JP6537611B2 - How to kill - Google Patents

Description

  The present invention relates to a method of killing spore forming bacteria.

  Spore-forming bacteria such as Bacillus and Clostridia form a strong shell structure and form spores having extremely high resistance to heat and drugs. It is known that certain spore forming bacteria produce toxins when they enter the human body. For example, in the medical field, linen products such as sheets and pillow covers are mainly heat-disinfected, but heat-disinfection can not kill spore forming bacteria that are heat resistant. For this reason, nosocomial infection with spore forming bacteria through linen products has caused damage to the dead.

  In order to kill spore forming bacteria, sterilization with high pressure steam is often carried out, or a strong chemical bactericide such as sodium hypochlorite is used at a high concentration. Also, in food processing, severe heat treatment or low temperature distribution method is taken as a measure against spore.

US 2014/03 08 162 discloses antiseptic and rinse compositions for use in laundry detergents. Also, US 2014/0238445 discloses a method of cleaning, disinfecting and rinsing, including a cleaning step using a composition containing phosphinosuccinic acid and a disinfecting and rinsing step using a composition containing percarboxylic acid. ing. Furthermore, WO 2013/079308 discloses the sterilization of bacteria containing spores. US2004 / 0,058,878 discloses are disclosed for improving the stop effect by combining germinating agent and quaternary ammonium salts.

The present invention relates to a method of killing which performs the following step 1, step 2 and step 3.
Step 1: contacting the spore forming bacteria with dipicolinic acid or a salt thereof;
Step 2: a step of contacting the spore-forming bacteria with a cationic surfactant; and Step 3: a step of heating the spore-forming bacteria to 50 ° C. or higher.
(However, step 3 shall be ended after step 1 and step 2 are started).

  The present invention relates to a germicidal auxiliary composition comprising dipicolinic acid or a salt thereof and a cationic surfactant.

Detailed Description of the Invention

  The techniques disclosed so far do not have a sufficient bactericidal effect on spores, that is, dormant spore-forming bacteria, and the effect of reducing the number of spore bacteria is insufficient.

  High concentrations of disinfectants and sterilization methods using high-pressure steam sterilization are not suitable for disinfecting a wide range of environments such as hospitals, and there may be equipment damage and toxicity to the human body. There are restrictions on the use environment.

  In food processing, severe heat treatment or low temperature distribution is used as a measure against spores, but problems such as deterioration of flavor and quality, and power cost also arise.

  The present invention relates to a killing method capable of efficiently and effectively disinfecting spore forming bacteria which are harmful in the medical and food fields. Furthermore, the present invention relates to a slaughtering aid composition used to kill spore forming bacteria. In the present invention, "slaughter" is mainly intended to germinate and kill bacteria having a spore.

  MEANS TO SOLVE THE PROBLEM As a result of conducting earnest examination in view of the said subject, this inventor is a mild by keeping dipicolinic acid (2, 6-pyridine dicarboxylic acid) or its salt, and a cationic surfactant in contact with a spore forming microbe. The inventors have found that heating can effectively kill cells, and have led to the present invention.

  According to the present invention, there is provided a method of killing which can kill spore forming bacteria. In addition, the present invention provides a slaughtering aid composition used to kill spore forming bacteria.

Spore-forming bacteria are bacteria that are resistant to certain heat treatments and drying by forming spores in the absence of nutrients. A spore shows a strong shell structure formed by bacteria, formed by bacteria, and distinguished from the bacteria itself. The "spore-forming bacteria" to be slaughtered according to the present invention are general spore-forming bacteria which are present in medical fields, foods, and beverage products. For example, bacteria of the genus Bacillus such as Bacillus cereus and Bacillus subtilis, bacteria of the genus Clostridium such as Clostridium difficile, bacteria of the genus Amphibacillus And Sporasarcina bacteria, Geobacillus bacteria, Aeribacillus bacteria, Alicyclobacillus bacteria and the like. When these spore-forming bacteria form spores, the spore-forming bacteria have thermostability. Here, heat resistance means, as shown by the spore forming bacteria provided in the examples of the present invention, a spore having an initial number of bacteria of 10 ⁷ to 10 ⁹ CFU / mL for 30 minutes at 80 ° C. When heated above, it means that 10 ⁷ CFU / mL or more of spore forming bacteria can survive.

In the present invention, "slaughter" is mainly intended to germinate and kill bacteria having a spore. Germination means that the spores of the spore forming bacteria germinate, and thereby refers to the phenomenon of becoming a cell having normal growth and metabolic ability. Killing also includes reducing the overall number of bacteria with spores present in the population. The killing effect is not particularly limited. For example, as shown in the examples, the viable count of bacteria having spores is 10 ⁷ CFU / mL, the initial number of which is 10 ⁸ CFU / mL. Indicates a state of less than.

The present invention has an excellent germicidal effect by performing the following step 1, step 2 and step 3.
Step 1: contacting the spore forming bacteria with dipicolinic acid or a salt thereof;
Step 2: a step of contacting the spore-forming bacteria with a cationic surfactant; and Step 3: a step of heating the spore-forming bacteria to 50 ° C. or higher.
(However, step 3 shall be ended after step 1 and step 2 are started).

  In the present invention, the order of steps 1, 2 and 3 is not particularly limited. However, after the process 1 and the process 2 are started, the process 3 is completed. That is, in the series of operations including Step 1, Step 2 and Step 3, it is not included to start either Step 1 or Step 2 or Step 2 after Step 3 is completed.

  The reason why the germicidal method of the present invention has an excellent germicidal effect is considered as follows, though it is not clear. By performing Step 1, Step 2 and Step 3, the spores germinate. It is assumed that the cationic surfactant enhances the affinity between dipicolinic acid or its salt and the spore forming bacteria, and dipicolinic acid or its salt is likely to enter the spore forming bacteria, and that germination is effectively promoted upon heating. Ru. The germinated spore-forming bacteria can be easily sterilized at steps 2 and 3.

  In addition, since the action of dipicolinic acid or a salt thereof in a spore-forming bacterium is not an action mediated by a specific receptor specific to the species, it is considered to have a killing effect regardless of the type of spore-forming bacterium.

  In addition, it can confirm having germinated here by confirming the heat resistant fall of a microbe. The decrease in heat resistance can be confirmed by, for example, treating the spore forming bacteria under an environment of 80 ° C. and examining the number of surviving bacteria.

  In the present invention, when dipicolinate is used, the counter ion of dipicolinic acid is not particularly limited, and examples thereof include alkali metal ions such as sodium ion and potassium ion; alkaline earth metal ions such as calcium ion and magnesium ion Ru.

  In the present invention, the term "cationic surfactant" refers to a surfactant whose hydrophilic moiety is charged to a cation when it is dissolved in a solution, and is not limited to, but is not limited to, primary ammonium salt, secondary ammonium salt And tertiary ammonium salts, quaternary ammonium salts and the like. More specifically, the cationic surfactant of the present invention is alkyl trimethyl ammonium salt, alkyl triethyl ammonium salt, alkyl dimethyl ethyl ammonium salt, alkyl methyl diethyl ammonium salt, dialkyl dimethyl ammonium salt, dialkyl diethyl ammonium salt, dialkyl ethyl methyl Examples thereof include ammonium salts, quaternary ammonium salts such as benzalkonium salts, alkyl pyridinium salts and alkyl benzethonium salts, and primary ammonium salts such as alkylamine salts.

  Here, the alkyl trimethyl ammonium salt, the alkyl triethyl ammonium salt, the alkyl dimethyl ethyl ammonium salt and the alkyl methyl diethyl ammonium salt refer to a compound represented by the following general formula (1).

^Wherein any one of R 11 ^{to R 14,} a hydroxyl group, an ester group, an amide group hydrocarbon group having 3 or more carbon atoms which may have a ^remaining three of R 11 ^{to R 14} represents a methyl group or an ethyl group, X ^- represents an inorganic or organic anionic compounds. ]

Here, the carbon number of the hydrocarbon group having 3 or more carbon atoms is preferably 6 or more, more preferably 10 or more, more preferably 12 or more, from the viewpoint of further improving the germicidal effect. 14 or more, and preferably 22 or less, more preferably 20 or less, more preferably 18 or less. The hydrocarbon group having 3 or more carbon atoms may be a linear or branched alkyl group or alkenyl group, preferably a linear alkyl group. As X ⁻ , halogen ions such as chloride ion and bromide ion, sulfate ion, phosphate ion, hydrogen phosphate ion, dihydrogen phosphate ion, nitrate ion, carbonate ion, hydrogen carbonate ion, acetate ion and the like can be mentioned. Preferably it is an inorganic ion, more preferably it is a halogen ion, more preferably it is a chloride ion.

