JP5700620B2 - Method for controlling the methanogenic activity of hydrogen-utilizing methane bacteria - Google Patents

Description

The present invention relates to a method for controlling the methanogenic activity of hydrogen-utilizing methane bacteria. More specifically, the present invention is suitable for controlling the methanogenic activity of a hydrogen-assimilating methane bacterium that is a microorganism that produces methane (CH ₄ ) from hydrogen (H ₂ ) and carbon dioxide (CO ₂ ). Regarding the method.

  In recent years, high-efficiency material conversion technology utilizing the functions of living organisms has begun to attract attention for the construction of a resource recycling society, and various studies are being promoted. As such a representative technique, a technique for methane fermentation treatment of organic waste such as garbage is known. The methane fermentation treatment is a method in which organic waste is put into a fermentation broth and fermented under anaerobic conditions to convert the organic waste into biogas containing methane gas. Since methane fermentation treatment can significantly reduce the volume of organic waste, it can solve the recent problem of tightness in landfill sites, and is a highly useful technology that can recover methane gas as energy.

By the way, it is known that the fermented liquid used for methane fermentation treatment contains hydrogen-utilizing methane bacteria as methane bacteria responsible for methane production. A hydrogen-utilizing methane bacterium is a methane bacterium that has a respiration mode that grows by acquiring energy in the process of generating methane (CH ₄ ) from hydrogen (H ₂ ) and carbon dioxide (CO ₂ ), and the methane fermentation process. It plays an important role in producing methane as the final product in the final stage.

In addition, the hydrogen-assimilating methane bacterium uses a function capable of generating methane (CH ₄ ) from hydrogen (H ₂ ) and carbon dioxide (CO ₂ ) to convert carbon dioxide, which is a global warming gas, into methane gas. It can also be used to convert energy resources. For example, in Patent Document 1, carbon dioxide in exhaust gas is dissolved in water or an alkaline solution, and dissolved oxygen in the obtained carbon dioxide solution is reduced in concentration, and then hydrogen-assimilating methane fermentation is performed using this as a substrate. As a result, carbon dioxide in exhaust gas is recovered and reduced, and this is converted into methane useful as a fuel source, thereby recovering methane effective as an energy resource while protecting the global environment.

  As described above, hydrogen-utilizing methane bacteria have extremely excellent functions and are considered to be extremely useful microorganisms in the industry.

JP 2004-321857 A

  To efficiently implement methane production by hydrogen-utilizing methane bacteria, establish a method for efficiently growing hydrogen-utilizing methane bacteria and a method for enhancing the methane-producing activity of hydrogen-utilizing methane bacteria themselves. Is important. However, until now, these techniques have not been established.

  That is, in order to grow hydrogen-utilizing methane bacteria, it is necessary to adjust the redox potential to a low level by chemical adjustment such as adding a reducing agent such as cysteine or sodium sulfide to the culture solution. In the method, the growth rate of hydrogen-utilizing methane bacteria could not be made sufficiently fast, and it was difficult to say that hydrogen-utilizing methane bacteria could grow efficiently.

  Moreover, until now, no method has been disclosed for increasing the methane production activity of hydrogen-utilizing methane bacteria.

  By the way, methane gas produced by hydrogen-assimilating methane bacteria is known as a gas having a global warming effect about 20 times that of carbon dioxide. Hydrogen-utilizing methane bacteria are widely inhabited on the earth. For example, in remote areas, even if methane gas is recovered as an energy resource, there is a cost for transporting methane gas. There is also a request to suppress the generation of methane gas. In addition, when the supply amount of methane gas becomes excessive in the future, it may be possible to request to suppress the generation of methane gas as a warming gas without collecting the methane gas. From such a viewpoint, it is necessary to establish not only a method for increasing the methane producing activity of hydrogen-utilizing methane bacteria but also a method for reducing the methane producing activity of hydrogen-utilizing methane bacteria.

  Therefore, an object of the present invention is to provide a technique that can enhance the methane producing activity of the hydrogen-assimilating methane bacterium itself and can promote the growth of the hydrogen-assimilating methane bacterium.

  Another object of the present invention is to provide a technique that can reduce the methane production activity of the hydrogen-assimilating methane bacterium itself.

In order to solve such a problem, the present inventor has made various studies on a culture solution containing hydrogen-utilizing methane bacteria at a cell density at a stationary stage growth stage. That is, when hydrogen-utilizing methane bacteria are included at the cell density at the stationary stage, no further increase in cell density can occur. If the effect is confirmed, this effect can be said to be an effect exhibited by the improvement or reduction of the methane production activity of the hydrogen-assimilating methane bacterium itself. As a result, the methane production activity per individual hydrogen-utilizing methane bacterium is controlled by bringing the electrode into contact with the culture solution and controlling the oxidation-reduction potential of the culture solution within a certain range by applying a potential to this electrode. It was found that control to increase or decrease is possible.

