JP2018090514A - Fine bubble mixed liquid having bactericidal effect - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide highly-versatile fine bubble mixed liquid capable of maintaining stably an excellent bactericidal effect.SOLUTION: There is provided fine bubble mixed liquid having a diameter of 60-200 nm, formed by mixing fine bubbles containing reactive oxygen species inside, and also having a bactericidal effect.SELECTED DRAWING: Figure 1

Description

  The present invention relates to a fine bubble mixture having a bactericidal effect.

  For example, oxygen nanobubble water disclosed in Patent Document 1 is known as a microbubble mixed liquid having a bactericidal effect. This oxygen nanobubble water contains oxygen nanobubbles containing oxygen in bubbles, and the bubble diameter can be drastically reduced by giving physical stimulation such as ultrasonic waves to the bubbles containing oxygen. Has been.

JP 2005-246294 A

  However, since the oxygen nanobubble water disclosed in Patent Document 1 stabilizes the bubble diameter by the action of the electrolyte mixed in the aqueous solution, there is a possibility that microbubbles cannot be maintained if the components of the aqueous solution change. there were. The aqueous solution is desirably set to a salt concentration in the range of 0.01 to 3.5%. However, the oxygen nanobubble water obtained thereby contains salt, and its use may be limited. was there.

  Then, this invention aims at provision of the fine bubble liquid mixture which can maintain a favorable disinfection effect stably and is highly versatile.

  The object of the present invention is achieved by a microbubble mixture having a diameter of 60 to 200 nm and having a bactericidal effect, in which microbubbles containing active oxygen species are mixed.

  ADVANTAGE OF THE INVENTION According to this invention, while being able to maintain a favorable sterilization effect stably, the highly versatile fine bubble liquid mixture can be provided.

It is a longitudinal cross-sectional view which shows an example of the manufacturing apparatus which manufactures the fine bubble liquid mixture which concerns on one Embodiment of this invention. It is a principal part disassembled perspective view of the manufacturing apparatus shown in FIG. It is a principal part enlarged view which shows the operation state of the manufacturing apparatus shown in FIG. It is a longitudinal cross-sectional view which shows the modification of the manufacturing apparatus shown in FIG. It is a principal part side view which shows the other modification of the manufacturing apparatus shown in FIG. It is a figure which shows the antibacterial effect with respect to Bacillus subtilis, Escherichia coli, and Aspergillus niger of this invention. It is a figure which shows the change of the amount of dissolved oxygen at the time of the production | generation of a bubble liquid mixture. It is a figure which shows the carbon dioxide blowing test result done after the production | generation of a bubble liquid mixture. It is a figure which shows the dissolved oxygen concentration change after the production | generation of a bubble liquid mixture. It is a figure which shows the antibacterial effect when Escherichia coli is mist-processed using a bubble liquid mixture. It is a figure which shows the antibacterial effect when black mushroom mold is mist-processed using a bubble liquid mixture. It is a figure which shows the change of the antibacterial activity with respect to Bacillus subtilis when a bubble liquid mixture is processed with superoxide demutase or catalase. It is a figure which shows the change of the antibacterial activity with respect to colon_bacillus | E._coli when a bubble liquid mixture is processed with superoxide demutase or catalase. It is a figure which shows the relationship with oxygen, a superoxide radical, hydrogen peroxide, a hydroxyl radical, and an enzyme. It is a figure of the result of having measured the hydrogen peroxide content by the color development reaction after processing the air bubble mixture with superoxide demutase or catalase. Comparison chart of average particle size of oxygen water and ultra fine bubble water (UFB). Comparison of the number of bubbles contained in oxygen water and ultra fine bubble water (UFB). It is a figure which shows the time-dependent change after manufacture of oxygen water, and a zeta potential with time. It is a figure which shows the time-dependent change of the elapsed days after manufacture of oxygen water, and the amount of superoxide radicals.