  Here, dialkyl dimethyl ammonium, dialkyl diethyl ammonium salt and dialkyl ethyl methyl ammonium salt refer to a compound represented by the following general formula (2).

[Wherein, any two of R ^{21 to} R ²⁴ are a hydrocarbon group having 3 or more carbon atoms which may have a hydroxyl group, an ester group or an amide group, and the remaining two of R ^{21 to} R ²⁴ represents a methyl group or an ethyl group, X ^- represents an inorganic or organic anionic compounds. Here, any two of R ^{21 to} R ²⁴ may not have the same hydroxyl group, an ester group, or a hydrocarbon group having 3 or more carbon atoms which may have an amide group. ]

Here, the carbon number of the hydrocarbon group having 3 or more carbon atoms is preferably 6 or more, more preferably 10 or more, and preferably 22 or less, more preferably 20, from the viewpoint of further improving the germicidal effect. More preferably, it is 18 or less, more preferably 14 or less. The hydrocarbon group having 3 or more carbon atoms may be a linear or branched alkyl group or alkenyl group, preferably a linear alkyl group. As X ⁻ , halogen ions such as chloride ion and bromide ion, sulfate ion, phosphate ion, hydrogen phosphate ion, dihydrogen phosphate ion, nitrate ion, carbonate ion, hydrogen carbonate ion, acetate ion and the like can be mentioned. Preferably it is an inorganic ion, more preferably it is a halogen ion, more preferably it is a chloride ion.

The benzalkonium salt refers to a compound represented by the following general formula (3).

[Wherein, R ³¹ represents a hydrocarbon group, and X ⁻ represents an inorganic or organic anionic compound. ]

The carbon number of R ³¹ is preferably 6 or more, more preferably 10 or more, more preferably 12 or more, more preferably 14 or more, and preferably 22 or less, more preferably 20 or less, more Preferably it is 18 or less. R ³¹ may be a linear or branched alkyl group or alkenyl group, preferably a linear alkyl group. X ⁻ is preferably a halogen ion such as chloride ion or bromide ion, a sulfate ion, a phosphate ion, a hydrogen phosphate ion, a dihydrogen phosphate ion, a nitrate ion, a carbonate ion, a hydrogen carbonate ion, an acetate ion, etc. Be Preferably it is an inorganic ion, more preferably it is a halogen ion, more preferably it is a chloride ion.

  Here, "alkyl" in the case of referring to alkyl pyridinium salt, alkyl benzethonium salt or alkyl amine salt preferably refers to a hydrocarbon group having 3 or more carbon atoms. The carbon number of the hydrocarbon group having 3 or more carbon atoms is preferably 6 or more, more preferably 10 or more, and preferably 22 or less, more preferably 20 or less, more preferably 18 or less. The hydrocarbon group having 3 or more carbon atoms may be a linear or branched alkyl group or alkenyl group, preferably a linear alkyl group.

  As the cationic surfactant, dialkyldimethyl ammonium salts, alkyl trimethyl ammonium salts or benzalkonium salts are preferable, dialkyl dimethyl ammonium salts and alkyl trimethyl ammonium salts are more preferable, and alkyl trimethyl ammonium salts are further preferable.

  Here, the salt is not limited, but chlorides, halides such as bromides, sulfates, phosphates, hydrogen phosphates, dihydrogen phosphates, nitrates, carbonates, hydrogencarbonates, acetates, etc. Can be mentioned. It is preferably an inorganic salt, more preferably a halide, more preferably a chloride.

  As the cationic surfactant, dialkyldimethyl ammonium chloride, alkyl trimethyl ammonium chloride or benzalkonium chloride is preferable, dialkyl dimethyl ammonium chloride and alkyl trimethyl ammonium chloride are more preferable, and alkyl trimethyl ammonium chloride is more preferable.

  The molar ratio of dipicolinic acid to cationic surfactant ([molar concentration of dipicolinic acid] / [molar concentration of cationic surfactant]) is preferably 1/1000 or more, more preferably 1/100 or more, More preferably 1/50 or more, more preferably 1/20 or more, more preferably 1/10 or more, more preferably 1/5 or more, more preferably 1/2 or more, And preferably 1000 or less, more preferably 100 or less, more preferably 50 or less, more preferably 20 or less, more preferably 10 or less, more preferably 5 or less, more Preferably it is 2 or less.

<Step 1 and Step 2>

Step 1 of the present invention is a step in which a spore forming bacterium and dipicolinic acid or a salt thereof are brought into contact. Step 2 of the present invention is a step in which a spore forming bacterium and a cationic surfactant are brought into contact with each other. Here, step 1 and step 2 include any of the following modes.
(1) An embodiment in which step 1 and step 2 are simultaneously carried out, ie, an embodiment in which contact between spores forming bacteria, dipicolinic acid or a salt thereof and a cationic surfactant are simultaneously initiated and simultaneously terminated.
(2) A mode in which the process 1 is performed first and the process 2 is performed after the process 1 ends. For example, after step 1 in which the spore forming bacteria and dipicolinic acid or a salt thereof are brought into contact with each other, dipicolinic acid or a salt thereof is removed, and then a cationic surfactant is added to the spore forming bacteria and the cationic surfactant. And can be kept in contact with each other.
(3) A mode in which the step 2 is performed first and the step 1 is performed after the step 2 is completed. For example, after step 2 in which the spore forming bacteria and the cationic surfactant are brought into contact with each other, the cationic surfactant is removed, and then dipicolinic acid or a salt thereof is added to the spore forming bacteria and dipicolinic acid or its salt. It can be in contact with salt.
(4) The aspect which performs the process 1 previously and starts the process 2 without complete | finishing the process 1. Here, it is possible to first contact the spore forming bacteria with dipicolinic acid or a salt thereof and then contact the cationic surfactant without removing dipicolinic acid or a salt thereof.
(5) A mode in which step 2 is performed first and step 1 is started without ending step 2. Here, it is possible to first contact the spore forming bacteria with the cationic surfactant, and then contact with dipicolinic acid without removing the cationic surfactant.

  The dipicolinic acid or its salt and the cationic surfactant may be provided in liquid or solid state, respectively. In addition, dipicolinic acid or a salt thereof and a cationic surfactant are preferably provided as a germicidal aid composition from the viewpoint of convenience.

  Here, the term “contact” refers to a method of keeping in contact in a solution, or, when spore forming bacteria are present on a solid surface, killing of the liquid described later, such as a solution containing dipicolinic acid or a salt thereof and a cationic surfactant. The method of apply | coating a sprout adjuvant composition to the solid surface is mentioned. As a method of applying to a solid surface, a method of spraying and a method of using a tool such as a brush or a sponge may be mentioned. Here, the solid surface is not particularly limited, but refers to a hard surface or the like.

  When the spore-forming bacteria and dipicolinic acid or a salt thereof are brought into contact with each other in a solution, the content of dipicolinic acid or a salt thereof in the solution upon contact is preferably from the viewpoint of further improving the germicidal effect. 0.05 mM or more, more preferably 0.5 mM or more, more preferably 3 mM or more, more preferably 4 mM or more, more preferably 6 mM or more, more preferably 8 mM or more, and stability of the solution containing dipicolinic acid From the viewpoint, it is preferably 1 M or less, more preferably 200 mM or less, more preferably 100 mM or less, more preferably 50 mM or less, more preferably 20 mM or less. In addition, when the spore forming bacteria and dipicolinic acid or a salt thereof are brought into contact with each other in a solution, the content of dipicolinic acid or a salt thereof in the solution is preferably 0.05 mM to 1 M in view of the above aspects. More preferably, it is 0.05-200 mM, More preferably, it is 3-100 mM, More preferably, it is 3-25 mM. Here, in the case of dipicolinate, the contents are all converted to acid (hereinafter, the same meaning unless otherwise noted).