Further, the inventors of the present application have conducted intensive studies and brought the electrode into contact with a culture solution containing hydrogen-utilizing methane bacteria at the cell density at the growth stage before the logarithmic growth phase, and cultivated by applying a potential to the electrode. It was also found that the growth of hydrogen-utilizing methane bacteria can be promoted by controlling the redox potential of the liquid to a potential that can increase the methane production activity per individual hydrogen-assimilating methane bacterium.

Based on these findings, the inventor of the present application uses, as a reference potential, a redox potential at which a hydrogen-assimilating methane bacterium that can be formed in the environment by an inherent or added chemical component can be grown. The electrode is brought into contact with an environment containing, and an electric potential is applied to the electrode so that the environmental oxidation-reduction potential is larger than -0.8 V including -0.8 V on the silver / silver chloride electrode potential reference. By controlling, the methane production activity per individual hydrogen-assimilating methane bacterium can be made higher than the methane production activity per individual hydrogen-assimilable methane bacterium at the reference potential. by controlling the redox potential of such increases on the positive side than the reference potential, hydrogen-utilizing methane bacteria per individual methanogenic active hydrogen-utilizing methane bacteria 1 at reference potential individuals skilled Than Ri methanogenic activity led to the finding that derived that can be reduced, thereby completing the present invention.

That is, the method for controlling the methanogenic activity of the hydrogen-utilizing methane bacterium of the present invention is based on the oxidation-reduction potential at which the hydrogen-assimilating methane bacterium that can be formed in the environment by the inherent or added chemical components can grow. The electrode is brought into contact with an environment containing hydrogen-utilizing methane bacteria, and an electric potential is applied to the electrode so that the environmental redox potential is -0.8 V including -0.8 V on the basis of the silver / silver chloride electrode potential. Is controlled to be larger on the negative side, so that the methane production activity per individual hydrogen-utilizing methane bacterium is higher than the methane production activity per individual hydrogen-utilizing methane bacterium at the reference potential. . The environmental redox potential is preferably -0.8 V based on the silver / silver chloride electrode potential. Moreover, it is preferable that the environment which contacts an electrode has reference electric potential.

Next, the method for controlling the methanogenic activity of the hydrogen-assimilating methane bacterium of the present invention is based on the oxidation-reduction potential at which the hydrogen-assimilating methane bacterium that can be formed in the environment by chemical components that are inherent or added can grow. The potential is controlled so that the electrode is brought into contact with an environment containing a hydrogen-utilizing methane bacterium and having a reference potential, and the potential is applied to the electrode so that the oxidation-reduction potential of the environment becomes larger on the plus side than the reference potential. Thus, the methane producing activity per individual hydrogen-utilizing methane bacterium is made lower than the methane producing activity per individual hydrogen-utilizing methane bacterium at the reference potential. In this case, it is preferable that the oxidation-reduction potential of the environment in this case is specifically set to a potential greater than -0.2V including -0.2V on the basis of the silver / silver chloride electrode potential.

  Here, in the method for controlling the methane production activity of the hydrogen-assimilating methane bacterium of the present invention, it is preferable that the hydrogen-assimilating methane bacterium is contained in the environment at a cell density corresponding to a stationary stage growth stage.

According to the present invention, since the methane production activity per individual hydrogen-utilizing methane bacterium can be controlled to be higher than the methane production activity per individual hydrogen-utilizing methane bacterium at the reference potential, It becomes possible to enhance the methane production activity of the entire environment by increasing the methane production activity of each cell of the existing hydrogen-utilizing methane bacteria group. In addition, when the cell density of hydrogen-utilizing methane bacteria present in the environment is smaller than the cell density at the stationary stage growth stage, the growth rate is accelerated over the growth rate at the reference potential. It is also possible.

Further, according to the present invention, since the methane production activity per individual hydrogen-utilizing methane bacterium can be controlled to be lower than the methane production activity per individual hydrogen-utilizing methane bacterium at the reference potential, It becomes possible to reduce the methanogenic activity of each cell of the hydrogen-assimilating methanogen group present in the environment, and greatly reduce the methanogenic activity of the entire environment.

It is a figure which shows the time-dependent change of the methane production amount in the various setting electric potential in Example 1. FIG. It is a figure which shows the number of microbes of the culture solution in the various setting electric potential in Example 2. FIG. It is a figure which shows the amount of methane production | generation in the various setting potential in Example 2. FIG. It is a figure which shows the structure of the experimental apparatus used in the Example. It is sectional drawing which shows an example of the activation control apparatus concerning 1st embodiment A. It is sectional drawing which shows an example of the activation control apparatus concerning 1st Embodiment B. It is sectional drawing which shows an example of the activation control apparatus concerning 1st Embodiment C. It is sectional drawing which shows an example of the activation control apparatus concerning 1st Embodiment D. It is sectional drawing which shows an example of the activation control apparatus concerning 2nd embodiment. It is sectional drawing which shows an example of the other form of an active control apparatus.

  Hereinafter, embodiments for carrying out the present invention will be described in detail with reference to the drawings.