  Hereinafter, embodiments of the present invention will be described with reference to the accompanying drawings. FIG. 1 is a longitudinal sectional view showing an example of a production apparatus for producing a fine bubble mixture according to an embodiment of the present invention. As shown in FIG. 1, the production apparatus 1 for a fine bubble mixture includes a storage tank 4 that stores a liquid L, and a bubble supply device 6 that supplies bubbles to the liquid L in the storage tank 4.

  The bubble supply device 6 includes a drive motor 10 fixed to the upper surface of the support plate 2 placed in the upper opening of the storage tank 4, a cylindrical rotary cylinder 20 that is rotationally driven by the drive motor 10, and the rotary cylinder 20. An outer cylinder 30 that hangs down from the support plate 2 so as to accommodate the liquid, a circulation device 40 that circulates the liquid stored in the storage tank 4, and a gas-liquid mixing unit that mixes bubbles with the liquid L circulated by the circulation device 40 50. In this embodiment, the support plate 2 is disposed so as to seal the storage tank 4, but the support plate 2 may be configured to allow communication between the inside and the outside of the storage tank 4.

  The drive motor 10 is configured such that both upper and lower ends of the output shaft 11 protrude from the casing 12, and the output shaft 11 is formed with an introduction path 11 a penetrating the center portion in the axial direction. An upper end portion of the output shaft 11 is connected to an introduction portion 14 through a rotary joint 13 so as to be airtight and relatively rotatable.

  The lower end portion of the output shaft 11 passes through a through hole 2a formed in the support plate 2 and is connected and fixed to the upper end portion of the rotating cylinder 20, and is supported and rotated so as to hang down the rotating cylinder 20. . The rotating cylinder 20 is made of, for example, a metal material such as stainless steel, and the ejection portion 22 including a plurality of ejection holes 22a is formed on the lower outer peripheral surface. The ejection holes 22a are formed at equal intervals along the circumferential direction so as to uniformly inject the bubble mixture introduced into the rotary cylinder 20 from the introduction path 11a of the output shaft 11 toward the periphery, Further, the ejection portions 22 are formed at equal intervals in the axial direction so as to have a predetermined height (for example, about 150 mm). Although the diameter of the ejection hole 22a is not specifically limited, For example, it is 0.1-1.5 mm. It is preferable that the outer peripheral surface of the rotating cylinder 20 is formed smoothly without providing a stirring blade or the like.

  A stirring member 24 is provided in the lower part inside the rotary cylinder 20. As shown in an exploded perspective view in FIG. 2, the stirring member 24 has a plurality of flat plate-like rotary blades 24 a arranged on a lower disk 24 b having a diameter substantially the same as the outer diameter of the rotary cylinder 20. In the case where a large number of rotor blades 24a are provided, it is preferable to arrange them radially at equal intervals. An upper disk 24c having a diameter substantially the same as the inner diameter of the rotary cylinder 20 is attached to the upper part of the rotary blade 24a. As shown by arrows in FIG. 2, the stirring member 24 is inserted from below the rotary cylinder 20 so that the rotary blades 24a are positioned between the circumferentially adjacent ejection holes 22a, and the lower disk 24b rotates. It is fixed to the rotating cylinder 20 so as to close the lower opening of the cylinder 20. The upper disk 24c has a communication hole 24d formed in the center, and the bubble mixture introduced into the rotary cylinder 20 passes through the communication hole 24d and is agitated by the rotary blade 24a and ejected from the ejection hole 22a. Is done. The shape and arrangement of the rotary blade 24a are not particularly limited as long as a swirl flow can be generated inside the rotary cylinder 20. For example, instead of the flat plate-like rotary blade 24a, a curved or spiral shape is used. Rotating blades such as airfoils may be used.