The contact between the initial number of bacteria and the concentration of dipicolinic acid or its salt is not particularly limited. For example, from the viewpoint of further improving the killing effect, the concentration of dipicolinic acid or a salt thereof relative to the initial number of bacteria of 10 ⁸ CFU / mL is preferably 0.05 mM or more, more preferably 0.5 mM or more, more preferably 3 mM or more , More preferably 4 mM or more, more preferably 6 mM or more, more preferably 8 mM or more, and preferably 1 M or less, more preferably 200 mM or less, more preferably 100 mM or less, more preferably 50 mM or less, more preferably Is less than 20 mM.

  When the spore forming bacteria and the cationic surfactant are brought into contact with each other in the solution, the content of the cationic surfactant in the solution at the time of the contact is preferably 0. 01 mM or more, more preferably 0.05 mM or more, more preferably 0.1 mM or more, more preferably 0.5 mM or more, more preferably 3 mM or more, more preferably 4 mM or more, more preferably 6 mM or more, more preferably 8 mM or more And preferably not more than 1000 mM, more preferably not more than 500 mM, still more preferably not more than 300 mM, more preferably not more than 100 mM. In addition, when the spore-forming bacteria and the cationic surfactant are brought into contact with each other in the solution, the content of the cationic surfactant in the solution is preferably 0.01 to 1000 mM, preferably from the above viewpoints. Preferably, 0.05 to 1000 mM, more preferably 0.1 to 1000 mM, more preferably 0.5 to 500 mM, more preferably 3 to 300 mM, more preferably 4 to 100 mM, more preferably 6 to 100 mM, more preferably 8 to 100 mM.

  When the spore forming bacteria and the cationic surfactant are brought into contact with each other in the solution, the content of the cationic surfactant in the solution at the time of the contact is preferably 0.01 from the viewpoint of further improving the germicidal effect. % By mass, more preferably 0.05% by mass, and preferably 10% by mass or less, more preferably 5% by mass or less, still more preferably 1% by mass or less. In addition, when the spore forming bacteria and dipicolinic acid or its salt and the cationic surfactant are brought into contact in a solution, the content of the cationic surfactant in the solution is preferably 0. The content is 01 to 10% by mass, more preferably 0.05 to 5% by mass, and still more preferably 0.05 to 1% by mass.

The contact between the initial number of bacteria and the concentration of the cationic surfactant is not particularly limited. For example, from the viewpoint of further improving the killing effect, the concentration of the cationic surfactant with respect to the initial number of bacteria of 10 ⁸ CFU / mL is preferably 0.01 mM or more, more preferably 0.05 mM or more, more preferably 0.1 mM Or more, more preferably 0.5 mM or more, more preferably 1 mM or more, more preferably 10 mM or more, more preferably 100 mM or more, and preferably 1000 mM or less, more preferably 500 mM or less, more preferably 300 mM or less Preferably it is 250 mM or less.

  These preferred concentrations are the same when dipicolinic acid or a salt thereof and the cationic surfactant are simultaneously brought into contact with the spore forming bacteria or are separately brought into contact.

  The temperature of Step 1 or Step 2 which is performed independently of Step 3 before the start of Step 3 of the present invention is preferably independently 0 ° C. or more, more preferably, from the viewpoint of further improving the germicidal effect. Is 10 ° C. or more, more preferably 15 ° C. or more, more preferably 20 ° C. or more, and preferably less than 50 ° C., more preferably 49 ° C. or less, more preferably 45 ° C. or less, more preferably 40 ° C. or less More preferably, it is 30 ° C. or less. The contact temperature between the spore forming bacteria and the solution containing dipicolinic acid or a salt thereof or a cationic surfactant in step 1 and step 2 performed independently of step 3 is preferably 15 ° C. or higher from the same viewpoint. It is less than 50 ° C, more preferably 15 ° C to 40 ° C, more preferably 15 to 30 ° C.

  These preferable temperatures are the same as in the case where Step 1 and Step 2 are carried out simultaneously or in the case where they are carried out separately. However, when Step 1 and Step 2 are carried out separately, Step 1 and Step 2 are carried out. The temperature of 2 may be different or the same.

  The times of step 1 or step 2 performed independently of step 3 of the present invention are each independently preferably 1 second or more, more preferably 5 seconds or more, from the viewpoint of further improving the germicidal effect. Is 30 seconds or more, more preferably 1 minute or more, more preferably 3 minutes or more, more preferably 5 minutes or more, more preferably 8 minutes or more, more preferably 10 minutes or more, more preferably 15 minutes or more, more preferably 20 minutes or more, more preferably 25 minutes or more, and preferably 24 hours or less, more preferably 12 hours or less, more preferably 6 hours or less, more preferably 3 hours or less, more preferably 60 minutes or less, more Preferably it is 50 minutes or less, More preferably, it is 40 minutes or less, More preferably, it is 35 minutes or less. In addition, the contact time of dipicolinic acid or a salt thereof with the spore forming bacteria in step 1 is preferably 1 second to 24 hours, more preferably 1 second to 6 hours, more preferably 1 second to It is 3 hours, more preferably 1 second to 60 minutes, more preferably 5 seconds to 60 minutes, more preferably 30 seconds to 60 minutes.

<Step 3>
Step 3 of the present invention is a step of heating the spore forming bacteria to 50 ° C. or higher.

  Step 3 of the present invention is preferably a step of heating the spore forming bacteria at 50 ° C. or more and 250 ° C. or less.

  The temperature for heating the spore-forming bacteria is 50 ° C. or more, more preferably 55 ° C. or more, more preferably 60 ° C. or more, more preferably 65 ° C. or more, more preferably 70 ° C. from the viewpoint of further improving the germicidal effect. More preferably, 75 ° C. or more, and preferably 250 ° C. or less, more preferably 200 ° C. or less, more preferably 150 ° C. or less, more preferably 120 ° C. or less, more preferably 100 ° C. or less, more preferably It is 90 ° C. or less, more preferably 85 ° C. or less. The temperature at the time of heating in step 3 of the present invention is preferably 50 ° C. to 250 ° C., more preferably 50 ° C. to 100 ° C., more preferably 55 ° C. to 100 ° C., more preferably 60 ° C. C. to 100.degree. C., more preferably 65.degree. C. to 90.degree. C., more preferably 70.degree. C. to 90.degree. C., more preferably 75.degree. C. to 85.degree.

  Usually, the spore forming bacteria is difficult to kill at 100 ° C. or less unless it germinates. However, in the present invention, since germination is promoted by step 1 and step 2, a killing effect can be obtained even at 100 ° C. or lower in step 3. In the present invention, it is also possible to kill at temperatures higher than 100 ° C.

  The heating time in step 3 of the present invention is preferably 3 minutes or more, more preferably 5 minutes or more, more preferably 8 minutes or more, more preferably 10 minutes or more, from the viewpoint of further improving the germicidal effect. Preferably it is 20 minutes or more, more preferably 25 minutes or more, and preferably 3 hours or less, more preferably 90 minutes or less, more preferably 70 minutes or less, more preferably 50 minutes or less, more preferably 40 minutes or less More preferably, it is 35 minutes or less. The heating time in step 3 of the present invention is preferably 3 minutes to 90 minutes, more preferably 5 minutes to 70 minutes, more preferably 8 minutes to 50 minutes, more preferably 10 minutes to 50, from the above viewpoint Minutes, more preferably 20 to 40 minutes, more preferably 25 to 35 minutes. This time may include the same period as step 1 and / or step 2.

  The heating in step 3 of the present invention is not particularly limited as long as the temperature of the system containing the spore forming bacteria is consequently increased. For heating, for example, a method of applying dry heat, moist heat, boiling, hot water, steam heat or the like to the spore forming bacteria may be mentioned. Here, for example, a method of immersing the spore forming bacteria in hot water, a method of placing a solid such as a hard surface on which the spore forming bacteria exist, a liquid containing the spore forming bacteria, etc. A method by heat treatment, a method by which the temperature of the spore rises as a result of irradiation of a laser, etc. are exemplified.

  The pH of the solution at 24 ° C. in step 1, step 2 and step 3 is not particularly limited, but is preferably 3 or more, more preferably 4 or more, still more preferably 6 or more, from the viewpoint of safety Preferably, it is 12 or less, more preferably 9 or less, more preferably 8.5 or less. In addition, the pH of the solution at 24 ° C. is preferably 3 to 12, more preferably 3 to 9, and more preferably 4 to 8.5, from the above viewpoints.

