Antimicrobial composition for plant culture medium

Abstract

The present invention relates to an antimicrobial composition for plant culture medium that exhibits excellent antimicrobial effects due to the high physicochemical stability thereof, allowing the active ingredients to remain intact and unaffected during high-temperature and high-pressure sterilization processes. Moreover, the composition can be conveniently added to plant culture media, enhancing user convenience. Therefore, the composition can be effectively used to prevent microbial contamination in plant tissue culture media or hydroponic nutrient solutions.

Description

Composition for antimicrobial activity in plant media

[0001] The present invention relates to a composition for antimicrobial use in plant media.

[0002] This invention was made possible with the support of the Ministry of SMEs and Startups under the "2023 First-Time Youth Startup in Lifetime" support project, Project No. 20135745, "High-value-added floriculture kit based on plant tissue culture technology for floriculture farmers and rare plant gardeners" project (May 1, 2023 - December 31, 2023) of the Korea Institute of Startup & Entrepreneurship Development.

[0003] Typically, plant tissue culture uses either a gel-like or solid medium containing a gelling agent, such as agarose or gellan gum, added to the culture medium, or a liquid medium that does not contain a gelling agent. The medium is sterilized while placed in a culture vessel, and then plant tissue is placed on the medium in an aseptic environment, and plant tissue culture is performed in a culture chamber. Sterilization of the medium prior to plant tissue culture is necessary to prevent the proliferation of microorganisms due to the contamination of the culture vessel and the subsequent inhibition of plant tissue growth.

[0004] For sterilization of media and culture vessels, filtration sterilization using a microporous filter or high-temperature, high-pressure sterilization under high-temperature, high-pressure conditions in an autoclave are generally performed. However, after sterilization, the inside of the culture vessel may become contaminated with microorganisms due to unexpected introduction of microbial spores into the culture vessel through air, tools, the worker's body, etc. while moving or working on the medium and culture vessel, or due to introduction of microorganisms remaining on the plant.

[0005] Therefore, to prevent microbial contamination, antibiotics such as penicillin G and streptomycin and / or antifungal agents such as benzimidazole are conventionally added during medium preparation and sterilization is performed. However, filtration sterilization is very cumbersome to perform, and high-temperature and high-pressure sterilization is problematic because it can destroy the molecular structure of antibiotics, reducing their antibacterial efficacy. In addition, antibiotics exhibit phytotoxicity, which has the disadvantage of lowering culture efficiency.

[0006] Meanwhile, hydroponics, a representative example of a plant production process that uses media in addition to plant tissue culture, is nutrient solution contamination, which is a major problem in hydroponics, which grows plants in a closed and isolated environment. Therefore, microbial growth within the nutrient solution is generally suppressed by methods such as sterilizing the nutrient solution using ultraviolet (UV) or ozone generators, or continuously circulating the nutrient solution to suppress microbial growth. However, these methods require large-scale facility investment, and some methods, such as UV sterilization, do not exhibit sufficient sterilization effects.

[0007] Accordingly, the inventors of the present invention conducted research to develop an antimicrobial composition that can be easily used by simply adding it to a plant medium and has an excellent effect in inhibiting microbial growth in the plant medium, and completed the present invention.

[0008] One object of the present invention is to provide a plant medium antimicrobial composition comprising 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one, potassium sorbate, methylparaben and propylparaben.

[0009] One aspect of the present invention provides a plant medium antimicrobial composition comprising 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one, potassium sorbate, methylparaben, and propylparaben.

[0010] A composition for antimicrobial activity in a plant medium, wherein the weight ratio of 5-chloro-2-methyl-4-isothiazolin-3-one and 2-methyl-4-isothiazolin-3-one in the first paragraph is 1:1 to 5:1.

[0011] According to one specific example of the present invention, the composition further comprises water, and may comprise, based on the total weight of the water, 0.0750 to 0.1875 wt% of 5-chloro-2-methyl-4-isothiazolin-3-one; 0.0250 to 0.0625 wt% of 2-methyl-4-isothiazolin-3-one; 0.45 to 0.60 wt% of potassium sorbate; 0.30 to 0.15 wt% of methylparaben; and 0.30 to 0.15 wt% of propylparaben.