  The outer cylinder 30 is formed in a straight cylinder shape made of, for example, a resin such as acrylic or a metal such as stainless steel, and is arranged coaxially with a gap between the outer cylinder 30 and the outer peripheral surface of the rotating cylinder 20, and the upper end portion is a support plate. 2 is fixed. The lower end of the outer cylinder 30 substantially coincides with the lower end of the inner cylinder 20, and the ejection portion 22 of the rotating cylinder 20 is covered with the outer cylinder 30. A spiral guide plate (not shown) may be provided on the inner peripheral surface of the outer cylinder 30 so that a swirling flow of the liquid L is likely to occur inside the outer cylinder 30.

  The circulation device 40 is connected to the discharge port 4 a formed at the bottom of the liquid storage tank 4 and the introduction part 14, and the liquid L in the liquid storage tank 4 is interposed between the pipe 42 and the liquid storage tank 4. And a circulation pump 44 to be taken out from the outside. The gas-liquid mixing unit 50 is disposed downstream of the circulation pump 44 in the middle of the pipe 42, and is pressurized and supplied from an air supply device 52 such as a compressor or a gas cylinder to the liquid L passing through the inside. Mix the gas.

  The gas supplied from the air supply device 52 is air, oxygen, molecular oxygen, triplet oxygen, ozone, ozonide, hydrogen peroxide, hydroperoxyl, hydrogen oxide, hydroxy radical, alkoxyl radical, carbon monoxide, It is preferable to use a gas body containing oxygen atoms such as carbon dioxide and nitric oxide.

  The configuration of the gas-liquid mixing unit 50 is not particularly limited, and for example, a pore blowing method in which bubbles are mixed into the liquid through the pores of the porous film, and gas and liquid are swirled at high speed to be mixed. A swirl flow system, a venturi system that introduces gas using a negative pressure generated by the passage of the liquid L, or the like may be used. In addition, the gas-liquid mixing unit 50 can suck the outside air and mix it with the liquid L without providing the air supply device 52. The bubble liquid mixture discharged from the gas-liquid mixing unit 50 is supplied into the liquid storage tank 4 from the ejection unit 22 of the rotary cylinder 20. Furthermore, the gas-liquid mixing unit 50 may be configured to include a liquid electrolysis device that generates bubbles by electrolyzing the liquid. In this configuration, the generated bubbles are separated from the liquid that has not been electrolyzed. You may supply in the piping 42 in the mixed state.

  A branch pipe 53 is connected to the pipe 42 constituting the circulation path, and by adjusting the opening degree of the on-off valve 54 provided in the middle of the branch pipe 53, a part of the bubble mixed liquid flowing through the pipe 42 is reduced. It can be continuously removed from the branch pipe 53. The pipe 42 is configured to be able to supply the liquid L from the liquid supply device 55, and the liquid L corresponding to the amount discharged from the branch pipe 53 is continuously adjusted by adjusting the opening degree of the on-off valve 56. Can be replenished.

  Next, a method for producing a fine bubble mixture having a bactericidal effect using the fine bubble mixture production apparatus 1 having the above-described configuration will be described. As shown in FIG. 1, a liquid L such as water is stored in the storage tank 4, and substantially the entire rotating cylinder 20 is immersed in the liquid. Next, when the drive motor 10 and the circulation pump 44 are operated, the rotary cylinder 20 rotates around the axis together with the rotary blade 24a as shown by an arrow in FIG. 3, and is introduced into the rotary cylinder 20 from the circulation device 40. The bubble mixture is swirled by the rotor blades 24a and ejected from the ejection section 22.

  When the rotating cylinder 20 rotates at a high speed, the liquid level S of the liquid L becomes a mortar-shaped recess centered on the rotating cylinder 20, and the liquid level S reaches the vicinity of the ejection part 22 around the rotating cylinder 20. Decrease and move up and down around here. For this reason, the jet stream F of the bubble mixed liquid discharged from each of the ejection holes 22a is stirred while colliding with the liquid surface S and taken into the liquid, so that the bubbles become fine and diffuse into the liquid L. Since the amount of decrease in the liquid level S due to the rotation of the rotary cylinder 20 increases as the rotary cylinder 20 rotates at high speed, the rotational speed of the rotary cylinder 20 is adjusted so that the liquid level S is maintained in the vicinity of the ejection portion 22. It is preferable (for example, 3600 to 15000 revolutions per minute, or 15000 revolutions per minute or more). The gap generated between the rotating cylinder 20 and the outer cylinder 30 (length D in FIG. 2) is preferably set as appropriate so that the desired liquid level S can be easily lowered, for example, 10 to 20 mm or 20 mm or more. It is.