  The method for controlling the methane production activity of the hydrogen-assimilating methane bacterium according to the present invention uses, as a reference potential, a redox potential at which the hydrogen-assimilating methane bacterium that can be formed in the environment by an inherent or added chemical component can grow. An electrode is brought into contact with an environment containing hydrogen-utilizing methane bacteria, and an electric potential is applied to the electrode to reduce the environmental redox potential to -0.8 V including -0.8 V on the basis of the silver / silver chloride electrode potential. By controlling so that it may become large in the side, the methanogenic activity of the hydrogen-assimilating methane bacterium is made higher than that of the hydrogen-assimilating methane bacterium at the reference potential.

  In addition, the method for controlling the methanogenic activity of the hydrogen-utilizing methane bacterium of the present invention uses the oxidation-reduction potential at which the hydrogen-assimilating methane bacterium that can be formed in the environment by the inherent or added chemical components can grow as a reference potential. The electrode is brought into contact with an environment containing a hydrogen-utilizing methane bacterium and having a reference potential, and the potential is applied to the electrode to control the environmental redox potential to be larger than the reference potential. The methane producing activity of the hydrogen assimilating methane bacterium is made lower than the methane producing activity of the hydrogen assimilating methane bacterium at the reference potential.

  That is, in the present invention, the oxidation-reduction potential at which a hydrogen-assimilating methane bacterium that can be formed in the environment by an inherent or added chemical component can be used as a reference potential, and is in contact with the environment containing the hydrogen-assimilating methane bacterium. By controlling the oxidation-reduction potential of the environment so that it includes -0.8 V and becomes larger than -0.8 V on the basis of the silver / silver chloride electrode potential, the hydrogen-assimilating methane bacteria The methanogenic activity of the hydrogen-utilizing methane bacterium at the reference potential is higher than that of the hydrogen-utilizing methane bacterium, and the oxidation-reduction potential of the environment containing the hydrogen-assimilating methane bacterium is electrically controlled to a larger potential on the plus side than the reference potential. By doing so, the methane producing activity of the hydrogen assimilating methane bacterium is made lower than the methane producing activity of the hydrogen assimilating methane bacterium at the reference potential.

Thus, the example which reported that control which raises or reduces the methane production activity of the cell body of hydrogen-utilizing methane bacterium itself can be made, and it has been clarified for the first time in this application. The methane generation activity means the ability to generate methane (CH ₄ ) from hydrogen (H ₂ ) and carbon dioxide (CO ₂ ). If the methane generation activity increases, hydrogen (H ₂ ) and carbon dioxide (CO ₂₎ increases the speed of generating methane (CH ₄₎ from the rate of generating methane (CH ₄₎ from when decreased methanogenic activity and hydrogen _{(H 2)} carbon dioxide (CO ₂₎ is reduced .

  Hereinafter, details of the method of the present invention will be specifically described.

The “reference potential” in the present invention is an oxidation-reduction potential at which hydrogen-utilizing methane bacteria that can be formed in the environment by chemical components that are inherent or added can grow. Since hydrogen-utilizing methane bacteria are microorganisms that perform anaerobic respiration, it is said that they do not grow unless they are in a low redox potential environment, for example, a redox potential environment that is larger than ΔE ′ ₀ = −0.33 V on the minus side. ing. In an environment where hydrogen-utilizing methane bacteria are growing in nature, a low oxidation-reduction potential at which hydrogen-utilizing methane bacteria can grow is formed by chemical components inherent in the environment. In addition, a low redox potential at which the hydrogen-assimilating methane bacterium can grow is also formed in the methane fermentation broth containing the hydrogen-assimilating methane bacterium. When hydrogen-utilizing methane bacteria are grown in an environment in which hydrogen-utilizing methane bacteria cannot grow (including culture media, etc.), the hydrogen It is controlled to the redox potential that allows methanogenic methane bacteria to grow. For example, it is possible to control the oxidation-reduction potential at which hydrogen-utilizing methane bacteria can grow by adding bubbling of hydrogen, bubbling of an inert gas such as nitrogen, or addition of a reducing agent such as cysteine or sodium sulfide.

  In other words, the oxidation-reduction potential at which a hydrogen-assimilating methane bacterium that can be formed in the environment by chemical components that are inherent or added is the potential of the hydrogen-assimilating methane bacterium already growing by chemical components that are inherent in the environment. The oxidation-reduction potential of an environment controlled to a viable oxidation-reduction potential, hydrogen-assimilating methane bacteria cannot grow only with chemical components that are inherent in the environment. It means the oxidation-reduction potential of the environment controlled to a viable oxidation-reduction potential.

  In the present invention, the oxidation-reduction potential at which a hydrogen-assimilating methane bacterium that can be formed in the environment by an inherent or added chemical component can be grown as a reference potential, and the methane producing activity of the hydrogen-assimilating methane bacterium at the reference potential Based on the above, by controlling the redox potential of the environment containing hydrogen-utilizing methane bacteria electrically, the methane production activity of the hydrogen-utilizing methane bacteria is controlled to be increased or decreased.