[0012] According to one specific example of the present invention, 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one, potassium sorbate, methylparaben and propylparaben may be present in an amount of 1.15 wt% based on the total weight of water.

[0013] According to one specific example of the present invention, the plant medium may be a medium for plant tissue culture.

[0014] According to one specific example of the present invention, the composition may be included in an amount of 0.5 to 10.0 ml per 1 L of the plant tissue culture medium.

[0015] According to one specific example of the present invention, the plant medium may be a nutrient solution for hydroponics.

[0016] According to one specific example of the present invention, the composition may be included in an amount of 0.1 to 1.5 wt% based on the total weight of the hydroponic nutrient solution.

[0017] According to one specific example of the present invention, the composition may have antibacterial activity against Staphylococcus aureus and Botrytis cinerea.

[0018] According to the composition for antimicrobial activity in plant media, it has high physicochemical stability, so that the active ingredient remains intact during the high-temperature and high-pressure sterilization process, exhibiting excellent antimicrobial effects. In addition, it is non-toxic to plants and can be easily added to plant media, so it is convenient to use, and can be effectively utilized to prevent microbial contamination of plant tissue culture media or hydroponic nutrient solutions.

[0019] Figure 1 is a photograph showing contamination of a plant tissue culture medium by microorganisms 14 days after treatment with a composition for antimicrobial activity in a plant medium according to one specific example of the present invention.

[0020] One aspect of the present invention provides a plant medium antimicrobial composition comprising 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one, potassium sorbate, methylparaben, and propylparaben.

[0021] The term "plant medium" used in the present invention refers to a medium used in plant production, and includes a tissue culture medium applied to inducing proliferation, growth and differentiation of plant tissues or callus, a liquid or solid medium that provides an environment such as moisture and nutrients necessary for plant individuals to proliferate, grow and reproduce through the roots of the plant, and a medium that supports mechanical strength so that the roots of the plant can physically support them. The plant medium of the present invention may be, for example, a medium for plant tissue culture and a nutrient solution for hydroponic cultivation of plants.

[0022] In the present invention, “antimicrobial” or “antimicrobial activity” means a property of resisting microorganisms such as bacteria or fungi, and more specifically, it means a property of an antibiotic or the like to inhibit the growth or proliferation of microorganisms.

[0023] In the present invention, the "antimicrobial composition" means a composition having an activity of inhibiting the growth of microorganisms such as bacteria or fungi. In the present invention, the antimicrobial composition means a composition having an activity of inhibiting the growth of microorganisms such as bacteria or fungi, the composition containing the components 5-chloro-2-methyl-4-isothiazolin-3-one and 2-methyl-4-isothiazolin-3-one as main ingredients.

[0024] Healthy plants generally do not contain internal microorganisms, but they can become infected through diseased tissue or wounds in the roots or aboveground parts of the plant due to their structural characteristics. Even a single bacterial infection within the inoculated tissue can lead to contamination during culture. Internal infections in plants are most often caused by rod-shaped bacteria, particularly Bacillus licheniformis and Bacillus subtilis. Their spores can withstand harsh environments such as heat, dryness, and low temperatures, as well as ultraviolet light.

[0025] The antimicrobial composition of the present invention has excellent antibacterial effects against endophytic bacteria such as Bacillus licheniformis and Bacillus subtilis, and thus can effectively prevent contamination of a sterilized medium by endophytic bacteria. In addition, the antimicrobial composition of the present invention has excellent antibacterial activity against microorganisms that are problematic in plant media, such as Staphylococcus aureus and Botrytis cinerea, a fungus that causes gray mold disease, and thus can effectively prevent contamination of a sterilized medium by various external microorganisms.

[0026] In addition, the antimicrobial composition of the present invention may be used by additionally including an existing antibacterial agent and / or antifungal agent, and may further include, for example, an antibacterial / antifungal agent of the penicillin series, quinolone series, tetracycline series, sulfonamide series, and macrolide series, but is not limited thereto.