  The pH adjustment can be performed when preparing dipicolinic acid or a salt thereof or a cationic surfactant solution in advance. Alternatively, it can be carried out after adding dipicolinic acid or a salt thereof or a cationic surfactant to a solution containing the spore forming bacteria, while being in contact with the spore forming bacteria. As a pH adjuster, commonly used acids and bases, for example, inorganic acids such as hydrochloric acid and sulfuric acid, organic acids such as lactic acid and citric acid or their salts, inorganic bases such as sodium hydroxide and potassium hydroxide, triisopropanol Organic bases such as amines are mentioned. As the pH adjuster, as the acid, an inorganic acid such as hydrochloric acid or sulfuric acid is preferable, and as the base, an inorganic base such as sodium hydroxide or potassium hydroxide is preferable.

  In the method of killing of the present invention, step 3 can be completed by quenching the spore-forming bacteria heated in step 3 with water containing ice or a cooled container or the like. Alternatively, step 3 can be completed by cooling the heat naturally in a while for a while for a while for the spore forming bacteria warmed in step 3. Here, the heated spore-forming bacteria may be in the state of being present in the liquid.

  When the spore forming bacteria are present in the liquid, the method of killing the present invention may have a step of stirring the spore forming bacteria before or after Step 1, Step 2, and Step 3 or those steps. Step 1, step 2 and step 3 can be repeated.

  The step 3 can be performed after the step 1 and the step 2 are completed (aspect 1).

  In the aspect 1, “performing the step 3 after the end of the step 1 and the step 2” is to carry out the step 3 after removing the dipicolinic acid or the salt thereof and the cationic surfactant, and in the step 1 and the step 2 With the end, it means. Aspect 1 is preferably within 24 hours, more preferably within 12 hours, more preferably within 6 hours, more preferably within 1 hour, more preferably within 30 minutes, still more preferably, after Step 1 and Step 2. Transfer to step 3 within 15 minutes, more preferably within 5 minutes, and even more preferably within 1 minute.

  You may have the period which performs the process 1 or the process 2, and the process 3 simultaneously (aspect 2).

  In the embodiment 2, “having a period in which step 1 or step 2 and step 3 are simultaneously performed” is to carry out step 3 after removing dipicolinic acid or a salt thereof or a cationic surfactant, step 1 or step With the end of 2 means. Aspect 2 is preferably within 24 hours, more preferably within 12 hours, more preferably within 6 hours, more preferably within 1 hour, more preferably within 30 minutes, more preferably within 2 hours after Step 1 or Step 2. Transfer to step 3 within 15 minutes, more preferably within 5 minutes, and even more preferably within 1 minute.

It may have a period in which step 1 and step 2 and step 3 are simultaneously performed (aspect 3). Aspect 3 can be divided as follows.
Embodiment 3-1 Step 1 and / or Step 2 are started prior to Step 3. Embodiment 3-2 Step 1 and Step 2 and Step 3 are started at the same time. Embodiment 3-3 Step 3 before Step 1 and Step 2 Embodiment 3-4 Starting Embodiments of Step 1 or Step 2 and Step 3 Simultaneously In embodiment 3, from the viewpoint of further improving the germicidal effect, embodiment 3-1 is preferable, but not limited thereto.

 The aspect (aspect 3) which has the period which performs the process 1, the process 2, and the process 3 simultaneously is preferable from a viewpoint of improving the slaughtering method of this invention from a slaughtering method further.

<Stericide adjuvant composition>
The germicide composition of the present invention is a composition comprising dipicolinic acid or a salt thereof and a cationic surfactant. The slaughtering aid composition of the present invention is used for warming pretreatment performed for the purpose of germination. It is also distinguished from bactericidal compositions which are intended to kill using the composition itself at ambient temperature (about 20 ° C. to about 38 ° C.). In particular, it refers to a composition that can exert a germicidal effect by heating after Step 1 and Step 2 of the present invention, or simultaneously with Step 1 and Step 2. In the present invention, a slaughtering aid composition may be used in Step 1 and Step 2.

  The slaughtering agent composition includes a liquid composition or a solid composition.

  The liquid germicide composition comprises dipicolinic acid or a salt thereof, a cationic surfactant, and a solvent.

  When the germicide composition is a liquid, the solvent contained includes water or a hydrophilic solvent. Examples of hydrophilic solvents include monohydric alcohols such as ethanol, methanol and isopropanol; polyhydric alcohols such as glycerin, ethylene glycol and propylene glycol; carbitols such as methyl carbitol and ethyl carbitol. The solvent is preferably water or a mixture of water and a hydrophilic solvent, more preferably water, from the viewpoint of further improving the germicidal effect.

  As the liquid spore assistant composition, from the viewpoint of convenience, preferably a solution containing dipicolinic acid or a salt thereof and a cationic surfactant, more preferably an aqueous solution containing dipicolinic acid or a salt thereof and a cationic surfactant is there.

  The liquid germicide composition may further contain a pH adjuster. As a pH adjustor contained in the liquid septic aid composition, generally used acids and bases, for example, inorganic acids such as hydrochloric acid and sulfuric acid, organic acids such as lactic acid and citric acid or salts thereof, sodium hydroxide And inorganic bases such as potassium hydroxide, and organic bases such as monoethanolamine, triethanolamine, triisopropanolamine and the like. As the pH adjuster, as the acid, an inorganic acid such as hydrochloric acid or sulfuric acid is preferable, and as the base, an inorganic base such as sodium hydroxide or potassium hydroxide is preferable.

  The pH at 24 ° C. of the liquid blasting aid composition, preferably a solution comprising dipicolinic acid or a salt thereof and a cationic surfactant, more preferably an aqueous solution comprising dipicolinic acid or a salt thereof and a cationic surfactant, is particularly limited Although not preferred, it is preferably 3 or more, more preferably 4 or more, still more preferably 6 or more, and from the viewpoint of safety, preferably 12 or less, more preferably 9 or less, more preferably 8.5 It is below. In addition, the pH of the solution at 24 ° C. is preferably 3 to 12, more preferably 3 to 9, and more preferably 4 to 8.5, from the above viewpoints.

  The liquid germicide composition preferably contains the content of dipicolinic acid in a solution containing dipicolinic acid or a salt thereof and a cationic surfactant, more preferably in an aqueous solution containing dipicolinic acid or a salt thereof and a cationic surfactant Is preferably 0.05 mM or more, more preferably 0.5 mM or more, more preferably 3 mM or more, more preferably 4 mM or more, more preferably 6 mM or more, more preferably 8 mM or more from the viewpoint of further improving the germicidal effect. And preferably 1 M or less, more preferably 200 mM or less, more preferably 100 mM or less, more preferably 50 mM or less, from the viewpoint of the stability of the solution containing dipicolinic acid or a salt thereof and a cationic surfactant. More preferably, it is 20 mM or less. In addition, the content of dipicolinic acid or a salt thereof in the liquid germicide composition is preferably from 0.05 mM to 1 M, more preferably from 0.05 mM to 200 mM, more preferably from the viewpoints described above. 3 mM to 100 mM, more preferably 3 mM to 15 mM, more preferably 4 mM to 15 mM.

  The liquid germicide composition is preferably of cationic surfactant in a solution comprising dipicolinic acid or a salt thereof and a cationic surfactant, more preferably in an aqueous solution comprising dipicolinic acid or a salt thereof and a cationic surfactant The content is preferably 0.05 mM or more, more preferably 0.5 mM or more, more preferably 3 mM or more, more preferably 4 mM or more, more preferably 6 mM or more, from the viewpoint of further improving the germicidal effect. It is 8 mM or more and preferably 1 M or less, more preferably 200 mM or less, more preferably 100 mM or less, more preferably 50 mM or less, more preferably 20 mM or less. In addition, the content of the cationic surfactant in the liquid germicidal aid composition is preferably 0.1 to 1 M, more preferably 0.5 to 500 mM, more preferably 3 to 3 from the above viewpoints. It is 300 mM, more preferably 4 to 100 mM, more preferably 6 to 100 mM, 8 to 100 mM.

  The content of the cationic surfactant in the liquid mashing aid composition is preferably 0.01% by mass or more, more preferably 0.05% by mass or more, from the viewpoint of further improving the germicidal effect. Preferably it is 10 mass% or less, More preferably, it is 5 mass% or less, More preferably, it is 1 mass% or less. In addition, the content of the cationic surfactant in the liquid slaughtering aid composition is preferably 0.01 to 10% by mass, more preferably 0.05 to 5% by mass, more preferably from the above viewpoints. Is 0.05 to 1% by mass.