  In the method of the present invention for enhancing the methane production activity, the oxidation-reduction potential of the environment containing hydrogen-utilizing methane bacteria is not limited. That is, even in an environment having an oxidation-reduction potential at which hydrogen-utilizing methane bacteria cannot grow, the method of the present invention can increase methane production activity and further promote its growth.

  In the method of the present invention for reducing the methane production activity, the oxidation-reduction potential of the environment containing hydrogen-utilizing methane bacteria is limited to the oxidation-reduction potential at which hydrogen-utilizing methane bacteria can grow. That is, in an environment having a redox potential where hydrogen-utilizing methane bacteria cannot grow, hydrogen-utilizing methane bacteria cannot live, so the method of the present invention in the case of reducing methane production activity is applied. There is no significance. However, in an environment having an oxidation-reduction potential at which hydrogen-utilizing methane bacteria can grow, the methane producing activity of hydrogen-utilizing methane bacteria can be easily reduced by the method of the present invention.

  The “environment” in the present invention includes not only soil, groundwater, sludge, methane fermentation broth, but also a culture medium such as a culture solution in which hydrogen-utilizing methane bacteria live or can live. The “environment” preferably contains a lot of moisture. In this case, it becomes easy to improve the controllability of the environmental oxidation-reduction potential by the electrode, and the effect of the present invention can be easily obtained. Moreover, you may make it add additives, such as a nutrient source of hydrogen utilization methane bacteria, to an environment as needed.

The hydrogen-assimilating methane bacterium to which the method of the present invention can be applied is not particularly limited, and has any breathing mode that generates methane (CH ₄ ) from hydrogen (H ₂ ) and carbon dioxide (CO ₂ ). Applicable to hydrogen-utilizing methane bacteria. For example, but not limited to Methanothermobacter thermautotrophicus. In addition, since hydrogen (H ₂ ) and carbon dioxide (CO ₂ ) are necessary for hydrogen-utilizing methane bacteria to produce methane, these gases are not present in the environment or are insufficient. If it is, it is necessary to supply appropriately. Note that the supply ratio of hydrogen (H ₂ ) and carbon dioxide (CO ₂ ) is preferably about 4: 1 in terms of molar ratio.

  In the method of the present invention for enhancing the methane production activity, an electric potential is applied to the electrode, and the environmental redox potential is -0.8 V including -0.8 V on the basis of the silver / silver chloride electrode potential, and is more negative than -0.8 V. Therefore, the methane producing activity of the hydrogen assimilating methane bacterium is made higher than the methane producing activity of the hydrogen assimilating methane bacterium at the reference potential. Here, if the potential of the electrode is excessively increased to the negative side, water is electrolyzed vigorously and electric energy input to an unintended reaction is consumed. Moreover, sufficient improvement effect of methane production activity can be obtained even at −0.8V. Therefore, the oxidation-reduction potential of the environment is more preferably −0.8 V to −1.4 V, and further preferably −0.8 V.

  In the method of the present invention in the case of reducing the methane production activity, the hydrogen-assimilating methane bacterium is controlled by applying a potential to the electrode and controlling the oxidation-reduction potential of the environment to be larger on the plus side than the reference potential. The methanogenic activity of methane is higher than the methanogenic activity of hydrogen-utilizing methane bacteria at the reference potential. Specifically, the environmental oxidation-reduction potential may be set to a larger potential ((environmental oxidation-reduction potential)> − 0.45 V) than −0.45 V on the basis of the silver / silver chloride electrode potential. It is preferable to set a large potential on the plus side including -0.2V. However, if the potential of the electrode is increased too much on the plus side, the input electric energy is wasted, and it is more preferable to set the voltage to -0.2V to + 0.2V.

  According to the method of the present invention for enhancing the methane production activity, the growth rate of hydrogen-utilizing methane bacteria in the environment is increased more than the growth rate at the reference potential, so that the hydrogen-utilizing methane bacteria can be more efficiently propagated. And can be used for methane production. Moreover, the methanogenic activity of the hydrogen-assimilating methane bacterium can be made higher than the methanogenic activity of the hydrogen-assimilating methane bacterium at the reference potential. Even if it is contained in the environment at a corresponding cell density, it can be obtained. Therefore, even in an environment where hydrogen-utilizing methane bacteria are contained at an extremely high density, the methane-producing activity of the hydrogen-assimilating methane bacteria is increased and the methane-producing ability of the environment as a whole is extremely excellent. be able to.