[0027] When the antimicrobial composition of the present invention is used in addition to an existing antibacterial agent and / or antifungal agent, not only can it exhibit antimicrobial activity against a wider variety of microorganisms due to the different mechanisms of antibacterial / antifungal action, but the antimicrobial activity can also be further enhanced, and sufficient antimicrobial activity can be exhibited even with a smaller amount compared to when the existing antibacterial / antifungal agent is used alone, so that the problem of plant toxicity caused by the use of the existing antibacterial / antifungal agent alone can be alleviated or resolved.

[0028] The antimicrobial composition according to the present invention may be prepared in the form of, for example, a directly sprayable solution, powder, or suspension, or as a highly concentrated aqueous, oily, or other suspension, dispersion, emulsion, oily dispersion, paste, dust, dustable material, or granule, but is not limited thereto.

[0029] According to one specific example of the present invention, the weight ratio of 5-chloro-2-methyl-4-isothiazolin-3-one and 2-methyl-4-isothiazolin-3-one may be 1:1 to 5:1.

[0030] When 5-chloro-2-methyl-4-isothiazolin-3-one is contained in an amount of less than 1 weight per 1 weight of 2-methyl-4-isothiazolin-3-one, or 5-chloro-2-methyl-4-isothiazolin-3-one is contained in an amount of more than 5 weight per 1 weight of 2-methyl-4-isothiazolin-3-one, the antibacterial and antifungal properties of the composition may not be sufficient, and the synergistic effect with potassium sorbate, methylparaben and / or propylparaben may not be sufficient. Therefore, the weight ratio of 5-chloro-2-methyl-4-isothiazolin-3-one and 2-methyl-4-isothiazolin-3-one is preferably 1:1 to 5:1, more preferably 2:1 to 4:1, and most preferably 3:1.

[0031] According to one specific example of the present invention, the composition further comprises water, and may comprise, based on the total weight of the water, 0.075 to 0.1875 wt% of 5-chloro-2-methyl-4-isothiazolin-3-one; 0.025 to 0.0625 wt% of 2-methyl-4-isothiazolin-3-one; 0.45 to 0.60 wt% of potassium sorbate; 0.30 to 0.15 wt% of methylparaben; and 0.30 to 0.15 wt% of propylparaben.

[0032] When 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one and methylparaben are 0.0375 wt%, 0.0125 wt% and 0.35 wt%, respectively, based on the total weight of water, no synergistic effect on antibacterial and antifungal activity is observed in the range of 0.40 to 0.60 wt% of potassium sorbate and 0.35 to 0.15 wt% of propylparaben. When 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one and methylparaben are 0.0750 wt%, 0.0250 wt% and 0.30 wt%, respectively, based on the total weight of water, no synergistic effect on antibacterial and antifungal activity is observed in the range of 0.40 to 0.55 wt% of potassium sorbate and 0.35 to 0.20 wt% of propylparaben. When 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one and methylparaben are 0.1125 wt%, 0.0375 wt% and 0.25 wt%, respectively, based on the total weight of water, potassium sorbate and propylparaben are 0.40 to 0.45 wt% In the range of 0.35 to 0.30 wt%, no synergistic effect on antibacterial and antifungal activity is observed. When 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one and methylparaben are respectively 0.1500 wt%, 0.0500 wt% and 0.20 wt% based on the total weight of water, no synergistic effect on antibacterial and antifungal activity is observed at 0.40 wt% of potassium sorbate and 0.35 wt% of propylparaben, and at 0.60 wt% of potassium sorbate and 0.15 wt% of propylparaben. When 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one and methylparaben are 0.1875 wt%, 0.0625 wt% and 0.15 wt%, respectively, based on the total weight of water, no synergistic effect on antibacterial and antifungal activity is observed in the ranges of 0.40 wt% potassium sorbate and 0.35 wt% propylparaben, and 0.55 to 0.60 wt% potassium sorbate and 0.20 to 0.15 wt% propylparaben.