  Thus, according to the manufacturing apparatus 1 of the fine bubble mixed liquid of the present embodiment, the bubble mixed liquid ejected from the ejection part 22 by the rotation of the rotary cylinder 20 is mixed with the liquid L while performing liquid drainage. In addition, a fine bubble mixed liquid in which fine bubbles such as microbubbles and nanobubbles are mixed can be efficiently generated.

  Further, since the rotary cylinder 20 is radially arranged so that the rotary blades 24 extend from the inner peripheral surface toward the center, the rotational resistance is increased as compared with the case where the rotary blades are provided on the outer peripheral surface of the rotary cylinder 20. Can be suppressed, and power saving can be achieved.

  The liquid L can be appropriately selected other than water depending on the application. The liquid L may be at normal temperature, but is preferably low temperature in order to maintain fine bubbles for a long time, for example, preferably 10 ° C. or less, more preferably 8 ° C. or less. In order to keep the liquid L in the liquid storage tank 4 at a low temperature, a heat exchanger for cooling is arranged along the outer periphery of the liquid storage tank 4 or so as to be immersed in the liquid L in the liquid storage tank 4. Also good. It is preferable that the liquid L maintains the low temperature state even after the fine bubbles are mixed.

  Moreover, in this embodiment, although the outer cylinder 30 is arrange | positioned so that the desired mortar-shaped liquid level S may arise easily at the time of rotation of the rotation cylinder 20, the outer cylinder 30 is not essential, for example, When the storage tank 4 has a small cylindrical shape, a configuration in which the outer cylinder 30 is not provided may be used.

  In the present embodiment, the bubble mixture can be stably ejected from the ejection portion 22 by forming the introduction path 11a for introducing the bubble mixture into the rotary cylinder 20 in the output shaft 11 of the drive motor 10. However, the introduction path 11a may be other than the output shaft 11, or may be configured to introduce the bubble mixed liquid from the upper part or the side part of the rotary cylinder 20 via a mechanical seal or the like.

  Moreover, the structure which provided further the support member 60 which supports the lower end part of the rotary cylinder 20 rotatably may be sufficient as the manufacturing apparatus 1 as shown in FIG. The support member 60 includes a holding plate 62 at the lower end portion of the support cylinder 61, and a sliding bearing 63 is provided on the holding plate 62, and the distal end portion 20 a of the rotating cylinder 20 is supported by the sliding bearing 63. The support cylinder 61 is appropriately formed with a flow hole 61a through which the liquid L flows inside and outside. According to this configuration, the rotary cylinder 20 can be rotated more stably, and a desired fine bubble mixture can be easily generated.

  The manufacturing apparatus 1 shown in FIG. 1 is configured so that the fine bubble mixture can be continuously taken out from the branch pipe 53. However, as shown in FIG. It is possible to adopt a batch-type configuration that is not provided.

  In addition, the manufacturing apparatus 1 shown in FIG. 1 has the rotary cylinder 20 directly connected to the output shaft 11 of the drive motor 10, but as shown in FIG. You may connect via the shaft coupling 71. FIG. In this configuration, a base plate 73 is provided on the upper surface of the support plate 2 via a plurality of support columns 72, and a through hole 73 a through which the output shaft 11 is inserted is formed in the base plate 73, thereby driving on the base plate 73. A casing 12 of the motor 10 can be mounted. In order to maintain an airtight state inside the liquid storage tank 4, it is preferable that a through-hole seal 74 through which the rotary shaft 20 is inserted in an airtight manner is provided in the through hole 2 a of the support plate 2.