  The composition of the present invention is a composition containing dipicolinic acid or a salt thereof and a cationic surfactant, water or other solvent, and containing neither peracetic acid, sulfoperoxycarboxylic acid nor dicarboxylic acid diester. possible. Here, “not included” indicates that the slaughtering aid composition does not have a killing effect, preferably 1000 ppm or less, more preferably 100 ppm or less, more preferably 10 ppm or less, more preferably 1 ppm or less, more preferably It is substantially 0 ppm. Here, "substantially" is below the detection limit.

  The liquid germicide composition of the present invention, preferably a solution containing dipicolinic acid or a salt thereof and a cationic surfactant, more preferably an aqueous solution containing dipicolinic acid or a salt thereof and a cationic surfactant, powdery dipicolin It can be prepared by mixing, preferably dissolving, the acid or its salt or cationic surfactant with water or another solvent.

  The solid germicide composition includes a composition comprising dipicolinic acid or a salt thereof and a cationic surfactant or dipicolinic acid or a salt thereof, a cationic surfactant, and a solidifying agent.

  The solidifying agent contained in the solid germinating agent composition includes, but is not limited to, polyethylene glycol having a number average molecular weight of 1,000 to 100,000; carnauba wax, candelilla wax, jojoba oil Waxes such as beeswax and lanolin; hydrocarbons having 15 or more carbon atoms such as paraffin, vaseline, ceresin and microcrystalline wax; higher fatty acids having 12 to 22 carbon atoms such as lauric acid, myristic acid and stearic acid; cetyl alcohol, C14-C22 higher alcohols, such as stearyl alcohol, are mentioned.

  The solid germicide composition can further contain a pH adjuster. As a pH adjuster contained in the solid slaughter aid composition, commonly used acids and bases, for example, inorganic acids such as hydrochloric acid and sulfuric acid, organic acids such as lactic acid and citric acid or their salts, sodium hydroxide And inorganic bases such as potassium hydroxide, and organic bases such as monoethanolamine, triethanolamine, triisopropanolamine and the like. As the pH adjuster, as the acid, an inorganic acid such as hydrochloric acid or sulfuric acid is preferable, and as the base, an inorganic base such as sodium hydroxide or potassium hydroxide is preferable.

  When the spore forming agent composition of the present invention is used in step 1 and step 2 of the spore forming method of the present invention, the content of dipicolinic acid in the solution containing the spore forming bacteria and the spore forming agent composition is From the viewpoint of further improving the germicidal effect, preferably 0.05 mM or more, more preferably 0.5 mM or more, more preferably 3 mM or more, more preferably 4 mM or more, more preferably 6 mM or more, more preferably 8 mM or more And from the viewpoint of the stability of the solution containing dipicolinic acid and the cationic surfactant, preferably 200 mM or less, more preferably 100 mM or less, more preferably 30 mM or less, more preferably 25 mM or less, more preferably 20 mM or less It is preferable to use it as it is. In addition, when the spore forming agent composition of the present invention is used in step 1 and step 2 of the spore-forming method of the present invention, it contains dipicolinic acid in a solution containing a spore forming bacterium and a spore forming agent composition. Preferably, the amount is preferably 0.05 to 200 mM, more preferably 0.5 to 100 mM, more preferably 3 to 30 mM, more preferably 3 to 25 mM, taking the above aspects into consideration.

  When used in step 1 and step 2 of the spore-forming method of the present invention, the liquid spore-accumulating agent composition contains the cationic surfactant content in a solution containing the spore-forming bacteria and the spore-forming agent composition Preferably, the content of the cationic surfactant in the solution containing dipicolinic acid and the cationic surfactant, more preferably in the aqueous solution containing the dipicolinic acid and the cationic surfactant, from the viewpoint of further improving the germicidal effect, It is preferably 0.1 mM or more, more preferably 0.5 mM or more, more preferably 3 mM or more, more preferably 4 mM or more, more preferably 6 mM or more, more preferably 8 mM or more, and preferably 1000 mM or less, more preferably Is 500 mM or less, more preferably 300 mM or less, more preferably 100 mM or less. In addition, the content of the cationic surfactant at the time of using the liquid germicide composition is preferably 0.1 to 1000 mM, more preferably 0.5 to 500 mM, more preferably from the above viewpoints. Is 3 to 300 mM, more preferably 4 to 100 mM, more preferably 6 to 100 mM.

  When used in step 1 and step 2 of the spore-forming method of the present invention, the liquid spore-accumulating agent composition contains the cationic surfactant content in a solution containing the spore-forming bacteria and the spore-forming agent composition However, from the viewpoint of further improving the germicidal effect, it is preferably 0.01% by mass or more, more preferably 0.05% by mass or more, and preferably 10% by mass or less, more preferably 5% by mass or less, more preferably Is 1% by mass or less. In addition, the content of the cationic surfactant at the time of using the liquid slaughtering aid composition is preferably 0.01 to 10% by mass, more preferably 0.05 to 5%, in consideration of the above aspects. %, More preferably 0.05 to 1% by mass.

  The dipicolinic acid or a salt thereof of the present invention can be used either synthetically or by fermentation using a method of extraction from a microorganism. As the cationic surfactant of the present invention, either commercially available one or synthetic one can be used.

  The method and the adjuvant composition of the present invention can be used for sterilization cleaning of food processing equipment, sterilization cleaning of clothes, and the like.

  With regard to the embodiments described above, the present invention discloses the following germicidal methods and germicidal adjuvant compositions.

<Item 1>
Killing method to perform the following step 1, step 2 and step 3:
Step 1: contacting the spore forming bacteria with dipicolinic acid or a salt thereof;
Step 2: a step of contacting the spore-forming bacteria with a cationic surfactant; and Step 3: a step of heating the spore-forming bacteria to 50 ° C. or higher (however, step 3 is terminated after starting step 1 and step 2) It shall be).

<Section 2>
The killing method according to <Item 1>, having a period in which Step 1, Step 2, and Step 3 are simultaneously performed.

<Section 3>
The killing method according to <Item 1>, wherein the steps 2 and 3 are performed after the end of the step 1.

<Section 4>
The killing method according to <Item 1>, wherein step 1 and step 3 are performed after the end of step 2.

<Section 5>
The cationic surfactant is any of a primary ammonium salt, a secondary ammonium salt, a tertiary ammonium salt, or a quaternary ammonium salt, according to any one of <Item 1> to <Item 4>. How to kill.

<Section 6>
The cationic surfactant is a quaternary ammonium salt such as alkyl trimethyl ammonium salt, dialkyl dimethyl ammonium salt, benzalkonium salt, alkyl pyridinium salt, alkyl benzethonium salt, or a primary ammonium salt such as alkyl amine salt The killing method according to any one of Items 1 to 5.

<Section 7>
Cationic surfactant is a quaternary ammonium salt represented by the general formula (1), <claim 1> to quit method according to any one of <6.>.

^Wherein any one of R 11 ^{to R 14,} a hydroxyl group, an ester group, an amide group hydrocarbon group having 3 or more carbon atoms which may have a ^remaining three of R 11 ^{to R 14} represents a methyl group or an ethyl group, X ^- represents an inorganic or organic anionic compounds. ]

<Section 8>
The cationic surfactant includes alkyl trimethyl ammonium salt, alkyl triethyl ammonium salt, alkyl dimethyl ethyl ammonium salt, alkyl methyl diethyl ammonium salt, dialkyl dimethyl ammonium salt, dialkyl diethyl ammonium salt, dialkyl ethyl methyl ammonium salt, benzalkonium salt, alkyl The killing method according to any one of <Item 1> to <Item 7>, which is at least one selected from the group consisting of pyridinium salts and alkyl benzethonium salts.

<Section 9>
The method according to any one of <1> to <8>, wherein the spore forming bacteria, dipicolinic acid or a salt thereof, and the cationic surfactant are brought into contact with each other in the steps 1 and 2.

<Section 10>
Item 10. The method according to Item 9, wherein the content of dipicolinic acid or a salt thereof in the solution is 0.05 mM or more and 200 mM or less.

<Item 11>
The killing method according to <Item 9>, wherein the content of the cationic surfactant in the solution is 0.1 mM or more and 1000 mM or less.

<Section 12>
The concentration of dipicolinic acid or its salt when contacting the spore forming bacteria with dipicolinic acid or its salt is 0.05 mM or more, more preferably 0.5 mM or more, more preferably 3 mM or more, more preferably 4 mM or more, more preferably Is 6 mM or more, more preferably 8 mM or more, and preferably 200 mM or less, more preferably 100 mM or less, more preferably 50 mM or less, more preferably 20 mM or less, according to any one of <Item 1> to <Item 11> How to kill.

