Method for promoting peracetic acid decomposition using metal compound, and method for culturing microorganism using same

Abstract

The present invention relates to a method for decomposing peracetic acid using a metal compound and a method for culturing a microorganism using the same, and more particularly, to a method for decomposing peracetic acid using a metal compound and a method for culturing a microorganism using the same, whereby a microorganism can be cultured in a sterilized culture medium by significantly rapidly decomposing peracetic acid when using the method of the present invention.

Description

Technical Field

[0001] The present invention relates to a peracetic acid decomposition method using a metal compound and a microorganism cultivation method using the above method. Background Art

[0002] In the bio-industry, sterilization of culture media and reactors is very important when mass-producing microbial raw materials and useful metabolites derived from microorganisms.

[0003] Therefore, conventionally, an autoclave, which is a sterilization method using high temperature and high pressure, has been mainly used to sterilize a culture medium and a reactor for culturing microorganisms.

[0004] However, when a high-pressure sterilizer is used to sterilize a culture medium and a reactor, the sugar component of the culture medium reacts with a protein (peptone or yeast extract) which is one of the other components of the culture medium at a high temperature, inducing the production of toxic substances such as hydroxymethyl furfural (HMF) that inhibit the growth of microorganisms, thereby posing a problem of inhibiting the growth of microorganisms. In addition, the material of the microbial reactor has a limiting point that it needs to be made of a material that is resistant to heat and pressure, such as metal or glass.

[0005] In order to overcome the above problems and limitations, a filtration method is used. However, this method has the inconvenience of sterilizing the culture vessel itself separately during filtration, and there is a high risk of clogging and re-contamination during the filtration process. In addition, the size of the filter is limited and the cost is high, resulting in a problem of low industrial applicability.

[0006] In order to solve this problem, a sterilization method using peracetic acid (PAA) has been developed as disclosed in Korean Patent Publication No. 10-2015-0097295. Korean Patent Publication No. 10-2015-0097295 discloses a method for sterilizing a photobioreactor using peracetic acid when pure culture of microalgae that requires light is carried out. However, the above-mentioned prior art documents do not disclose a method for sterilizing a culture medium containing sugar and a nitrogen source (peptone, yeast extract, whey, etc.). In addition, in the method of sterilizing a photobioreactor and a culture medium using peracetic acid, when the culture medium components include sugar and a nitrogen source, a high concentration of peracetic acid is required to achieve complete sterilization. In particular, when a culture medium to which a nitrogen source is added is sterilized together, nitrogen oxides (NO3-) are generated due to peracetic acid, and there is a problem that the generated nitrogen oxides bind to iron ions and inhibit the catalytic reaction for decomposing peracetic acid. In this case, it takes more than one month to decompose the peracetic acid reactant only by the natural decomposition of peracetic acid and the catalytic reaction based on iron and HEPES.

[0007] On the other hand, as disclosed in Korean Patent Publication No. 10-2020-0083909, when sterilization is performed by adding iron ions, alkali metal hydroxide, EDTA and sugar to a peracetic acid mixture and reacting them to decompose the peracetic acid mixture into acetic acid, water and oxygen, there are technical limitations in that the decomposition of peracetic acid takes a long time and the acetic acid as a decomposition product inhibits or kills some microorganisms.

[0008] In view of this, the present inventors have developed a method for promoting the decomposition of peracetic acid using a metal compound and a method for culturing microorganisms using the same in order to solve the problems of the prior art, thereby completing the present invention. Summary of the invention

[0009] Technical issues

[0010] The object of the present invention is to provide a method for decomposing peracetic acid, comprising: a step of mixing peracetic acid with water; and a step of adding iron ions, an alkali metal hydroxide, ethylenediaminetetraacetic acid (EDTA), sugar and a metal compound to the mixture.

[0011] Another object of the present invention is to provide a method for sterilizing a culture medium, comprising: a step of adding peracetic acid to a culture medium containing sugar and a nitrogen source and sterilizing the culture medium; and a step of adding iron ions, alkali metal hydroxides, ethylenediaminetetraacetic acid (EDTA) and a metal compound as a peracetic acid decomposition catalyst to the sterilized culture medium.

[0012] Another object of the present invention is to provide a method for culturing microorganisms, comprising: adding a culture medium sterilized by the above method to a bioreactor; and inoculating the microorganisms into the added culture medium and culturing them in pure form.