  According to the configuration shown in FIG. 5, by providing the shaft coupling 71, it is easy to cut the edge between the inside of the liquid storage tank 4 and the pipe 42, and the maintainability can be improved.

  In order to confirm the effect of the fine bubble mixture produced by the production apparatus 1, the following test was performed. FIG. 6 is an antibacterial test result of a microbubble mixed solution related to Bacillus subtilis, Escherichia coli, and Aspergillus niger that forms spores. As shown in FIG. 6, when a metal piece to which Bacillus subtilis, Escherichia coli, and Aspergillus niger were adhered was submerged for 1 minute using 2 liters of a fine bubble mixture, Bacillus subtilis almost died out. Escherichia coli and Aspergillus niger Was sterilized.

  The above test was carried out by mixing and immobilizing the artificial bone material titanium (diameter 5 mm x 10 mm) with Escherichia coli, Bacillus subtilis, and Aspergillus niger for 2 days. , The artificial bone material is rolled onto the LB agar medium for Bacillus subtilis, the 802 agar medium for Bacillus subtilis, and the potato agar medium is transferred to the potato agar medium. Was measured. Specifically, it was immersed in 2 liters of the bubble mixture sampled 3 hours after the production for 1 minute, washed 3 times with 5 ml of sterilized water, cultured in each medium, and antibacterial activity was measured. Moreover, the oxidation power was measured by the iodinated starch reaction method.

  As a result of measuring the oxidation activity by the iodinated starch reaction, it showed an ozone oxidation activity of about 0.35 ppm. However, this level of oxidation activity does not show antibacterial activity using Bacillus subtilis, Escherichia coli, and Aspergillus niger. It was also revealed that the peak of oxidative activity did not match the antibacterial activity.

  Moreover, the change of the dissolved oxygen amount when producing | generating a bubble liquid mixture is shown in FIG. Although active oxygen can be retained even if oxygen gas is bubbled as it is in blowing oxygen, it is more preferable that the amount of active oxygen produced is improved by using a bubble mixing nozzle.

  Further, the temperature in the liquid storage tank at the time of generating the bubble mixed liquid is desirably 10 ° C. or less, more preferably 8 ° C. or less.

  FIG. 8 shows the result of examining how much the retained oxygen retention effect of the bubble mixture is maintained by conducting a carbon dioxide gas blowing test on the generated bubble mixture using a gas-liquid mixing nozzle. The bubble mixture produced by this test had dissolved oxygen exceeding 60 ppm in about 15 minutes after the start of production, but the time required to expel dissolved oxygen using carbon dioxide gas was 1 hour or more. When carbon dioxide gas is blown directly without blowing oxygen gas, almost all dissolved oxygen disappears in about a few minutes, so once the bubble mixture is generated using oxygen gas, it is stably maintained. It revealed that.

  FIG. 9 shows the results of verifying how the dissolved oxygen concentration changes after the bubble mixture is generated. It became clear that the dissolved oxygen concentration was kept constant if the inside of the reservoir was maintained at 10 degrees or less after the bubble mixture was generated. The amount of dissolved oxygen was about 10 ppm in about 12 hours after generation, and the amount of dissolved oxygen tended to be higher than at the time of generation. Thereafter, the dissolved oxygen amount gradually decreases, but it takes more than one month to reach the saturated oxygen amount.

  In FIG. 6, when the metal fragment with microorganisms attached to the bubble mixture was treated for 1 minute, almost all the test bacteria were killed. FIG. 10 shows the results of verifying the antibacterial effect when the bubble mixture is placed in an ultrasonic humidifier and mist is generated. When Escherichia coli was used as a test strain, the antibacterial effect was revealed to be about 2 to 3 orders of magnitude higher. However, the antibacterial effect did not change much even when the mist treatment was performed for about 10 minutes to 1 hour.