[0033] Meanwhile, when 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one and methylparaben are 0.0750 wt%, 0.0250 wt% and 0.30 wt%, respectively, based on the total weight of water, a synergistic effect on antibacterial and antifungal activity is observed at 0.60 wt% of potassium sorbate and 0.15 wt% of propylparaben. When 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one and methylparaben are 0.1125 wt%, 0.0375 wt% and 0.25 wt%, respectively, based on the total weight of water, a synergistic effect on antibacterial and antifungal activity is exhibited at 0.50 to 0.60 wt% of potassium sorbate and 0.25 to 0.15 wt% of propylparaben. When 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one and methylparaben are 0.1500 wt%, 0.0500 wt% and 0.20 wt%, respectively, based on the total weight of water, a synergistic effect on antibacterial and antifungal activity is exhibited at 0.45 to 0.55 wt% of potassium sorbate and 0.30 to 0.20 wt% of propylparaben. When 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one and methylparaben are 0.1875 wt%, 0.0625 wt% and 0.15 wt%, respectively, based on the total weight of water, a synergistic effect on antibacterial and antifungal activity is exhibited at 0.45 to 0.50 wt% of potassium sorbate and 0.30 to 0.25 wt% of propylparaben.

[0034] According to one specific example of the present invention, 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one, potassium sorbate, methylparaben and propylparaben may be present in an amount of 1.15 wt% based on the total weight of water.

[0035] According to one specific example of the present invention, the plant medium may be a medium for plant tissue culture.

[0036] According to one specific example of the present invention, the composition may be included in an amount of 0.5 to 10.0 ml per 1 L of the plant tissue culture medium.

[0037] When the plant medium antimicrobial composition of the present invention is used in an amount of less than 0.5 ml per 1 L of a plant tissue culture medium, the antimicrobial activity against microorganisms in the medium may not be sufficient, and when it is used in an amount exceeding 10.0 ml, the antimicrobial activity does not significantly increase compared to when 10.0 ml is used. Therefore, the antimicrobial composition of the present invention is preferably used in an amount of 0.5 to 10.0 ml per 1 L of a plant tissue culture medium, and more preferably, in an amount of 1 to 4 ml.

[0038] According to one specific example of the present invention, the plant medium may be a nutrient solution for hydroponics.

[0039] According to one specific example of the present invention, the composition may be included in an amount of 0.1 to 1.5 wt% based on the total weight of the hydroponic nutrient solution.

[0040] When the plant medium antimicrobial composition of the present invention is used in an amount of less than 0.1 wt% based on the total weight of the nutrient solution for hydroponics, the antimicrobial activity against microorganisms in the nutrient solution may not be sufficient, and when it is used in an amount exceeding 1.5 wt%, the antimicrobial activity does not significantly increase compared to when it is used in an amount of 1.5 wt%. Therefore, the plant medium antimicrobial composition of the present invention is preferably used in an amount of 0.1 to 1.5 wt% based on the total weight of the nutrient solution for hydroponics, and more preferably, in an amount of 0.15 to 0.50 wt%.

[0041] The present invention will be described in more detail below through one or more examples. However, these examples are intended to exemplify the present invention and the scope of the present invention is not limited to these examples.

[0042]

[0043] Example 1. Confirmation of the mixing ratio of potassium sorbate, methylparaben, and propylparaben that exhibit a synergistic effect with a 3:1 mixture of 5-chloro-2-methyl-4-isothiazolin-3-one / 2-methyl-4-isothiazolin-3-one.

[0044] 1-1. Check the mixing ratio of each ingredient that produces a synergistic effect.

[0045] The mixing ratio of 5-chloro-2-methyl-4-isothiazolin-3-one / 2-methyl-4-isothiazolin-3-one (MCI / MI) 3:1 mixture and potassium sorbate, methylparaben, and propylparaben showing synergistic effects were analyzed.

[0046] Specifically, the plant tissue culture medium was prepared by adding 20 g / L of sucrose to the Murashing & Skoog medium basic salts and adjusting the pH to 5.7, and placing the mixture in a culture vessel. Thereafter, a plant medium antimicrobial composition was prepared by mixing a mixture of MCI / MI 3:1, potassium sorbate, methylparaben, and propylparaben in a combination selected from the ratios in Table 1 so that the total weight of water would be 1.15 wt%. Thereafter, the prepared plant medium antimicrobial composition was added to the plant tissue culture medium at a concentration of 1 ml / L, sterilized under high temperature and high pressure, and then the stopper of the culture vessel was removed to allow the inflow of microorganisms through air. The degree of contamination of the medium was visually measured on the 14th day to evaluate the antimicrobial effect. - means that no contamination was observed visually, and + means that contamination was observed visually.