[0013] Technical Solution

[0014] In order to achieve the above object, the present invention provides a method for decomposing peracetic acid, which comprises: a step of mixing peracetic acid with water; and a step of adding iron ions, alkali metal hydroxide, ethylenediaminetetraacetic acid (EDTA), sugar and a metal compound to the mixture.

[0015] Furthermore, the present invention provides a method for sterilizing a culture medium, comprising: a step of adding peracetic acid to a culture medium comprising sugar and a nitrogen source and sterilizing the culture medium; and a step of adding iron ions, alkali metal hydroxides, ethylenediaminetetraacetic acid (EDTA) and metal compounds as peracetic acid decomposition catalysts to the sterilized culture medium.

[0016] Furthermore, the present invention provides a method for culturing microorganisms, comprising: adding a culture medium sterilized by the above method to a bioreactor; and inoculating the microorganisms into the added culture medium and culturing them in pure form.

[0017] Effects of the Invention

[0018] The present invention relates to a method for decomposing peracetic acid using a metal compound and a method for culturing microorganisms using the method. When the method of the present invention is used, peracetic acid present in a culture medium can be decomposed at a significantly rapid rate within a few minutes, thereby culturing microorganisms in a sterilized culture medium. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 This is a diagram showing the growth state of Euglena gracilis at different concentrations of acetic acid added to the culture medium. The culture conditions of each culture bottle correspond to acetic acid concentrations of 0%, 0.05%, 0.15%, 0.1%, 0.2%, and 0.25% (w / v) from the left to the right (reference point and comparative examples 2 to 6).

[0020] Figure 2 This is a diagram showing the growth state of Euglena gracilis at different concentrations of sodium acetate added to the culture medium. The culture conditions of each culture bottle correspond to sodium acetate concentrations of 0%, 0.05%, 0.1%, 0.15%, 0.2%, and 0.25% (w / v) from the left to the right (reference point and Examples 11 to 15).

[0021] Figure 3 This is a graph showing the growth state of Euglena gracilis at different concentrations of acetic acid added to the culture medium (Comparative Examples 2 to 6) and the growth state of species of Euglena gracilis at different concentrations of sodium acetate added to the culture medium (Examples 11 to 15). The Y axis represents absorbance, SA represents sodium acetate, and SS represents acetic acid.

[0022] Figure 4 The growth states of species of Euglena gracilis were observed at different concentrations of sodium bicarbonate added to the culture medium (Examples 16 to 21 and Comparative Example 7), and the result values ​​in Table 5 are shown in a graph. DETAILED DESCRIPTION

[0023] Hereinafter, the present invention will be described in detail by referring to the accompanying drawings and by examples of the present invention. However, the following implementation examples are disclosed by way of examples of the present invention, and when it is determined that the detailed description of a well-known technology or structure known to ordinary technicians unnecessarily confuses the main purpose of the present invention, its detailed description may be omitted, and thus the present invention is not limited thereto. The present invention can be deformed and applied within the description of the claims described below and the equivalent scope interpreted therefrom.

[0024] Throughout the specification, when certain parts are "connected (connected, in contact with, combined)" with other parts, this includes not only the case of "direct connection" but also the case of "indirect connection" with other parts in between. Furthermore, when certain parts "include" certain structural elements, this means that other structural elements may also be included, rather than excluding other structural elements, unless there is a special record to the contrary.

[0025] The terms used in this specification are only used to illustrate specific embodiments and are not used to limit the present invention. Singular expressions include plural expressions as long as there is no clear difference in the context. In this specification, it should be understood that the terms "including" or "having" are used to refer to the existence of features, numbers, steps, actions, structural elements, accessories or combinations of these recorded in the specification, rather than excluding the existence or addibility of one or more other features or numbers, steps, actions, structural elements, parts or combinations of these in advance.

[0026] Furthermore, the terminology used in this specification is used to appropriately describe the preferred embodiments of the present invention, which may vary according to the intention of the user, the operator, or the conventions of the field to which the present invention belongs. Therefore, this terminology should be defined based on the content of the full text of this specification. In the full text of the specification, when some parts "include" certain structural elements, this means that other structural elements may also be included, rather than excluding other structural elements, as long as there is no special record to the contrary.

