV was diluted 10-fold with differentiation medium and filtered using a 1 mL syringe and Millex HV Filter unit (Millipore, SLHVR04NL). 5 μL of the filtered fecal solution (corresponding to 2.8×10 ⁶ HNV genome copies) was mixed with 45 μL of a suspension (treatment solution) of a composition containing a specific concentration of sodium percarbonate shown in Table 1 in a PA microcentrifuge tube (Beckman Coulter, 357448) and reacted at a specific temperature and time shown in Table 1. Next, 1.45 mL of basal medium containing 1 mg/mL bovine serum albumin (Sigma-Aldrich, A9647) was added to the drug-treated fecal solution. The centrifuge tube was placed in a fixed-angle rotor TLA-55 (Beckman Coulter) and ultracentrifuged for 1.5 hours at Rmax with an Optima MAX-TL (Beckman Coulter) at a centrifugal force of 186047×g (corresponding to 55,000 rpm), and the supernatant was removed. The pellet was suspended in 100 μL of differentiation medium to prepare an infection solution for hSIO. The infection solution prepared by the above method was applied to hSIO after differentiation induction for 6 to 7 days from which the existing medium in the well had been removed. Incubation was performed at 37° C. for 3 hours. After washing three times with 300 μL of basal medium, 250 μL of differentiation medium containing porcine bile extract (Sigma-Aldrich, B8631-100G) added to a final concentration of 125 ppm was added, and the cells were cultured under conditions of 37 ° C. and 5% CO ₂ until the timing of sampling. 10 μL of supernatant was collected immediately after the start of culture (day 0) and after 3 days of culture (day 3). The collected supernatant was stored at -80 ° C. until subjected to RT-qPCR. In addition, the infection solution for hSIO prepared by performing the same operation using basal medium instead of the drug solution was used as a drug-free control.

(4) RT-qPCR
The number of HNV genome copies in the collected supernatant was quantified using the Norovirus Detection Kit G1/G2 (Toyobo, FIK-273). The procedure was performed according to the protocol. PCR amplification and data measurement were performed using a LightCycler 480II (Roche). there was.
Based on the measured viral load, A: HNV is inactivated to the lower limit of detection, B: HNV is detected but reduced compared to the control (no drug treatment), C: equivalent to the control (no drug treatment). The results were evaluated on a three-point scale: HNV proliferation, cytopenia, or cytopenia.

Claims (5)

A human norovirus inactivation agent comprising a composition containing sodium percarbonate and sodium carbonate as active ingredients (excluding compositions containing one or more compounds selected from the group consisting of compounds represented by the following general formula (I) and compounds represented by the following general formula (II)), which is used by dissolving in water so that the sodium percarbonate concentration is 0.2% by mass or more and 0.479% by mass or less and the sodium carbonate concentration is 0.1% by mass or more and 1% by mass or less.

[In the formula, R ¹⁰ and R ²⁰ each independently represent an alkyl group or an alkenyl group having 7 to 13 carbon atoms. M ¹ and M ² each independently represent a hydrogen atom or a salt-forming cation.] The human norovirus inactivating agent according to claim 1, wherein the temperature of the water is 20 to 50°C. A method for inactivating human norovirus, comprising a step of immersing or contacting an object (excluding humans) contaminated with human norovirus with a treatment liquid containing a composition containing sodium percarbonate and sodium carbonate (excluding compositions containing one or more compounds selected from the group consisting of compounds represented by the following general formula (I) and compounds represented by the following general formula (II) ) and water, wherein the sodium percarbonate concentration in the treatment liquid is 0.2% by mass or more and 0.479% by mass or less, and the sodium carbonate concentration is 0.1% by mass or more and 1% by mass or less.

[In the formula, R ¹⁰ and R ²⁰ each independently represent an alkyl group or an alkenyl group having 7 to 13 carbon atoms. M ¹ and M ² each independently represent a hydrogen atom or a salt-forming cation.] The method according to claim 3, in which the substrate is immersed in a treatment solution at 20 to 50°C for 5 to 120 minutes. The method according to claim 3 or 4, wherein the object is a textile product.