[0047]

[0048] Classification Additive Concentration (Wt%)MCI / MI 3:1 Mixed FormulationMCI0.0375, 0.0750, 0.1125, 0.1500, 0.1875MI0.0125, 0.0250, 0.0375, 0.0500, 0.0625Subtotal0.05, 0.10, 0.15, 0.20, 0.25Potassium Sorbate0.40, 0.45, 0.50, 0.55, 0.60Methylparaben0.15, 0.20, 0.25, 0.30, 0.35Propylparaben0.15, 0.20, 0.25, 0.30, 0.35

[0049]

[0050] As a result, the synergistic effect was confirmed according to the mixing ratio of each component, as shown in Table 2.

[0051]

[0052] Potassium sorbate addition concentration (wt%) 0.40 0.45 0.50 0.55 0.60 MCI / MI 3:1 mixed preparation addition concentration (wt%) 0.05+++++ 0.35 Methylparaben addition concentration (wt%) 0.10++++-0.30 0.15++--- 0.25 0.20+---+0.20 0.25+--++ 0.15 0.35 0.30 0.25 0.20 0.15 Propylparaben addition concentration (wt%)

[0053]

[0054] Through these results, it was confirmed that when a mixture of 0.10 to 0.25 wt% of MCI / MI 3:1 based on the total weight of water, 0.45 to 0.6 wt% of potassium sorbate, 0.30 to 0.15 wt% of methylparaben, and 0.30 to 0.15 wt% of propylparaben was added to a plant tissue culture medium at a concentration of 1 ml / L, a synergistic effect could be exhibited in preventing contamination of the medium by microorganisms during a 14-day plant tissue culture process.

[0055]

[0056] 1-2. Confirmation of antimicrobial activity according to treatment concentration of plant medium antimicrobial composition

[0057] The antimicrobial activity of the plant medium antimicrobial composition of the present invention was analyzed according to the treatment concentration for the plant tissue culture medium.

[0058] Specifically, the antimicrobial activity was analyzed in the same manner as in Example 1-1, except that the plant medium antimicrobial composition, in which MCI / MI 3:1 mixture, potassium sorbate, methylparaben, and propylparaben were mixed at 0.15 wt%, 0.50 wt%, 0.25 wt%, and 0.25 wt%, respectively, based on the total weight of water, was treated at a concentration of 0, 1, 2, or 4 ml / L in the plant tissue culture medium, at a mixing ratio in which the synergistic effect was confirmed in Example 1-1.

[0059] As a result, while contamination by microorganisms was observed from the 7th day in the untreated group, no contamination by microorganisms was observed until the 14th day in the group treated with the plant medium antimicrobial composition of the present invention at all concentrations (Fig. 1).

[0060]

[0061] Example 2. Confirmation of excellent antimicrobial activity and low phytotoxicity of a plant medium antimicrobial composition in apple dwarf rootstock tissue culture.

[0062] In order to confirm the excellent antimicrobial activity and low phytotoxicity of the plant medium antimicrobial composition that was confirmed to exhibit a synergistic effect in Example 1, it was applied to tissue culture of M.26, an apple rootstock variety.