[0027] The term "sterilization" as used in the present invention means killing vegetative cells and spores of microorganisms.

[0028] The term "metal compound" used in the present invention may mean a compound including metal ions such as sodium, calcium, potassium, etc., which can be combined with acetic acid.

[0029] The term “alkali metal” used in the present invention may mean an element belonging to Group 1 in the periodic table, and specifically may include lithium (Li), sodium (Na), and potassium (K) except hydrogen.

[0030] The term "peracetic acid mixture" used in the present invention may mean a mixture generated by mixing peracetic acid with water. In the present invention, the peracetic acid mixture may include peracetic acid, acetic acid and hydrogen peroxide.

[0031] The term "peracetic acid reactant" used in the present invention may mean that iron ions, alkali metal hydroxide, EDTA (Ethylene diamine tetraacetic acid), sugar and metal compounds are added to the above-mentioned peracetic acid mixture. In the present invention, the peracetic acid reactant may include hydrogen peroxide, acetic acid or sodium acetate (CH3COONa), etc. On the other hand, the above-mentioned peracetic acid reactant may include sodium acetate (CH3COONa), calcium acetate ((CH3COO)2Ca) or potassium acetate (CH3COOK) according to the added metal compound, but is not limited thereto.

[0032] The term "culture medium" used in the present invention is also referred to as a culture vessel or culture solution, which is a substance developed for the purpose of proliferation or cultivation of microorganisms to maintain a nutritional state similar to the natural environment in which microorganisms usually live and reproduce, and means a liquid or solid substance made by adding all nutrients required for growth. The culture of microorganisms in the culture medium varies depending on the selection of essential nutrients, including oxygen or gases required for culture, water, pH and temperature, and important nutrients may include carbon, nitrogen, inorganic phosphates and sulfur, trace metals, water and vitamins.

[0033] As an embodiment for achieving the above technical problem, the present invention provides a method for decomposing peracetic acid, characterized in that it includes: a step of mixing peracetic acid with water; and a step of adding iron ions, alkali metal hydroxide, ethylenediaminetetraacetic acid (EDTA), sugar and a metal compound to the mixture, wherein the metal compound includes any one or more selected from the group consisting of sodium bicarbonate (NaHCO3), disodium hydrogen phosphate (Na2HPO4), calcium carbonate (CaCO3), calcium lactate (Ca(CH3CHOHCOO)2·5H2O) and dipotassium hydrogen phosphate (K2HPO4).

[0034] In the above-mentioned peracetic acid decomposition method of the present invention, the above-mentioned iron ion can be Fe 2+ Ion or Fe 3+ Ions, the above-mentioned iron ions can be derived from salts containing Fe ions such as FeCl3, FeCl2 and / or FeSO4.

[0035] The alkali metal hydroxide may be LiOH, NaOH or KOH, but is not limited thereto.

[0036] The sugar may be dextrose, glucose, sucrose and / or molasses, but is not limited thereto.

[0037] The present invention is characterized in that the above-mentioned metal compound includes any one or more selected from the group consisting of sodium bicarbonate (NaHCO3), disodium hydrogen phosphate (Na2HPO4), calcium carbonate (CaCO3), calcium lactate (Ca(CH3CHOHCOO)2·5H2O) and dipotassium hydrogen phosphate (K2HPO4).

[0038] The present invention is characterized in that the concentration of the above-mentioned metal compound can be above 0.005% (w / v), preferably above 0.025% (w / v), more preferably above 0.05% (w / v), and most preferably above 0.08% (w / v).

[0039] The present invention is characterized in that the above-mentioned peracetic acid decomposition method of the present invention can significantly promote the overall peracetic acid decomposition reaction by removing acetic acid that hinders the growth of microorganisms included in the peracetic acid reactant. In one embodiment of the present invention, in order to remove acetic acid included in the peracetic acid reactant, any one or more selected from the group consisting of sodium bicarbonate (NaHCO3), disodium hydrogen phosphate (Na2HPO4), calcium carbonate (CaCO3), calcium lactate (Ca(CH3CHOHCOO)2·5H2O) and dipotassium hydrogen phosphate (K2HPO4) are used, but the metal compound may include a compound formed by ions that can be combined with acetic acid, such as sodium, potassium and / or calcium, and the scope of the rights is not limited thereto.