  Similarly, the results of the antibacterial effect of the mist treatment when Aspergillus niger is used as a test bacterium are shown in FIG. In the case of Aspergillus niger, mist treatment effectively showed antibacterial activity, and it was revealed that colony formation was suppressed to about 4 to 5 digits.

From the test results so far, reactive oxygen species may be one of the factors in the antibacterial activity test against spores. In addition, it has been reported that the bactericidal effect of calcined calcium (calcium monoxide) is reduced by adding catalase and superoxide demutase (SOD), which are active oxygen scavenging enzymes. Therefore, the bactericidal effect of the calcined scallop shell powder slurry on Bacillus subtilis spores may be related to reactive oxygen species in addition to pH and calcium ions. As for reactive oxygen species, the bactericidal effect of hydrogen peroxide on spores has been studied.

   Thus, FIG. 12 shows the results of examining whether the antibacterial effect is reduced by adding catalase and superoxide demutase (SOD) to the bubble mixture. In FIG. 12, when tested using Bacillus subtilis that forms spores, it was revealed that both catalase and superoxide demutase suppress their antibacterial effects. From this result, it was shown that the antibacterial component of the bubble mixture is an active oxygen species.

  Similarly, the result of using Escherichia coli as a test strain is shown in FIG. In E. coli, catalase, which reduces the effect of hydrogen peroxide, suppressed more effectively than superoxide demutase. On the other hand, superoxide demutase seems to have increased antibacterial activity compared to the control bubble mixture. This result suggests that the reactive oxygen species that can provide antibacterial effects differ between Bacillus subtilis and Escherichia coli.

  FIG. 14 shows the relationship between reactive oxygen species and enzymes. As the active oxygen species, a superoxide radical is generated first, and then hydrogen peroxide is generated from the superoxide radical. Similarly, it has a cascade relationship in which hydroxy radicals are generated from hydrogen peroxide.

  FIG. 14 shows that superoxide demutase decomposes from superoxide radicals to hydrogen peroxide. Therefore, catalase and superoxide demutase (SOD) were mixed in the bubble mixture, and hydrogen peroxide was detected by an origination reaction by 4-aminoantipyrine spectrophotometry using the enzyme. The result is shown in FIG. A strong color reaction was observed when superoxide demutase was mixed in the bubble mixture. This result revealed that superoxide radicals were generated as the main component in the bubble mixture.

    In the test of FIG. 15, detection was performed using a 4-aminoantipyrine colorimetric method using a peroxidase enzyme. The reaction of 4-aminoassocipyrizo is used for the determination of hydrogen peroxide and can be detected with high sensitivity in an aqueous solution. Color reaction occurs when 4-aminoantipyrine coexists with peroxidase and hydrogen peroxide. In this test, gas-liquid mixed water, superoxide demutase (5 U / ml), peroxidase, and 4-aminoantipyrine were mixed, and color development was confirmed after 3 hours at room temperature. Thereafter, the result of the color development reaction continued for more than one month is shown in FIG. On the other hand, gas-liquid mixed water and superoxide demutase were not mixed, and no color reaction occurred with peroxidase and 4-aminoantipyrine alone. When the hydrogen peroxide concentration was converted based on the standard color chart, it was 5 mg /? (5 ppm) or more. These results showed that superoxide radicals corresponding to 5 ppm were contained in the gas-liquid mixed water for a long period of time.

  Active oxygen species such as superoxide radicals and hydroxy radicals are difficult to detect directly because of their high reactivity. Therefore, for detection, it was reacted with a coloring reagent or a luminescent reagent, and the concentration was calculated by changing the reagent. First, quantification was attempted using a xanthine oxidase enzyme having the ability to show the ability to generate superoxide radicals as an index. Specifically, a xanthine oxidase enzyme and hypoxanthine as a substrate are mixed to generate a superoxide radical by an enzymatic reaction, which reacts specifically with the superoxide radical, MPEC (2-methyl-6-p-methoxy) A calibration curve of a superoxide radical generation source was prepared by detecting with phenylethynylimidazopyranodine). On the other hand, the emission intensity when the bubble mixture and MPEC were mixed was measured and converted as the superoxide radical generation source ability.