[0063] Specifically, growing points were collected from axillary buds of M.26, an apple dwarf rootstock cultivar, and the collected growing points were planted on LS (Linsmaier and Skoog, 1965) medium containing 1 mg / L of 6-benzylamino purine (BAP), 0.5 mg / L of indole acetic acid (IAA), and 30 g / L of sucrose, and shoots were grown for 90 days. After 90 days, in vitro growth characteristics such as the number of new shoots, new shoot height, contamination rate, and mortality rate were investigated. At this time, the plant medium antimicrobial composition was prepared by mixing MCI / MI 3:1 mixture, potassium sorbate, methylparaben, and propylparaben at 0.15 wt%, 0.50 wt%, 0.25 wt%, and 0.25 wt%, respectively, based on the total weight of water, and the prepared plant medium antimicrobial composition was treated at a concentration of 0, 0.5, 1, 2, or 4 ml / L in the LS medium, respectively. In addition, as a comparative group, penicillin G, carbenicillin, or kanamycin was treated to the medium at the concentrations shown in Table 3 below, respectively. The treatment concentration for each antibiotic was set with reference to the Antibiotics in Plant Tissue Culture Protocol (Merck).

[0064]

[0065] Classification concentration (ppm) Penicillin G 0, 0.5, 1, 2, 4 Carbenicillin 0, 500, 1000, 2000, 4000 Kanamycin 0, 40, 80, 160, 320

[0066]

[0067] As a result, it was confirmed that the number of new shoots and the height of new shoots significantly increased and the mortality rate was lower in the group treated with the plant medium antimicrobial composition of the present invention compared to the penicillin G, carbenicillin, or kanamycin treatment group. In addition, the contamination rate by microorganisms was also confirmed to be significantly lower, and it was confirmed that no contamination occurred at all in the 4㎖ / L treatment group. In particular, according to a two-way analysis of variance, it was confirmed that the interaction effect according to the plant medium antimicrobial composition of the present invention and its treatment concentration had a significant effect on the new shoot height, contamination rate, and mortality rate (Table 4).

[0068]

[0069] Treatment concentration, number of shoots (units / section), shoot length (mm), contamination rate (%), mortality rate (%), penicillin G0 ppm3.4 a 32.5 a 18.9 a 15.5 d 0.5ppm3.3 ab 28.0 b 14.1 b 20 cd 1ppm3.0 bc 27.5 b 12.4 bc 23.4 bc 2ppm2.9 c 26.5 b 10.3 c 27.0 ab 4ppm2.7 c 25.7 b 9.3 c 30.4 a Carbenicillin 0 ppm 3.2 a 32.5 a 18.9 a 15.5 c 500ppm2.7 b 30.0 ab 13.8 b 21.4 b 1000ppm2.5 bc 27.9 bc 11.6 bc 23.8 b 2000ppm2.4 bc 26.6c 9.7 cd 30.2 a 4000ppm2.2 c 25.9 c 7.5 d 35.0 a Kanamycin 0 ppm 3.0 a 32.5 a 18.9 a 15.5 d 40ppm2.9 a 30.7 ab 15.0 b 22.8 c 80ppm2.7 b 26.4 bc 13.2 bc 29.4 b 160ppm2.4 bc 23.8 c 12.1 bc 35.0 ab 320ppm2.1 c 22.7 c 10.4 c 38.6 a Composition for antimicrobial activity in plant culture medium 0㎖ / L3.3 a 32.5 a 18.9 a 15.5 b 0.5㎖ / L3.2 a 32.0 a 6.4 b 16.9 b 1㎖ / L3.2 a 31.9 a 4.5 bc 19.8 a 2㎖ / L3.2 a 31.9 a 1.8 cd 20.4 a 4㎖ / L3.0 a 31.8 a 0.0 d 21.4 a F-value two-way analysis of variance Antibiotic type (A) 14.44 ** 7.43 ** 15.27 ** 8.62 ** Antibiotic concentration (B) 23.24 ** 7.82 ** 22.59 **17.28 ** A×B1.62 NS 3.09 ** 3.24 ** 3.55 ** a~d Mean separation within columns by Duncan's multiple range test at P=0.05. NS,*,** Nonsignificant or significant at P≤0.05 or 0.01, respectively.

[0070]

[0071] Example 3. Confirmation of low phytotoxicity of a plant medium antimicrobial composition in tissue culture of various plants.

[0072] It was further analyzed whether the plant medium antimicrobial composition of the present invention exhibits low phytotoxicity when culturing various plant tissues.