[0040] In the step of mixing peracetic acid with water in the above-mentioned method for decomposing peracetic acid of the present invention, peracetic acid and water may be mixed at room temperature to generate a peracetic acid mixture, and the peracetic acid mixture may include hydrogen peroxide and acetic acid.

[0041] 1) CH3CO3H (peracetic acid) + H2O → H2O2 (hydrogen peroxide) + CH3CO2H (acetic acid)

[0042] On the other hand, hydrogen peroxide generated by the reaction of peracetic acid and water undergoes Fenton reaction with iron ions and is decomposed into acetic acid, water and oxygen as shown in the following reaction formula.

[0043] 2) 2CH3CO3H → 2CH3CO2H + O2

[0044] 3) 2H2O2→2H2O+O2

[0045] In this case, hydrogen peroxide can be used as an oxidizing agent, a bleaching agent, and a disinfectant, and can be used for sterilizing a culture medium in relation to the present invention.

[0046] In the above-mentioned method for decomposing peracetic acid of the present invention, the step of adding iron ions, alkali metal hydroxide, EDTA (Ethylene diamine tetraacetic acid), sugar and metal compound to the above-mentioned peracetic acid mixture can be performed at room temperature, thereby generating a peracetic acid reactant, and the above-mentioned peracetic acid reactant can include hydrogen peroxide, acetic acid or sodium acetate (CH3COONa), etc. On the other hand, the above-mentioned peracetic acid reactant can include sodium acetate (CH3COONa), calcium acetate ((CH3COO)2Ca) or potassium acetate (CH3COOK) according to the added metal compound, but is not limited thereto.

[0047] The following example shows that sodium acetate (CH3COONa) is produced by adding sodium bicarbonate (NaHCO3) to acetic acid (CH3COOH).

[0048] 4-1)CH3COOH+NaHCO3→CH3COONa+CO2+H2O

[0049] 4-2)2CH3COOH+Na2HPO4→2CH3COONa+H3PO4

[0050] The following example shows that calcium carbonate (CaCO3) is added to acetic acid (CH3COOH) to produce calcium acetate ((CH3COO)2Ca).

[0051] 4-3)2CH3COOH+CaCO3→(CH3COO)2Ca+CO2+H2O

[0052] 4-4)2CH3COOH+Ca(CH3CHOHCOO)2·5H2O+6O2→(CH3COO)2Ca+6CO2+6H2O

[0053] The following example shows that sodium acetate is produced by adding sodium bicarbonate to acetic acid (CH3COOH).

[0054] 4-5)CH3COOH+KH2PO4→CH3COOK+H2PO4 -

[0055] 4-6)2CH3COOH+K2HPO4→2CH3COOK+H3PO4

[0056] In the above-mentioned peracetic acid decomposition method of the present invention, an alkaline buffer solution may be used instead of the alkali metal hydroxide, or an alkaline buffer solution may be added. The alkaline buffer solution may be 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), but is not limited thereto.

[0057] As another embodiment, the present invention provides a method for sterilizing a culture medium, characterized in that it includes: a step of adding peracetic acid to a culture medium including a sugar and a nitrogen source and sterilizing it; and a step of adding iron ions, alkali metal hydroxides, EDTA (Ethylene diamine tetraacetic acid) and metal compounds as peracetic acid decomposition catalysts to the sterilized culture medium, wherein the metal compound includes any one or more selected from the group consisting of sodium bicarbonate (NaHCO3), disodium hydrogen phosphate (Na2HPO4), calcium carbonate (CaCO3), calcium lactate (Ca(CH3CHOHCOO)2·5H2O) and dipotassium hydrogen phosphate (K2HPO4).

[0058] In the method for sterilizing a culture medium of the present invention, specific details of the peracetic acid decomposition method can be appropriately applied without departing from the essence of the present invention.

[0059] Peracetic acid, peracetic acid mixture, and peracetic acid reaction product present in a medium sterilized by the medium sterilization method of the present invention can be decomposed, and the medium thus prepared can be used for microbial culture.

[0060] In the sterilization method of the culture medium of the present invention, the iron ion may be Fe 2+ Ion or Fe 3+ Ions, the above-mentioned iron ions can be derived from salts containing Fe ions such as FeCl3, FeCl2 and / or FeSO4.