  Further, according to the method of the present invention in the case of reducing the methane producing activity, the methane producing activity of the hydrogen assimilating methane bacterium can be lowered than the methane producing activity of the hydrogen assimilating methane bacterium at the reference potential, The effect can be obtained even when hydrogen-utilizing methane bacteria are contained in the environment at a cell density corresponding to the stationary stage growth stage. Therefore, by applying the present invention to an environment in which hydrogen-utilizing methane bacteria are contained at a cell density corresponding to the growth stage in the stationary phase, the methane producing activity of the hydrogen-utilizing methane bacteria is reduced. Thus, the methane production capacity of the environment as a whole can be greatly reduced, and in some cases, hydrogen-utilizing methane bacteria can be killed. By using this method, it becomes possible to meet the demand for suppressing the generation of methane gas as a warming gas. For example, if the method of the present invention is carried out by bringing an electrode into contact with an aqueous environment, the methane production activity of the hydrogen-assimilating methane bacterium can be reduced, and the methane production capacity of the entire environment can be greatly reduced. Even in an environment that is not an aqueous environment, if the method of the present invention is carried out by bringing the electrode into contact with water to such an extent that the oxidation-reduction potential can be controlled, the methanogenic activity of the hydrogen-assimilating methane bacterium can be increased. This can significantly reduce the overall methane production capacity of the environment.

  Below, about an example of embodiment of the method of this invention in the case of improving methane production activity, 1st embodiment at the time of using environment as a culture solution is described based on FIGS. A form is demonstrated based on FIG.

<First embodiment>
In the method of the present invention according to the first embodiment, the working electrode, the counter electrode, and the reference electrode are connected to a constant potential setting device, the culture solution and the electrolyte solution are brought into contact with each other through an ion exchange membrane, and the culture solution is contacted. By bringing the working electrode and the reference electrode into contact, bringing the counter electrode into contact with the electrolytic solution, and controlling the potential of the working electrode in a three-electrode system, the oxidation-reduction potential of the culture solution is increased to the minus side of the reference potential. I try to control it.

  The method of the present invention according to the first embodiment is performed by, for example, the activity control apparatus 1 shown in FIGS. That is, in the activity control apparatus 1 shown in FIGS. 5 to 8, one of the two tanks partitioned by the ion exchange membrane 6 is an activity control tank 7, and the other tank is a counter electrode tank 8. The control tank 7 contains the culture solution 4 containing hydrogen-utilizing methane bacteria, and the working electrode 9 and the reference electrode 11 are immersed therein. The counter electrode tank 8 contains the electrolyte solution 4a and the counter electrode 10 The working electrode 9 and the counter electrode 10 are immersed in the constant potential setting device 12 and the potential of the working electrode 9 is controlled by a three-electrode system so that the oxidation-reduction potential of the culture solution 4 is set to a minus side with respect to the reference potential. Control is performed so as to increase.

  In this way, by controlling the potential of the working electrode 9 by the three-electrode method, the potential of the working electrode 9 can be strictly controlled to the set potential. Specifically, a potential difference between the working electrode 9 and the reference electrode 11 is measured by a constant potential setting device (potentiostat) 12, and the working electrode 9 and the counter electrode 10 are adjusted so that the potential difference reaches the set potential. A current is passed between them so that no current flows through the reference electrode 11 serving as a reference. The details of the potential control by the three-electrode method are described in, for example, pages 6 to 9 of Electrochemical Measurement Method (above), Technical Bulletin Publishing Co., Ltd., 1st edition 15 printing, published in June 2004. ing. However, when the potential of the working electrode 9 can be controlled only by the voltage between the working electrode 9 and the counter electrode 10, the three-electrode system may not be used.

  Moreover, in the activity control apparatus 1 shown in FIGS. 5-8, the biogas containing the methane gas which retains in the space (head space) higher than the liquid level of the culture solution 4 in the activity control tank 7 of the activity control tank 7 is shown. A gas exhaust pipe 15a that leads to the outside (outside of the activity control device 1) is provided, and the biogas in the activity control tank 7 is recovered by the gas recovery means 15 that can be opened and closed by a valve 15b. ing. However, the biogas recovery method is not limited to this method. For example, without providing the gas recovery means 15, an opening is provided in the upper part of the activation control tank 7, and this opening is closed with an elastic material such as synthetic rubber (for example, silicone rubber). You may make it collect | recover biogas from a head space by inserting an injection needle. The elastic material such as synthetic rubber closes the hole when the injection needle is pulled out. Therefore, when the biogas is not collected, the biogas does not leak from the activity control tank 7 even if the injection needle is pulled out.

  Furthermore, in the activity control apparatus 1 shown in FIGS. 5 to 8, the culture solution 4 in the activity control tank 7 is guided out of the activity control tank 7 below the liquid level of the culture solution 4 in the activity control tank 7. The culture medium 4 is collected from the inside of the activity control tank 7 by a culture medium collecting means 16 provided with a culture medium discharge pipe 16a and opening and closing the culture medium discharge pipe 16a by a valve 16b. However, the method for collecting the culture solution 4 is not limited to this method. For example, without providing the culture solution collecting means 16, an opening may be provided in the activity control tank 7, closed with an elastic material such as synthetic rubber, and the culture solution 4 may be collected by inserting a syringe needle. Alternatively, it may be connected to a syringe at one end of a tube having both ends opened, and the other end is immersed in the culture solution 4 to collect the culture solution 4 through the tube. Also in these cases, biogas does not leak from the activity control tank 7.