[0073] Specifically, a plant medium antimicrobial composition was prepared by mixing MCI / MI 3:1 mixture, potassium sorbate, methylparaben, and propylparaben at 0.15 wt%, 0.50 wt%, 0.25 wt%, and 0.25 wt%, respectively, based on the total weight of water. A plant tissue culture medium was prepared by adding 20 g / L of sucrose and 1.0 mg / L of 2,4-D, a plant growth regulator, to Murashing&Skoog medium basal salts, adjusting the pH to 5.7, and placing the mixture in a culture vessel. Thereafter, the plant medium antimicrobial composition was added to the plant tissue culture medium in amounts of 0.05 wt% to 0.50 wt% in increments of 0.05 wt% based on the total weight of the plant tissue culture medium, and sterilized at high temperature and high pressure. Leaf discs were prepared by cutting cucumber, anthurium, and dahlia leaves into 1 cm × 1 cm pieces, 20 for each plant, and the leaf discs were placed on sterilized media. The shoot differentiation rate per disc was measured to evaluate plant toxicity. The differentiation rate was calculated by dividing the number of shoots generated per disc by the number of discs.

[0074]

[0075] Classification Addition Concentration (Wt%) 0.05 0.100 150 200 250 300 350 400 450 50 Cucumber 7.7 7.9 7.75 7.67 657 357 27.06 75 6.9 Anthurium 2.45 2.45 2.62 352 32 62 32 252 32 15 Dahlia 3.15 3.13 253 153 05 3.15 3.02 95 2.95 2.85

[0076]

[0077] As a result, it was confirmed that no phytotoxicity was observed during the tissue culture process of cucumber, anthurium, and dahlia at all added concentrations (Table 5).

[0078] Through these results, it was confirmed that the plant medium antimicrobial composition of the present invention can be effectively used to suppress microbial contamination of the medium that may occur during the tissue culture process of various plants without phytotoxicity.

[0079]

[0080] Example 4. Confirmation of antimicrobial activity of a plant medium antimicrobial composition in a nutrient solution for hydroponics.

[0081] 4-1. Confirmation of antimicrobial activity of plant medium antimicrobial composition against Staphylococcus aureus in nutrient solution for hydroponic cultivation

[0082] The antimicrobial activity of the plant medium antimicrobial composition of the present invention against Staphylococcus aureus in a nutrient solution for hydroponics was analyzed according to the concentration of addition.

[0083] Specifically, a composition for antimicrobial activity of plant media was prepared by mixing MCI / MI 3:1 mixture, potassium sorbate, methylparaben, and propylparaben at 0.15 wt%, 0.50 wt%, 0.25 wt%, and 0.25 wt%, respectively, based on the total weight of water. The nutrient solution for hydroponic cultivation was prepared by adding 3.2 × 10 Staphylococcus aureus to 1 L of Yamazaki standard solution. 5It was prepared by inoculating at a concentration of CFU / ㎖. Immediately after inoculation, the above plant medium antimicrobial composition was added to the hydroponic nutrient solution in an amount of 0.05 wt% to 0.50 wt% in increments of 0.05 wt% based on the total weight of the hydroponic nutrient solution, and left at 25°C for 30 minutes. Then, the antibacterial activity was evaluated by measuring the reduction rate of the microbial community according to the bacteriostatic reduction rate calculation method.

[0084]

[0085] Added concentration (weight %) Antibacterial power (%) 0.00 (control) -193.550.0564.500.1088.210.1591.170.2092.730.2596.100.3098.590.3599.120.4099.990.4599.990.5099.99

[0086]

[0087] As a result, it was confirmed that when the plant medium antimicrobial composition of the present invention was added to a nutrient solution for hydroponics in an amount of 0.25 wt% or more, it exhibited an antibacterial activity of 92% or more, and when added in an amount of 0.40 wt% or more, it exhibited an antibacterial activity of 99.99% (Table 6).

[0088] Through these results, it was confirmed that when the plant medium antimicrobial composition of the present invention is added at a concentration of 0.40 wt% or more based on the total weight of the nutrient solution for hydroponics, contamination of the nutrient solution by Staphylococcus aureus during hydroponics can be prevented.