[0061] In the above-mentioned method for sterilizing a culture medium of the present invention, the hydroxide of the alkali metal may be LiOH, NaOH or KOH, but is not limited thereto.

[0062] In the method for sterilizing the culture medium of the present invention, the sugar may be dextrose, glucose, sucrose and / or molasses, but is not limited thereto.

[0063] In the sterilization method of the above-mentioned culture medium of the present invention, it is characterized in that the concentration of the above-mentioned metal compound can be above 0.005% (w / v), preferably above 0.025% (w / v), more preferably above 0.05% (w / v), most preferably above 0.08% (w / v).

[0064] In the sterilization method of the culture medium of the present invention, an alkaline buffer may be used instead of the alkali metal hydroxide, or an alkaline buffer may be added. The alkaline buffer may be 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES: 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid).

[0065] As described in the above method of the present invention, if the medium is sterilized, there is an advantage that the medium can be stored for a long period of time without being contaminated even if high pressure sterilization is not performed before neutralization and the medium is placed in an open container.

[0066] As another embodiment, the present invention provides a method for culturing microorganisms, comprising: adding a culture medium sterilized by the above method to a bioreactor; and inoculating the added culture medium with microorganisms and culturing the culture medium in a pure manner.

[0067] In the method for culturing a microorganism of the present invention, the specific details of the method for sterilizing the culture medium described above can be appropriately applied without departing from the essence of the present invention.

[0068] In the microorganism culture method of the present invention, the above-mentioned microorganism can be Aurantiochytrium sp., Schizochytrium sp., Chlorella sp., Synechocystis sp., Debaryomyces sp., yeasts, lactic acid bacteria, Actinomycetes, Euglena, Mortierella, filamentous fungi and / or photosynthetic bacteria, but is not limited thereto.

[0069] The microorganism inoculation may mean an act of creating an environment in which the microorganisms can grow in order to cultivate the microorganisms, which may include streak smear method, plate smear method and pour plate method, but is not limited thereto.

[0070] The pure culture means a process of culturing microorganisms etc. in a state where only a single species exists in a medium. In this case, a method of confirming whether a pure culture is formed by confirming whether a single colony is formed by a streak smear method can be used.

[0071] According to one embodiment of the present invention, it can be confirmed that when sodium bicarbonate is added to the peracetic acid mixture, the decomposition rate of peracetic acid is significantly increased, and it can be confirmed that the decomposition rate of acetic acid gradually increases with the increase in the added concentration of sodium bicarbonate (Examples 1 to 6 and Comparative Example 1).

[0072] Hereinafter, the present invention will be described in detail by way of examples.

[0073] However, the following examples are for illustrating the present invention, and the contents of the present invention are not limited to the examples.

[0074] Example 1. Confirmation of promotion of decomposition of peracetic acid by sodium bicarbonate

[0075] A microbial culture medium was prepared by adding 5 g / L of yeast extract and 20 g / L of dextrose to 1 L of distilled water, and then peracetic acid (Dong Myung ONC, 160126) was added to a concentration of 0.05% (v / v).

[0076] After adding 96 μM FeCl 3 (Sigma, 7705-08-0), 5.5 mM NaOH (Sigma, 1310-73-2) and 96 μM EDTA (Sigma, 60-00-4) to the above solution, sodium bicarbonate (NaHCO 3 , SAMCHUN PURE CHEMICAL, S0343) was added to make its concentration reach 0.05% (w / v).

[0077] The degree of decomposition of peracetic acid was measured using a peroxide test (MQuant 1100810001) that can measure peroxide, which is a decomposition product of a peracetic acid solution.

[0078] Comparative Example 1: No sodium bicarbonate added

[0079] The experiment was performed in the same manner as in Example 1, but without the addition of sodium bicarbonate.

[0080] Examples 2 to 6. Sodium bicarbonate concentration difference

[0081] The experiment was performed in the same manner as in Example 1, specifically, sodium bicarbonate was added to a concentration of 0.1% (w / v) (Example 2), 0.15% (w / v) (Example 3), 0.2% (w / v) (Example 4), 0.25% (w / v) (Example 5), and 0.3% (w / v) (Example 6).