<Section 13>
The concentration of the cationic surfactant when the spore-forming bacteria come into contact with the cationic surfactant is 0.1 mM or more, more preferably 0.5 mM or more, more preferably 3 mM or more, more preferably 4 mM or more, more preferably Any one of <Item 1> to <Item 12> having a concentration of 6 mM or more, more preferably 8 mM or more, and preferably 1000 mM or less, more preferably 500 mM or less, more preferably 300 mM or less, more preferably 100 mM or less Killing method described.

<Section 14>
The concentration of the cationic surfactant when the spore-forming bacteria come into contact with the cationic surfactant is 0.01% by mass or more, more preferably 0.05% by mass or more, and preferably 10% by mass or less, more preferably The slaughter method according to any one of <Item 1> to <Item 13>, wherein the content is 5% by mass or less, more preferably 1% by mass or less.

<Section 15>
The method according to any one of <Item 1> to <Item 14>, wherein Step 1 and Step 2 are performed independently or simultaneously at 15 ° C. or more and less than 50 ° C.

<Section 16>
Step 1 and Step 2, which are carried out independently of Step 3, are each independently or simultaneously at 15 ° C. or more, more preferably 20 ° C. or more, and preferably less than 50 ° C., more preferably 49 ° C. or less The killing method according to any one of <Item 1> to <Item 15>, more preferably at 45 ° C. or less, more preferably at 40 ° C. or less, more preferably at 30 ° C. or less.

<Section 17>
The time of step 1, which is performed independently of step 3, ie, the contact time of the spore forming bacteria with dipicolinic acid or a salt thereof, is preferably 1 second or more, more preferably 5 seconds or more, more preferably 30 seconds or more More preferably, it is 1 minute or more, more preferably 3 minutes or more, more preferably 5 minutes or more, more preferably 8 minutes or more, more preferably 10 minutes or more, more preferably 15 minutes or more, more preferably 20 minutes or more More preferably 25 minutes or more, and preferably 60 minutes or less, more preferably 50 minutes or less, more preferably 40 minutes or less, more preferably 35 minutes or less A method of killing according to any one.

<Section 18>
The time of step 2 when performed independently of step 3, ie, the contact time of the spore forming bacteria with the cationic surfactant, is preferably 1 second or more, more preferably 5 seconds or more, more preferably 30 seconds or more More preferably, it is 1 minute or more, more preferably 3 minutes or more, more preferably 5 minutes or more, more preferably 8 minutes or more, more preferably 10 minutes or more, more preferably 15 minutes or more, more preferably 20 minutes or more More preferably 25 minutes or more, and preferably 60 minutes or less, more preferably 50 minutes or less, more preferably 40 minutes or less, more preferably 35 minutes or less A method of killing according to any one.

<Item 19>
The total contact time for contacting the spore forming bacteria with dipicolinic acid or a salt thereof in Step 1 when having a period simultaneously performed with Step 3 is preferably 5 minutes or more, more preferably 10 minutes or more, more preferably 15 minutes More preferably, it is 20 minutes or more, more preferably 25 minutes or more, and preferably 90 minutes or less, more preferably 80 minutes or less, more preferably 70 minutes or less, more preferably 65 minutes or less, more preferably The killing method according to any one of <Item 1> to <Item 18>, wherein the time is 60 minutes or less, more preferably 50 minutes or less, more preferably 40 minutes or less, more preferably 35 minutes or less.

<Item 20>
The total contact time of the spore-forming bacteria and the cationic surfactant in step 2 when there is a period simultaneously performed with step 3 is preferably 5 minutes or more, more preferably 10 minutes or more, more preferably 15 minutes or more, more preferably Is 20 minutes or more, more preferably 25 minutes or more, and preferably 90 minutes or less, more preferably 80 minutes or less, more preferably 70 minutes or less, more preferably 65 minutes or less, more preferably 60 minutes or less, The killing method according to any one of <Item 1> to <Item 19>, more preferably 50 minutes or less, more preferably 40 minutes or less, more preferably 35 minutes or less.

<Section 21>
The killing method according to any one of <Item 1> to <Item 20>, wherein the time for heating the spore forming bacteria to 50 ° C. or more in Step 3 is 3 minutes or more and 90 minutes or less.

<Section 22>
The temperature of step 3 is preferably 50 ° C. or more, more preferably 55 ° C. or more, more preferably 60 ° C. or more, more preferably 65 ° C. or more, more preferably 70 ° C. or more, more preferably 75 ° C. or more Preferably it is 250 ° C. or less, more preferably 200 ° C. or less, more preferably 150 ° C. or less, more preferably 120 ° C. or less, more preferably 100 ° C. or less, more preferably 90 ° C. or less, more preferably 85 ° C. or less The killing method according to any one of <Item 1> to <Item 21>.

<Section 23>
The heating time in step 3 is preferably 3 minutes or more, more preferably 5 minutes or more, more preferably 8 minutes or more, more preferably 10 minutes or more, more preferably 20 minutes or more, more preferably 25 minutes or more And is preferably 90 minutes or less, more preferably 70 minutes or less, more preferably 50 minutes or less, more preferably 40 minutes or less, more preferably 35 minutes or less. A method of killing according to any one.

<Section 24>
The killing method according to any one of <Item 1> to <Item 23>, having a period in which the step 1 and the step 2 are performed independently or simultaneously at 15 ° C. or more and less than 50 ° C. before the step 3 starts.

<Section 25>
The killing method according to any one of <Item 1> to <Item 24>, wherein the step 1 and the step 2 are performed independently or simultaneously for 1 second to 60 minutes before the start of the step 3.

<Item 26>
The killing method according to any one of <Item 1> to <Item 25>, wherein the step 1 and the step 2 are performed independently or simultaneously for 30 minutes to 90 minutes in total.

<Section 27>
An adjuvant composition comprising dicpicolinic acid or a salt thereof and a cationic surfactant.

<Section 28>
Liquid composition containing dipicolinic acid or a salt thereof preferably containing 0.05 mM or more, more preferably 0.5 mM or more, more preferably 3 mM or more, and preferably 200 mM or less, more preferably 100 mM or less, more preferably 30 mM or less The slaughtering aid composition according to <Item 27>, wherein

<Section 29>
A liquid composition containing preferably 0.1 mM or more, more preferably 0.5 mM or more, more preferably 3 mM or more, and preferably 1000 mM or less, more preferably 500 mM or less, more preferably 300 mM or less of a cationic surfactant. The slaughter auxiliary agent composition as described in <Item 27> or <Item 28>.

<Section 30>
The content of peracetic acid, sulfoperoxycarboxylic acid and dicarboxylic acid diester is preferably 1000 ppm or less, more preferably 100 ppm or less, more preferably dipicolinic acid or a salt thereof, a cationic surfactant, and water or other solvent. The germicidal auxiliary composition according to any one of <Item 27> to <29>, which is 10 ppm or less, more preferably 1 ppm or less, more preferably substantially 0 ppm.

  Hereinafter, the present invention will be more specifically described with reference to examples.

Preparation of bacteria Spore-forming bacteria: Bacillus subtilis 168 strain was used as a test strain. An aqueous dispersion having a content of spore forming bacteria of 10 ⁸ CFU / mL was prepared and used for the experiment. The spore forming bacteria in this aqueous dispersion were used in the experiment after confirming that 95% or more formed spores by microscopic observation.

Preparation of aqueous solution of dipicolinic acid Dipicolinic acid (hereinafter, also referred to as DPA) was prepared using Tris-HCl buffer (Wako Pure Chemical Industries, Ltd., a reagent (manufactured by Wako Pure Chemical Industries, Ltd., manufactured code 165-05342) and solvent as ion exchange water of 50 mM. Industrial Co., Ltd., manufacture code 318-90225), and sodium hydroxide was used as the pH adjuster. Preparation of the dipicolinic acid aqueous solution was performed at 24 ° C. While the pH of the aqueous dipicolinic acid solution was measured with a table-top pH meter type 9611 (manufactured by HORIBA), a pH adjuster was added dropwise to the aqueous dipicolinic acid solution to adjust the pH of the aqueous dipicolinic acid solution. In the examples, unless stated otherwise, the pH is the pH at 24 ° C. The experiment was carried out by adjusting to pH 7 (24 ° C.) unless otherwise noted.