  In addition to the gas recovery means 15 and the culture medium collection means 16, means for adding and supplying substances to the culture medium 4 may be provided. Specifically, an openable / closable substance introduction tube that can add and supply substances to the culture solution 4 from the outside of the activity control tank 7 may be provided. In this case, substances such as nutrients and neutralizing agents can be added to the culture solution 4 as necessary. Moreover, in order to maintain an anaerobic environment, gas can also be supplied and the carbon dioxide and hydrogen which are required for hydrogen utilization methane bacterium can also be supplied. However, the means for adding and supplying the substance to the culture solution 4 is not necessarily provided, and the gas recovery means 15 and the culture solution collecting means 16 may be used together as means for adding and supplying the substance to the culture solution 4. . Further, as described above, the injection needle of the syringe may be inserted into the elastic material, and the substance may be added to and supplied to the culture solution 4.

  Hereinafter, the case where the activation control device shown in FIG. 5 is used will be described as a first embodiment A, and the case where the activation control device shown in FIG. 6 is used will be described as a first embodiment B, which is shown in FIG. A case where the activation control device is used will be described as a first embodiment C, and a case where the activation control device shown in FIG. 8 is used will be described as a first embodiment D.

(First embodiment A)
In the activity control apparatus 1 shown in FIG. 5, a sealed container 20 is used as an activity control tank 7, a sealed container 21 that can be accommodated in the container 20 is used as a counter electrode tank 8, and the small container 21 is at least partially ionized. In addition to the exchange membrane 6, a gas discharge pipe 22 that discharges gas (gas generated from the counter electrode 10) out of the container 20 is provided. In the activation control device 1 shown in FIG. 5, the wiring connecting the counter electrode 10 and the constant potential setting device 12 passes through the gas exhaust pipe 22, but is not necessarily limited to this configuration. May be connected to the constant potential setting device 12 without passing through the gas discharge pipe 22.

  Therefore, according to the activity control device 1 shown in FIG. 5, biogas does not leak from the activity control tank 7. In addition, since the gas generated from the counter electrode tank 8 does not leak into the activation control tank 7, the gas generated from the counter electrode tank 8 is mixed into the biogas to reduce the methane concentration of the biogas, The gas generated from the tank 8 does not dissolve in the culture solution 4 and does not adversely affect the growth and function of the hydrogen-utilizing methane bacteria. Furthermore, since the activation control tank 7 has a sealed structure, there is an advantage that the activation control tank 7 can be easily controlled in an anaerobic environment. Further, since the gas generated from the counter electrode tank 8 can be discharged from a desired position, it can be recovered and reused in some cases.

  Further, by accommodating the small container 21 in the container 20, the small container 21 is immersed in the culture solution 4 accommodated in the container 20, and the ion exchange membrane 6 provided in at least a part of the small container 21 is cultured. Contact with liquid 4. In other words, the culture solution 4 comes into contact with the electrolyte solution 4 a through the ion exchange membrane 6.

  The sealed container 20 as the activity control tank 7 is a container having a size capable of accommodating the small container 21 with the sealed structure as the counter electrode tank 8, and the shape is not particularly limited. Examples of the material of the container include, but are not limited to, glass, plastic, an insulating metal, concrete, and the like. Further, a container in which a gas-impermeable film material is formed into a bag shape by heat sealing or the like may be used as the activity control tank 7.

  The small container 21 having a sealed structure as the counter electrode tank 8 is a container of a size that can be accommodated in the container 20 as the activity control tank 7, and includes the ion exchange membrane 6 at least in part. Here, the small container 21 may be a bag-like container formed entirely by the ion-exchange membrane 6, but only one side of the bag-like container may be constituted by the ion-exchange membrane 6, or a part of one surface may be further formed. You may make it comprise only the ion exchange membrane 6. FIG. When the ion exchange membrane 6 is partially used, other portions may be made of the same material as that of the container 20, or may be made of a membrane material other than the ion exchange membrane 6, such as a gas-impermeable membrane material. It may be configured so that gas from the small container 21 (gas generated from the counter electrode tank 8) does not leak into the container 20.

  The electrolyte solution 4a accommodated in the counter electrode tank 8 may include, for example, sodium ions or potassium ions. Since the culture solution 4 usually contains sodium ions, potassium ions, etc., the culture solution 4 can also be used as the electrolyte solution 4a.

  As the working electrode 9 and the counter electrode 10, for example, a conductive material such as a carbon plate can be used as appropriate. In the counter electrode 10, a reaction that complements the exchange of electrons with respect to the oxidation-reduction reaction in the working electrode 9 proceeds.

  The temperature of the activity control tank 7 (the temperature of the culture solution 4) is appropriately controlled to a temperature suitable for the growth of the hydrogen-assimilating methane bacteria that are the targets for activity control. For example, when a thermophilic hydrogen-assimilating methane bacterium such as Methanothermobacter thermautotrophicus is the target of activity control, it is preferably 40 ° C to 70 ° C, more preferably 50 ° C to 60 ° C. It is more preferable that the temperature is 55 ° C.