[0082] Example 7. Confirmation of promotion of decomposition of peracetic acid by sodium dihydrogen phosphate

[0083] The experiment was performed in the same manner as in Example 1, except that disodium hydrogen phosphate (Na2HPO4, SAMCHUN PURE CHEMICAL, S0893) was added at a concentration of 0.05% (w / v) instead of sodium bicarbonate.

[0084] Example 8. Confirmation of the promotion of peracetic acid decomposition by calcium carbonate

[0085] The experiment was performed in the same manner as in Example 1, except that calcium carbonate (CaCO3, SAMCHUN PURE CHEMICAL, S0076) was added at a concentration of 0.05% (w / v) instead of sodium bicarbonate.

[0086] Example 9. Confirmation of promotion of peracetic acid decomposition by calcium lactate

[0087] The experiment was performed in the same manner as in Example 1, except that calcium lactate (Ca(CH3CHOHCOO)2·5H2O, DAEJUNG CHEMICAL & METALS, 2513-4405) was added at a concentration of 0.05% (w / v) instead of sodium bicarbonate.

[0088] Example 10. Confirmation of promotion of decomposition of peracetic acid by dipotassium hydrogen phosphate

[0089] The experiment was performed in the same manner as in Example 1, except that dipotassium hydrogen phosphate (K2HPO4, SAMCHUN PURE CHEMICAL, P1098) was added at a concentration of 0.05% (w / v) instead of sodium bicarbonate.

[0090] Example 11. Confirmation of whether sodium acetate inhibits microbial growth

[0091] In order to confirm whether sodium acetate, which is a decomposition product of peracetic acid, inhibits the growth of microorganisms, the following experiment was performed.

[0092] The nutrient medium for culturing Euglena gracilis (containing grade 3 distilled water, KH2PO4, citrate, MgSO4·3H2O, vitamin B1, vitamin B6, vitamin B12, vitamin H, yeast extract, peptone and dextrose) was sterilized by autoclaving at 121°C for 15 minutes and kept ready for use. After adding 0.05% (w / v) of sodium acetate (CH3COONa) to the prepared nutrient medium, the Euglena gracilis strain (KCTC AG40099) was inoculated and stirred at 130 rpm at 28°C and 100 μE photons / m 2The cells were cultured under the conditions of 100 μg / mL / s and the growth curve was determined by the absorbance at 680 nm.

[0093] Examples 12 to 15. Difference in concentration of sodium acetate

[0094] The experiment was performed in the same manner as in Example 11, with sodium acetate added at concentrations of 0.1% (w / v) (Example 12), 0.15% (w / v) (Example 13), 0.2% (w / v) (Example 14), and 0.25% (w / v) (Example 15).

[0095] Comparative Example 2: Confirmation of whether acetic acid inhibits microbial growth

[0096] The experiment was performed in the same manner as in Example 11, adding acetic acid instead of sodium acetate.

[0097] Comparative Examples 3 to 6. Difference in Concentration of Acetic Acid

[0098] The experiment was performed in the same manner as in Comparative Example 2, except that acetic acid was added to the prepared nutrient medium in concentrations of 0.1% (w / v) (Comparative Example 3), 0.15% (w / v) (Comparative Example 4), 0.2% (w / v) (Comparative Example 5), and 0.25% (w / v) (Comparative Example 6) instead of sodium acetate.

[0099] Example 16. Determination of the effect of sodium bicarbonate on microbial growth for promoting the decomposition of peracetic acid

[0100] To confirm the effect of sodium bicarbonate added to the peracetic acid mixture on the growth of microorganisms, the following experiment was performed.

[0101] Specifically, a nutrient medium (containing grade 3 distilled water, KH2PO4, citrate, MgSO4·3H2O, vitamin B1, vitamin B6, vitamin B12, vitamin H, yeast extract, peptone and dextrose) for culturing Euglena gracilis was sterilized by autoclaving at 121°C for 15 minutes and kept ready for use, and a peracetic acid solution was added to the nutrient medium to a concentration of 0.05% (w / v). Then, 96 μM FeCl3 (Sigma, 7705-08-0), 5.5 mM NaOH (Sigma, 1310-73-2) and 96 μM EDTA (Sigma, 60-00-4) were added to the above medium, and then sodium bicarbonate (NaHCO3) was added to a concentration of 0.05% (w / v). The decomposition of the added peracetic acid was confirmed by using a peroxide test (MQuant 1100810001), and then Euglena gracilis was inoculated and cultured, and the growth curve of Euglena was measured by absorbance at a wavelength of 680 nm.