The test method was a solution of each concentration of xanthine oxidase (Sigma, 2 Units / mg) enzyme in 0.1 M potassium phosphate solution (pH 7.5). The actual reaction concentration was 0.72 mM hypoxanthine (Hypoxanthine, Tokyo Chemical Industry Co., Ltd.), 300 μM MPEC, 0.1 M potassium phosphate solution (pH 7.5), and the above enzyme solution was added to carry out the luminescence reaction. The amount of luminescence was detected using a Turner Biosystems Luminometer Model TD-20 / 20 (Promega Corporation) to detect the accumulated luminescence intensity for 1 minute.

  One unit of xanthine oxidase converts about 1.0 μmole of xanthine into uric acid at pH 7.5, 25 ° C. for 1 minute, and when hypoxanthine is used as a substrate, about 50% of the activity is obtained. When the amount of superoxide radicals in the gas-liquid mixed water is converted, oxygen water has the ability to generate 25-40-nano mole / min superoxide radicals.

    Hydrogen peroxide is known to be generated in vivo when Superoxide is decomposed by SOD. Because it is a relatively stable active oxygen, it is long in the living body and diffuses widely. In this test, detection was performed using the luminescent reagent Luminol-HRP (Horse Radish Peroxidase). The amount of luminescence was detected by detecting the integrated luminescence intensity for 10 seconds using TD-20 / 20 (Promega Corporation).

  For the quantification of hydrogen peroxide, first, a hydrogen peroxide solution 25 mM imidazole-nitric acid solution (pH 7.0) was used for dilution. Each 100 μ? Enzyme solution and 100 μ? Luminescent reagent (Luminol 10 mg + HRP 100 unit / 100 mL 0.2 M boric acid solution pH 9.5) were mixed at a ratio of 1: 1 to detect the amount of luminescence. The amount of luminescence was detected using a Turner Biosystems Luminometer Model TD-20 / 20 (Promega Corporation) for 10 seconds.

  The amount of hydrogen peroxide containing oxygen water accumulated 0.06-mg /? Or more immediately after generation. On the other hand, it decayed to 4.2 μg / liter one day after the generation of oxygen water, but the concentration of 3.9 μg / liter was maintained after 40 days.

    Hydroxy radicals are radicals generated by the reaction (Fenton reaction) of hydrogen peroxide with metal ions such as iron Fe 2+ and copper Cu + in the living body, and are the most reactive and short-lived among oxygen radicals. It is also directly involved in lipid peroxidation and gene damage. It is known that hydroxy radicals can be generated at the experimental level by the Fenton reaction, and that hydroxy radicals can be detected using Luminol. Therefore, we tried to detect the emission of hydroxy radicals with Luminol using this generation system.

  Specifically, solution 1 prepared by adjusting iron sulfate to 1 mM with distilled water, and DTPA (Diethylenetriamine-N, N, N ', N ", N" -pentaacetic acid, Tokyo Chemical Industry Co., Ltd.) as distilled water Solution 2 adjusted to 1 mM with 1 mM Luminol (Kishida was dissolved in 0.2 M boric acid solution pH 9.5 to prepare Solution 3 respectively. Iron sulfate solution: Chelating agent: Luminescent reagent = 1: 1: 1 100 μl of the mixed solution 4 and gas-liquid mixed water are mixed one by one to generate hydroxy radicals. At the same time as dispensing, TD-20 / 20 (Promega Corporation) is used to detect the accumulated luminescence intensity for 10 seconds. The calibration curve was prepared by diluting with a hydrogen peroxide solution 25 mM imidazole-nitric acid solution (pH 7.0) to prepare a solution with different hydrogen peroxide concentrations, and mixing 100 μl each with solution 4 in the same manner. Shows the relationship between emission intensity and hydroxy radical generation by generating radicals and emitting light. It created the amount line.