  In the present invention, by controlling the potential of the working electrode 9, the oxidation-reduction potential of the culture solution 4 is controlled to a large negative value including −0.8 V on the basis of the silver / silver chloride electrode potential. In addition, it is preferable to control the oxidation-reduction potential of the culture solution 4 to −0.8 V to −1.4 V, and it is more preferable to control to −0.8 V.

  Here, by adding the redox substance 3 to the culture solution 4, the controllability of the potential of the culture solution 4 can be improved, and the oxidation-reduction potential of the culture solution 4 can be brought close to or coincident with the potential of the working electrode 9. . And by providing the ion exchange membrane 6, permeation | transmission to the electrolyte solution 4a of the oxidation-reduction substance 3 contained in the culture solution 4 can be prevented. For example, by using a Nafion membrane (manufactured by DuPont) that is a membrane that transmits only monovalent cations as the ion exchange membrane 6, when the redox material 3 is iron ions, divalent iron ions or three Since the valent iron ions do not permeate the ion exchange membrane 6, they can remain in the culture solution 4 without allowing the redox material to permeate the electrolyte solution 4a. Therefore, when the potential of the working electrode 9 is controlled, the concentration ratio of the oxidized form and reduced form of the redox substance 3 in the culture solution 4 changes accordingly, and the followability of the potential of the culture solution 4 by the potential of the working electrode 9 is changed. Will improve.

  As the redox substance 3, a substance that can reversibly undergo a redox reaction with the working electrode 9 immersed in the culture solution 4 and that does not exhibit toxicity to hydrogen-utilizing methane bacteria is used. Can do. For example, a general iron ion is mentioned as a soil component as mentioned above. Here, in order to allow iron ions to stably exist in the culture solution 4, it is preferable that iron ions are coordinated with the chelating agent and added to the culture solution 4. As the chelating agent, any chelating agent capable of coordinating iron ions can be used. For example, diethylenetriaminepentaacetic acid (DTPA), ethylenediaminetetraacetic acid (EDTA), tetraethylenetriamine (TET), ethylenediamine (EDA), diethylenetriamine (DETA), citric acid, oxalic acid, crown ether, nitrilotetraacetic acid, disodium edetate, sodium edetate, trisodium edetate, penicillamine, trisodium pentetate calcium, pentetate, succil and edet Mention may be made of acid trientine. In addition to iron ions, quinone compounds such as potassium ferrocyanide and sodium anthraquinone disulfonate, and methyl viologen can be used. These substances also reversibly change into an oxidized form and a reduced form by an oxidation-reduction reaction. In particular, a quinone compound is a substance known as one of the soil components and is preferable. That is, by adding the soil itself to the culture solution 4, the oxidation-reduction potential of the culture solution 4 may be controlled by the oxidation-reduction substance 3 contained in the soil. However, the oxidation-reduction substance 3 is not limited to the above-described substances.

  Further, by providing the ion exchange membrane 6, the hydrogen-assimilating methane bacteria contained in the culture solution 4 can remain in the activity control tank 7. Therefore, by providing the ion exchange membrane 6, it is possible to make the hydrogen-assimilating methane bacteria stay in the culture solution 4, which is a suitable environment in which the oxidation-reduction potential is controlled, and to easily obtain the effects of the present invention. Can do.

  In some cases, the potential of the culture solution 4 can be controlled without adding the oxidation-reduction substance 3, so that the oxidation-reduction substance 3 does not necessarily have to be added. Further, since the potential of the culture solution 4 may be controlled without providing the ion exchange membrane 6, the ion exchange membrane 6 is not necessarily provided.

(First embodiment B)

  In the activity control apparatus 1 shown in FIG. 6, two tanks opened by partitioning a container 23 whose upper side is opened by an ion exchange membrane 6 are formed, and an upper open part of one tank as the activity control tank 7 Is closed by a gas-impermeable film or a gas-impermeable member 24. That is, the activation control device 1 shown in FIG. 6 has the same configuration as that in FIG. 5 except that the gas generated from the counter electrode vessel 8 is not leaked to the activation control vessel 7. Therefore, the same effect as that obtained when the activation control device shown in FIG. 5 is used can be obtained.

  As the gas impermeable film or the gas impermeable member 24, those generally used in various fields can be appropriately used. For example, examples of the gas impermeable member include, but are not limited to, glass, plastic, an insulating metal, concrete, and the like. Further, as the gas impermeable membrane, for example, an ion exchange membrane 6 can be used, but is not limited thereto.

  The counter electrode tank 8 may be left open, but has a sealed structure like the activation control tank 7 and includes a gas discharge pipe for discharging the gas generated in the counter electrode tank 8 to the outside of the counter electrode tank 8. You may do it. In this case, since the gas generated from the counter electrode tank 8 can be discharged from a desired position, it can be recovered and reused in some cases.