Surfactant The following surfactant was used.
Lauryl trimethyl ammonium chloride (Cortamine 24P, manufactured by Kao Corporation)
Alkyl benzyl dimethyl ammonium chloride (Sanisole C Kao Co., Ltd. product)
Sodium lauryl sulfate (Emar 0, manufactured by Kao Corporation)
Polyoxyethylene (3) sodium lauryl ether sulfate (Emar 20C, manufactured by Kao Corporation)
Polyoxyethylene (2) sodium lauryl ether sulfate (Emar E-27C, manufactured by Kao Corporation)
Sodium dodecyl benzene sulfonate ( Neoperex G-15, manufactured by Kao Corporation)
Polyoxyethylene lauryl ether (Emulgen 108, manufactured by Kao Corporation)
Polyoxyethylene (23) lauryl ether (Emulgen 123P, manufactured by Kao Corporation)
Polyoxyethylene lauryl ether (Emulgen 130K, manufactured by Kao Corporation)
Monolaurate polyethylene glycol (Emanon 1112, manufactured by Kao Corporation)
Polyoxyethylene hydrogenated castor oil (60E.O.) (Emanon CH-60, manufactured by Kao Corporation)
Lauryl glucoside (Mydol 12 manufactured by Kao Corporation)
Dodecyl trimethyl ammonium chloride (made by Tokyo Chemical Industry, hereinafter C12 trimethyl ammonium chloride)
Tetradecyl trimethyl ammonium chloride (made by Tokyo Chemical Industry, hereinafter C14 trimethyl ammonium chloride)
Hexadecyl trimethyl ammonium chloride (made by Tokyo Chemical Industry, hereinafter C16 trimethyl ammonium chloride)
Octadecyl trimethyl ammonium chloride (made by Tokyo Chemical Industry, hereinafter C18 trimethyl ammonium chloride)
Dioctadimethyl ammonium chloride (made by Tokyo Kasei, hereinafter C8 dimethyl ammonium chloride)
Didecyldimethyl ammonium chloride (made by Tokyo Chemical Industry, hereinafter C10 dimethyl ammonium chloride)
Didodecyl dimethyl ammonium chloride (made by Tokyo Chemical Industry, hereinafter C12 dimethyl ammonium chloride)
Ditetradecyldimethyl ammonium chloride (made by Tokyo Chemical Industry, hereinafter C14 dimethyl ammonium chloride)
Dihexadecyl dimethyl ammonium chloride (made by Tokyo Chemical Industry, hereinafter C16 dimethyl ammonium chloride)
Dioctadecyl dimethyl ammonium chloride (made by Tokyo Chemical Industry, hereinafter C18 dimethyl ammonium chloride)

Example 1-1:
10mM dipicolinic acid solution as described in stop test Table 1 (hereinafter, also referred to as DPA water solution), using a 0.10 wt% lauryl trimethyl ammonium chloride and spore-forming bacteria, subjected to quit tested by the following procedure , And evaluated the killing effect. The evaluation results are shown in Table 1. (In the table, "%" is "% by mass". The same shall apply hereinafter)

-procedure-

(1) Step 1 and Step 2: 100 μL of an aqueous dispersion of spore-forming bacteria (hereinafter referred to as spore solution; the same in the following examples), 800 μL of an aqueous solution of DPA and 100 μL of an aqueous solution of lauryltrimethylammonium chloride And used as a test solution. The final concentration of DPA was 10 mM, and the final concentration of lauryltrimethylammonium chloride was 0.1% by mass. Here, the final concentration means the concentration in the aqueous solution after mixing (in the state of 1 mL in total) (same in the following examples).
(2) Step 1 and Step 2: The test solution (pH 7.0) of the above (1) was allowed to stand at 24 ° C. for 30 minutes.
(3) Step 3: After the above (2), the test solution was heated as it is at 80 ° C. for 30 minutes.
The aluminum block thermostat MD-01N-110 of Major Science was used for the heating process.
(4) The test solution heat-treated in (3) is serially diluted with an LP aqueous solution (LP dilution solution (Digo, manufactured by Nippon Pharmaceutical Co., Ltd.)), and 100 μL of each dilution solution is smeared on LB agar medium (manufactured by BD) And cultured at 37 ° C. for 16 hours.
(5) After culture | cultivation of (4), the number X of viable bacteria X (CFU / mL) was calculated | required by the colony number which has grown.

Comparative Examples 1-1, 1-2
In the procedure (1) of Example 1-1, the aqueous solution shown in Table 1 was used instead of the aqueous solution containing a final concentration of 10 mM DPA and a final concentration of 0.1% by mass cationic surfactant. Other than that was carried out similarly to Example 1-1.

Example 2-1 and Comparative Examples 2-1 to 2-10:
The aqueous solution which mixed each surfactant aqueous solution shown in Table 2 in DPA aqueous solution was used. The final concentration of DPA in the aqueous solution was 10 mM, and the final concentration of each surfactant was 0.1% by mass. A slaughter test was conducted under the same conditions as in Example 1-1 to evaluate the slaughter effect. The evaluation results are shown in Table 2.

Examples 3-1 to 3-10:
The aqueous solution which mixed each surfactant aqueous solution shown in Table 3 in DPA aqueous solution was used. The final concentration of DPA and each surfactant in the aqueous solution was 10 mM. A slaughter test was conducted under the same conditions as in Example 1-1 to evaluate the slaughter effect. The evaluation results are shown in Table 3.

Comparative Examples 3-1 to 3-3:
The killing effect was evaluated under the same conditions as in Examples 3-1, 3-3, and 3-4 except that the temperature in step 3 was changed to RT (25 ° C.).

Examples 4-1 to 4-13:
The aqueous solution which mixed each surfactant aqueous solution shown in Table 4 in DPA aqueous solution was used. The final concentration of DPA in the aqueous solution was 10 mM, and the final concentration of each surfactant was the concentration shown in Table 4. A slaughter test was conducted under the same conditions as in Example 1-1 to evaluate the slaughter effect. The evaluation results are shown in Table 4.

Examples 5-1 to 5-3:
The aqueous solution which mixed C18 trimethyl ammonium chloride aqueous solution with DPA aqueous solution was used. The final concentration of DPA in the aqueous solution was 10 mM, and the final concentration of C18 trimethyl ammonium chloride was 20 mM. The heating temperature in step (3) was set to the temperature shown in Table 5. A slaughter test was conducted under the same conditions as in Example 1-1 to evaluate the slaughter effect. The evaluation results are shown in Table 5.

Examples 6-1 to 6-4
The aqueous solution which mixed C18 trimethyl ammonium chloride aqueous solution with DPA aqueous solution was used. The final concentration of DPA in the aqueous solution was 10 mM, and the final concentration of C18 trimethyl ammonium chloride was 1 mM. The pH (24 ° C.) of the test solution in procedures (1) and (2) was set to the value shown in Table 6. A slaughter test was conducted under the same conditions as in Example 1-1 to evaluate the slaughter effect. The evaluation results are shown in Table 6. Example 6-2 is a buffer system of phthalic acid and sodium hydroxide, Example 6-3 is a buffer system of boric acid and sodium hydroxide, and Example 6-4 is a buffer system of potassium chloride and sodium hydroxide. Using.

Example 7-1
-procedure-
(1) Step 1: 100 μL of spore fluid was mixed with 900 μL of DPA aqueous solution to prepare a test solution (pH 7.0). The final concentration of DPA was 10 mM.
(2) The test solution was allowed to stand at 24 ° C. for 30 minutes.
(3) The DPA solution was removed from the spore by the following method. The test solution in step 1 was centrifuged using a centrifuge (tabletop micro refrigerated centrifuge 3500, manufactured by Kubota Corporation, the same in the following) under the conditions of 15 krpm for 5 minutes, and the supernatant was removed.
(4) Step 2: The supernatant was removed, and immediately, 1 mL of C16 trimethyl ammonium chloride aqueous solution was added to a final concentration of 10 mM, and left at 24 ° C. for 30 minutes.
(5) Step 3: After the above (4), the test solution was heated at 80 ° C. for 30 minutes.
(6) The test solution of (5) was serially diluted with sterile water, and 100 μL of each diluted solution was smeared on LB agar medium (manufactured by BD) and cultured at 37 ° C. for 16 hours.
(7) After the culture of (6), the number of viable bacteria X (CFU / mL) was determined from the number of grown colonies.