[0102] Comparative Example 7: No sodium bicarbonate added

[0103] The experiment was performed in the same manner as Example 16, but without the addition of sodium bicarbonate.

[0104] Examples 17 to 21. Sodium bicarbonate concentration difference

[0105] The experiment was performed in the same manner as in Example 16, with sodium bicarbonate added to give concentrations of 0.1% (w / v) (Example 17), 0.15% (w / v) (Example 18), 0.20% (w / v) (Example 19), 0.25% (w / v) (Example 20), and 0.3% (w / v) (Example 21), respectively.

[0106] Experimental Example 1. Confirmation of the effect of promoting the decomposition of peracetic acid based on different concentrations of sodium bicarbonate

[0107] In order to confirm the effect of sodium bicarbonate and its concentration in the peracetic acid decomposition, an experiment was performed in the same manner as in Examples 1 to 6 and Comparative Example 1.

[0108] As a result, i) when sodium bicarbonate as a metal compound was added (Examples 1 to 6), it was confirmed that the decomposition rate of peracetic acid was significantly increased compared to when no sodium bicarbonate was added (Comparative Example 1) (Table 1), and ii) it was confirmed that the decomposition rate of peracetic acid increased as the sodium bicarbonate concentration increased to 0.05 to 0.25.

[0109] Specifically, it was confirmed that when 0.2% (w / v) or more of sodium bicarbonate was added (Examples 4 to 6), the decomposition of peracetic acid was completed within a few minutes, but when no sodium bicarbonate was added (Comparative Example 1), it took 48 hours for the peracetic acid to be completely decomposed (Table 1). Even when 0.05% (w / v) of sodium bicarbonate was added (Example 1), peracetic acid was completely decomposed within 3 hours, and thus the time required for decomposition was less than 1 / 8 of that in Comparative Example 1, where the decomposition time exceeded 24 hours, and thus the decomposition rate was significantly increased.

[0110] Table 1

[0111]

[0112] -NHC:NaHCO3

[0113] - The numbers in Tables 1 to 5 mean the concentration of peroxide present, for example ">100" means that more than 100 mg / L of peroxide remain.

[0114] - The numbers in brackets in Tables 1 to 5 mean the concentration of NaHCO 3 in % (w / v).

[0115] Experimental Example 2: Confirmation of the effect of promoting the decomposition of peracetic acid based on different types of metal compounds

[0116] In order to confirm the effect of the metal compound in the peracetic acid decomposition, an experiment was performed in the same manner as in Example 1, Examples 7 to 9, and Comparative Example 1.

[0117] As a result, when sodium bicarbonate (NaHCO3) as a metal compound was added (Example 1), peracetic acid decomposed quite quickly within 1 hour, but when Na2HPO4 was added (Example 7), peracetic acid was completely decomposed within 12 hours, thereby significantly increasing the decomposition rate of peracetic acid. In addition, calcium carbonate was completely decomposed within 1 hour like sodium bicarbonate (Example 8), and when calcium lactate or dipotassium hydrogen phosphate was added, it was also completely decomposed within 24 hours, thereby confirming that the decomposition rate was increased compared with the case where no metal compound was added (Examples 9 and 10) (Table 2).

[0118] Table 2

[0119]

[0120]

[0121] -NHC:NaHCO3

[0122] - The numbers in Tables 1 to 5 mean the concentration of peroxide present, for example ">100" means that more than 100 mg / L of peroxide remain.

[0123] - The concentrations of PAA and metal compounds in Table 2 are 0.05% (w / v) respectively.

[0124] Experimental Example 3: Confirming whether acetic acid or sodium acetate inhibits microbial growth

[0125] In order to confirm the effects of acetic acid (CH3COOH) as a decomposition product of peracetic acid and sodium acetate (CH3COONa) as a final product on the growth of microorganisms, experiments were performed in the same manner as in Examples 11 to 15 and Comparative Examples 2 to 6.

[0126] As a result, although the acetic acid concentration in Comparative Example 2 was 0.05% (w / v) and was not used as an inhibitor of the growth of Euglena, the growth of Euglena gracilis was inhibited by adding acetic acid at a concentration of 0.1% (w / v) or more (Comparative Examples 3 to 6). Figure 1 ).