[0089]

[0090] 4-2. Confirmation of the antifungal activity of a plant medium antimicrobial composition against Botrytis cinerea in a nutrient solution for hydroponics.

[0091] The antifungal activity of the plant medium antimicrobial composition of the present invention against Botrytis cinerea, a fungus causing gray mold disease in a nutrient solution for hydroponics, was analyzed according to the concentration of addition.

[0092] Specifically, the nutrient solution for hydroponics is 3.2×10 Botrytis cinerea in 1 L of Yamazaki standard solution. 5 It was prepared by inoculating at a concentration of CFU / ㎖. Immediately after inoculation, the plant medium antimicrobial composition of Example 4-1 was added to the hydroponic nutrient solution in an amount of 0.05 wt% to 0.50 wt% in increments of 0.05 wt% based on the total weight of the hydroponic nutrient solution, and left at 25°C for 30 minutes. Then, the antifungal activity was evaluated by measuring the reduction rate of the microbial community according to the bacteriostatic reduction rate calculation method.

[0093]

[0094] Added concentration (weight %) Antifungal activity (%) 0.00 (control) - 0.05 98.41 0.1099.50 0.1599.99 0.2099.99 0.2599.99 0.3099.99 0.3599.99 0.4099.99 0.4599.99 0.5099.99

[0095]

[0096] As a result, it was confirmed that when the plant medium antimicrobial composition of the present invention was added to a nutrient solution for hydroponics at 0.05 wt% or more, it exhibited an antifungal activity of 98.41% or more, and when it was added at 0.15 wt% or more, it exhibited an antifungal activity of 99.99% (Table 7).

[0097] Through these results, it was confirmed that when the plant medium antimicrobial composition of the present invention is added at a concentration of 0.15 wt% or more based on the total weight of the nutrient solution for hydroponics, contamination of the nutrient solution by Botrytis cinerea during hydroponics can be prevented.

[0098]

[0099] The present invention has been described above, focusing on specific embodiments thereof. Those skilled in the art will appreciate that the present invention can be implemented in modified forms without departing from its essential characteristics. Therefore, the disclosed embodiments should be considered illustrative rather than limiting. The scope of the present invention is set forth in the claims, not the foregoing description, and all differences within the scope equivalent thereto should be construed as being encompassed by the present invention.

Claims

A composition for antimicrobial use in plant media comprising 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one, potassium sorbate, methylparaben and propylparaben.

2. A composition for antimicrobial activity in a plant medium, wherein the weight ratio of 5-chloro-2-methyl-4-isothiazolin-3-one and 2-methyl-4-isothiazolin-3-one in paragraph 1 is 1:1 to 5:

1.

3. In the fourth paragraph, the composition further comprises water, Based on the total weight of the above water, 0.0750 to 0.1875 wt% of 5-chloro-2-methyl-4-isothiazolin-3-one; 0.0250 to 0.0625 wt% of 2-methyl-4-isothiazolin-3-one; 0.45 to 0.60 wt% potassium sorbate; 0.30 to 0.15 wt% methylparaben; and Propylparaben 0.30 to 0.15 wt% A composition for antimicrobial activity in plant media comprising:

4. A composition for antimicrobial activity in a plant medium, wherein 5-chloro-2-methyl-4-isothiazolin-3-one, 2-methyl-4-isothiazolin-3-one, potassium sorbate, methylparaben and propylparaben are 1.15 wt% based on the total weight of water in the third paragraph.

5. A plant medium antimicrobial composition according to claim 1, wherein the plant medium is a medium for plant tissue culture.

6. A plant medium antimicrobial composition according to claim 5, wherein the composition is contained in an amount of 0.5 to 10.0 ml per 1 L of the plant tissue culture medium.

7. A composition for antimicrobial use in a plant medium, wherein the plant medium is a nutrient solution for hydroponic cultivation.

8. A composition for antimicrobial activity in a plant medium, wherein the composition is included in an amount of 0.1 to 1.5 wt% based on the total weight of the hydroponic nutrient solution in the 7th paragraph.

9. A plant medium antimicrobial composition according to claim 1, wherein the composition has antimicrobial activity against Staphylococcus aureus and Botrytis cinerea.

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