  Hydroxy radicals containing oxygen water reached 0.053 μg / ml after 1 day of production, but disappeared after 30 days of production. Hydroxy radicals were highly reactive and disappeared instantaneously, indicating that they were retained in an aqueous solution for more than 1 day after generation.

  NTA (Nano Tracking Analysis) technology enables real-time observation of Brownian motion of nanoparticles in liquid. In addition, by measuring the speed of Brownian motion using dedicated software, a particle size distribution graph of particle size and number can be obtained. Therefore, using NanoSight (Nanosite Japan Quantum Design Co., Ltd.), various ultrafine bubble water and oxygen water bubble particle sizes were compared.

Visualization was performed by injecting the sample solution into the nanosite device, setting the module in the analyzer, and irradiating the laser. First, the laser is irradiated in the horizontal direction, and the side scattered light from the nanoparticles is visualized with an objective lens. This device visualizes the Brownian motion of nanoparticles. The particles moving in the liquid were automatically recognized and tracking analysis was performed. The movement trajectory was displayed with a red line for each particle, and the velocity was simultaneously analyzed for all the particles recognized on the screen. In the real-time display of the particle size distribution graph, the size of each particle is calculated from the moving speed of the particle by the Stokes-Einstein equation, and the result is shown in FIG.

  It is clear that the smaller the average particle size, the more stable it exists in water. Compared to UFB manufactured by S, I, and N, which have a proven track record as ultrafine bubble water, Nisshin Giken's oxygen water has a small average particle diameter of 71.8 nm. The minimum value was also measured for the standard deviation. This supported the long-term storage of active oxygen.

  In addition, the larger the number of bubbles contained, the higher the performance of UFB. FIG. 17 shows the measurement result of the number of bubbles using the nanosite. It became clear that the number of bubbles contained in oxygen water has the world's top class bubble density.

  Ultra fine bubbles have a colloidal side, and the particle surface is negatively charged. For this reason, fine bubbles repel each other. Because of this property, there is no bonding between fine bubbles, the bubble concentration does not decrease, and it exists stably for a long time. Therefore, the zeta potential of bubbles contained in oxygen water was measured using ELSZ-2000ZS (Otsuka Electronics), a zeta potential / particle size / molecular weight measurement system. The result is shown in FIG.

  When the zeta potential of oxygen water after 7 days, 96 days, 146 days, and 483 days from the generation was measured, an average zeta potential of −37.06 mV was measured even after 7 days. On the other hand, the average zeta potential of oxygen water from 96 days after generation to 483 days after generation also showed a high numerical value of −20 mV or less. As a result, it was revealed that the contained bubbles existed stably even after 1 year or more.

Since it became clear that bubbles exist stably in oxygen water that has been stored for a year or more, it reacts specifically with superoxide radicals to detect whether active oxygen superoxide radicals are also stable. The detection was attempted with the luminescent reagent MPEC (2-methyl-6-p-methoxyphenylethynylimidazopyranodine). The result is shown in FIG.

  In FIG. 19, after the oxygen water was generated, the content of superoxide radicals gradually decreased with time. However, the rate of decrease is very low, reaching the half-life of the amount of superoxide radicals at one year after the generation of oxygen water, and the active oxygen contained in oxygen water is stably maintained for a long time. It became clear.

DESCRIPTION OF SYMBOLS 1 Manufacturing apparatus 4 Liquid storage tank 6 Bubble supply apparatus 10 Drive motor 20 Rotating cylinder 22 Ejection part 24a Rotating blade 30 Outer cylinder 40 Circulating apparatus 50 Gas-liquid mixing part 60 Support member L Liquid

Claims (1)

  A fine bubble mixed liquid having a bactericidal effect, in which fine bubbles having a diameter of 60 to 200 nm or less and containing active oxygen species are mixed therein.