[0127] On the other hand, in the sodium acetate addition group (Examples 11 to 15), it was confirmed that Euglena grew smoothly under all conditions ( Figure 2 ), which confirmed that sodium acetate does not inhibit the growth of Euglena.

[0128] The numbers in Tables 3 and 4 represent the absorbance (AU, 680 nm) of the culture solution of Euglena gracilis. The result values ​​in Tables 3 and 4 are shown in the graph. Figure 3 middle.

[0129] Table 3

[0130]

[0131]

[0132] Table 4

[0133]

[0134] Experimental Example 4: Confirming whether sodium bicarbonate inhibits microbial growth

[0135] In order to confirm the effect of sodium bicarbonate used when decomposing peracetic acid and peracetic acid decomposition products into final products on the growth of Euglena gracilis, an experiment was performed in the same manner as in Comparative Example 7 and Examples 16 to 21.

[0136] As a result, it was confirmed that the growth of Euglena was high in all experimental groups of Comparative Example 7 in which only peracetic acid was added and Examples 16 to 21 in which peracetic acid and sodium bicarbonate were added. It was thus confirmed that sodium bicarbonate used to decompose peracetic acid and peracetic acid products had no particular effect on the growth of Euglena gracilis (Table 5 and Table 6). Figure 4 ).

[0137] Table 5

[0138]

[0139]

[0140] - The concentration of PAA in Table 5 is 0.05% (w / v).

Claims

1. A method for decomposing peracetic acid, characterized in that: include: The step of mixing peracetic acid in water; as well as The step of adding iron ions, alkali metal hydroxide, ethylenediaminetetraacetic acid, sugar and a metal compound to the mixture, The metal compound includes any one or more selected from the group consisting of sodium bicarbonate (NaHCO3), disodium hydrogen phosphate (Na2HPO4), calcium carbonate (CaCO3), calcium lactate (Ca(CH3CHOHCOO)2·5H2O) and dipotassium hydrogen phosphate (K2HPO4).

2. The method for decomposing peracetic acid according to claim 1, characterized in that: The iron ion is Fe 2+ Ion or Fe 3+ ion.

3. The method for decomposing peracetic acid according to claim 1, characterized in that: The sugar is dextrose, glucose, sucrose or molasses.

4. The method for decomposing peracetic acid according to claim 1, characterized in that: The concentration of the metal compound is 0.005% (w / v) or more.

5. The method for decomposing peracetic acid according to claim 1, characterized in that: Also includes: Step of adding alkaline buffer.

6. A method for sterilizing a culture medium, characterized in that: include: A step of adding peracetic acid to a medium comprising a sugar and a nitrogen source and sterilizing the medium; as well as The step of adding iron ions, alkali metal hydroxide, ethylenediaminetetraacetic acid and a metal compound as a peracetic acid decomposition catalyst to the sterilized culture medium, The metal compound includes any one or more selected from the group consisting of sodium bicarbonate (NaHCO3), disodium hydrogen phosphate (Na2HPO4), calcium carbonate (CaCO3), calcium lactate (Ca(CH3CHOHCOO)2·5H2O) and dipotassium hydrogen phosphate (K2HPO4).

7. The method for sterilizing a culture medium according to claim 6, characterized in that: The iron ion is Fe 2+ Ion or Fe 3+ ion.

8. The method for sterilizing a culture medium according to claim 6, characterized in that: The sugar is dextrose, glucose, sucrose or molasses.

9. The method for sterilizing a culture medium according to claim 6, characterized in that: The concentration of the metal compound is 0.005 (w / v) % or more.

10. The method for sterilizing a culture medium according to claim 6, characterized in that: Also includes: Step of adding alkaline buffer.

11. A method for culturing microorganisms, characterized in that: include: A step of adding the culture medium sterilized by the method according to claim 6 to the bioreactor; as well as The step of inoculating microorganisms into the added culture medium and culturing them purely.

12. The method for culturing microorganisms according to claim 11, characterized in that: The microorganism is one or more selected from the group consisting of Aurantiochytrium sp., Schizochytrium sp., Chlorella sp., Synechocystis sp., Debaryomyces sp., yeasts, lactic acid bacteria, Actinomycetes, Euglena, Mortierella, filamentous fungi and photosynthetic bacteria.

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