Example 7-2
-procedure-
(1) Step 2: 100 μL of spore solution was mixed with 900 μL of C16 trimethyl ammonium chloride aqueous solution to prepare a test solution (pH 7.0). The final concentration of C16 trimethyl ammonium chloride was 10 mM.
(2) The test solution was allowed to stand at 24 ° C. for 30 minutes.
(3) The C16 trimethyl ammonium chloride solution was removed from the spores by the following method. The test solution (2) was centrifuged using a centrifuge under the condition of 15 krpm for 5 minutes, and the supernatant was removed.
(4) Step 1: After removing the supernatant, 1 mL of DPA aqueous solution was immediately added to a final concentration of 10 mM, and left at 24 ° C. for 30 minutes.
(5) Step 3: After the above (4), the test solution was heated at 80 ° C. for 30 minutes.
(6) The test solution of (5) was serially diluted with sterile water, and 100 μL of each diluted solution was smeared on LB agar medium (manufactured by BD) and cultured at 37 ° C. for 16 hours.
(7) After the culture of (6), the number of viable bacteria X (CFU / mL) was determined from the number of grown colonies.

Example 7-3
-procedure-
(1) Step 1 and Step 2: 100 μL of spore fluid was mixed with DPA and 900 μL of an aqueous solution of C16 trimethyl ammonium chloride to prepare a test solution (pH 7.0). The final concentration of DPA and C16 trimethyl ammonium chloride was 10 mM.
(2) Step 3: Immediately after (1), the test solution was heated at 80 ° C. for 30 minutes.
(3) The test solution of (2) was serially diluted with sterile water, and 100 μL of each diluted solution was smeared on LB agar medium (manufactured by BD) and cultured at 37 ° C. for 16 hours.
(4) After the culture of (3), the number of viable bacteria X (CFU / mL) was determined from the number of grown colonies.

Comparative example 7-1
-procedure-
(1) 100 μL of spore solution was mixed with 900 μL of DPA solution to prepare a test solution (pH 7.0). The final concentration of DPA was 10 mM.
(2) Immediately after (1), the test solution was heated at 80 ° C. for 30 minutes.
(3) by the following method to remove DPA water solution from spores.
The test solution (2) was centrifuged using a centrifuge under the condition of 15 krpm for 5 minutes, and the supernatant was removed.
(4) The supernatant was removed, and immediately, 1 mL of a C16 trimethyl ammonium chloride aqueous solution was added to a final concentration of 10 mM, and allowed to stand at 24 ° C. for 30 minutes.
(5) The test solution of (4) was serially diluted with sterile water, and 100 μL of each diluted solution was smeared on LB agar medium (manufactured by BD) and cultured at 37 ° C. for 16 hours.
(6) After culture | cultivation of (5), the number X of viable bacteria (CFU / mL) was calculated | required by the colony number which has grown.

Comparative Example 7-2
-procedure-
(1) Step 3: 100 μL of spore fluid was heated at 80 ° C. for 30 minutes.
(2) Step 2: After (1), immediately after cooling to room temperature, it was mixed with 900 μL of a C16 trimethyl ammonium chloride aqueous solution and allowed to stand for 30 minutes to obtain a test solution (pH 7.0). The final concentration of C16 trimethyl ammonium chloride was 10 mM.
(3) The test solution of (2) was serially diluted with sterile water, and 100 μL of each diluted solution was smeared on LB agar medium (manufactured by BD) and cultured at 37 ° C. for 16 hours.
(4) After the culture of (3), the number of viable bacteria X (CFU / mL) was determined from the number of grown colonies.

Example 8-1:
A slaughter test was conducted in the same manner as in Example 3-1.

Comparative Example 8-1:
An aqueous solution of isophthalic acid having the same concentration was used instead of the aqueous DPA solution. Other than that was carried out similarly to Example 8-1.

Example 9-1
(1) Step 1 and Step 2: 100 μL of the spore solution was mixed with 800 μL of an aqueous solution of DPA and 100 μL of an aqueous solution of C16 trimethyl ammonium chloride to prepare a test solution (pH 7.0). The final concentration of DPA and C16 trimethyl ammonium chloride was 10 mM.
(2) Step 3: Immediately after the above (1), the test solution was heated at 80 ° C. for 30 minutes as it was. The aluminum block thermostat MD-01N-110 of Major Science was used for the heating process.
(3) The test solution heat-treated in (2) above is serially diluted with an LP aqueous solution (LP dilution solution (Digo, manufactured by Nippon Pharmaceutical Co., Ltd.)), and each dilution solution is diluted to 100 μL in LB agar medium (manufactured by BD) The cells were smeared and cultured at 37 ° C. for 16 hours.
(4) After the culture of (3), the number of viable bacteria X (CFU / mL) was determined from the number of grown colonies.

Comparative Example 9-1
The operation was carried out in the same manner as in Example 9-1, but without warming in step 3, it was allowed to stand at room temperature for 30 minutes.

Comparative Example 9-2
In place of the DPA and cationic surfactant aqueous solution of (1) step 1 and step 2 of Example 9-1, the spore solution was contacted with purified water. The heating in step 3 was performed in the same manner as in Example 9-1.

  The results of Example 9-1 and Comparative Examples 9-1 to 9-2 are shown in Table 9.

Examples 10-1 to 10-2
The aqueous solution which mixed C16 trimethyl ammonium chloride aqueous solution with DPA aqueous solution was used. The final concentration of DPA was as shown in Table 10, and the final concentration of C16 trimethyl ammonium chloride was 10 mM. A slaughter test was conducted under the same conditions as in Example 1-1 to evaluate the slaughter effect. The evaluation results are shown in Table 10.

  The slaughter method of the present invention can be used in a wide range of applications such as, for example, linen washing, food rot prevention and environmental purification.

Claims (10)

A killing method comprising the following step 1, step 2 and step 3:
Step 1: contacting the spore forming bacteria with dipicolinic acid or a salt thereof;
Step 2: A step of bringing a spore forming bacterium into contact with a cationic surfactant, wherein the cationic surfactant is an alkyltrimethyl ammonium salt represented by the general formula (1)

^Wherein any one of R 11 ^{to R 14,} a hydroxyl group, an ester group, an amide group hydrocarbon group having 3 or more carbon atoms which may have a ^remaining three of R 11 ^{to R 14} It represents a methyl group, X ^- represents an inorganic or organic anionic compounds. ]; And step 3: the spore-forming bacteria above 50 ° C., 3 minutes or more and 90 minutes or less step of heating (however, and ends the process 3 after starting the steps 1 and 2).   The method of killing according to claim 1, comprising a period in which the step 1, step 2 and step 3 are performed simultaneously. The molar ratio of the said dipicolinic acid with respect to the said cationic surfactant is 1/20 or more and 1000 or less by [molar concentration of dipicolinic acid] / [molar concentration of cationic surfactant]. How to kill. The method according to any one of claims 1 to 3 , wherein the spore forming bacteria, dipicolinic acid or a salt thereof, and a cationic surfactant are brought into contact with each other in the steps 1 and 2. The method according to claim 4 , wherein the content of dipicolinic acid or a salt thereof in the solution is 0.05 mM or more and 200 mM or less. The method according to claim 4 or 5 , wherein the content of the cationic surfactant in the solution is 0.1 mM or more and 1000 mM or less. A blasting aid composition comprising dipicolinic acid or a salt thereof and a cationic surfactant , wherein the cationic surfactant is an alkyltrimethyl ammonium salt represented by the general formula (1) Agent composition:

^Wherein any one of R 11 ^{to R 14,} a hydroxyl group, an ester group, an amide group hydrocarbon group having 3 or more carbon atoms which may have a ^remaining three of R 11 ^{to R 14} It represents a methyl group, X ^- represents an inorganic or organic anionic compounds. ] . The composition according to claim 7 , which is a liquid composition containing dipicolinic acid or a salt thereof of 0.05 mM or more and 200 mM or less. The composition according to claim 7 or 8 , which is a liquid composition containing 0.1 mM to 1000 mM of the cationic surfactant. 10. The method according to any one of claims 7 to 9, wherein a molar ratio of the dipicolinic acid to the cationic surfactant is [molar concentration of dipicolinic acid] / [molar concentration of cationic surfactant], 1/20 to 1000. The slaughtering aid composition according to item 1.