(First embodiment C)
The activation control device 1 shown in FIG. 7 is a U-shaped container in which two containers 25a and 25b each having an opening below the liquid level of the liquid to be accommodated are connected to each other through the ion exchange membrane 6. 25, one container 25a is used as an active control tank 7 with a sealed structure, and the other container 25b is opened as a counter electrode tank 8. In this case, the culture solution 4 and the electrolytic solution 4a are in contact with each other through the ion exchange membrane 6, and the space above the liquid level of the culture solution 4 in the activity control tank 7 and the liquid level of the electrolytic solution 4a in the counter electrode tank 8 are used. The upper space is further separated by the U-shaped structure of the container 25 itself. And since one container 25a is made into the airtight structure, the biogas generated from the activity control tank 7 is the activity control tank while preventing the gas generated from the counter electrode tank 8 from entering the activity control tank 7. 7 can be prevented from leaking. Therefore, the same effect as that obtained when the activation control device shown in FIG. 5 is used can be obtained.

  Note that the opening of the other container 25b in the activation control device 1 shown in FIG. 7 means that, for example, when the end of the other container 25b is completely opened, a sealed structure is formed as in the case of the one container 25a. It also means that a gas discharge pipe for discharging the gas generated in the counter electrode tank 8 to the outside of the counter electrode tank 8 is included. When the gas discharge pipe is provided, the gas generated from the counter electrode tank 8 can be discharged from a desired position, so that it can be easily recovered and reused.

(First embodiment D)
The active control device 1 shown in FIG. 8 has an H-shaped container in which two containers 26a and 26b each having an opening below the liquid level of the liquid to be accommodated are connected to each other through the opening through the ion exchange membrane 6. 26, one container 26a is used as an active control tank 7 with a sealed structure, and the other container 26b is opened as a counter electrode tank 8. Also in this case, the culture solution 4 and the electrolyte solution 4a are in contact with each other through the ion exchange membrane 6, and the space above the liquid level of the culture solution 4 in the activity control tank 7 and the electrolyte solution 4a in the counter electrode tank 8 are also in contact. The space above the liquid level is separated by the H-shaped structure of the container 26 itself. And since one container 26a of the H-shaped container 26 is made into the sealed structure, the activity control tank 7 becomes a sealed structure. Therefore, it is possible to prevent the biogas generated from the activation control tank 7 from leaking from the activation control tank 7 while preventing the gas generated from the counter electrode tank 8 from entering the activation control tank 7. Therefore, the same effect as that obtained when the activation control device shown in FIG. 5 is used can be obtained.

  In the present embodiment, the opening of the other container 26b is not limited to the case where the container 26 is completely opened. This also includes the case where a gas discharge pipe for discharging outside the electrode tank 8 is provided. When the gas discharge pipe is provided, the gas generated from the counter electrode tank 8 can be discharged from a desired position, so that it can be easily recovered and reused.

<Second Embodiment>
In the method of the present invention according to the second embodiment, the working electrode, the counter electrode, and the reference electrode are connected to a constant potential setting device, the culture solution and the counter electrode are brought into contact with each other through an ion exchange membrane, and the culture solution is contacted. The working electrode and the reference electrode are brought into contact with each other, and the potential of the working electrode is controlled by a three-electrode system, whereby the oxidation-reduction potential of the culture solution is controlled to be larger than the reference potential on the negative side. That is, the method according to the first embodiment is different from the method according to the first embodiment only in that the counter electrode is brought into direct contact with the ion exchange membrane without using an electrolytic solution.

  However, an ionic current can flow between the working electrode 9 and the counter electrode 10 via the ion exchange membrane 6 without using the electrolytic solution 4a as in the first embodiment. Further, the effect of retaining the hydrogen-assimilating methane bacteria in the culture solution 4 in the activity control tank 7 without moving (diffusing) to the counter electrode 10 side is also obtained. Furthermore, the effect of not allowing the redox material 3 in the culture solution 4 to permeate the counter electrode 10 side is also obtained. Therefore, according to the method according to the second embodiment, the same effect can be obtained under the same potential control conditions as in the first embodiment.

  The method of the present invention according to the second embodiment is implemented by, for example, an activation control device shown in FIG. In the activity control apparatus 1 shown in FIG. 9, the working electrode 9 and the reference electrode 11 are disposed in a sealed container 5 having at least a part of the ion exchange membrane 6, and the counter electrode 10 is disposed outside the container 5. The culture solution 4 is accommodated in the container 5, the working electrode 9 and the reference electrode 11 are immersed in the culture solution 4, and the ion exchange membrane 6 of the container 4 is at least a part when the culture solution 4 is accommodated in the container 5. Is provided at a position where it can come into contact with the ion exchange membrane 6, and the counter electrode 10 is arranged in contact with at least a part of the surface of the ion exchange membrane 6 opposite to the contact surface of the culture solution 4. . In the activity control apparatus 1 shown in FIG. 9, an opening 5 a is provided below the liquid level of the culture solution 4 in the container 5, the opening 5 a is closed by the ion exchange membrane 6, and ion exchange outside the container 5 is performed. It is assumed that the counter electrode 10 is disposed in contact with at least a part of the surface of the film 6. That is, in the activation control device 1 shown in FIG. 9, the entire container 5 functions as the activation control tank 7.

  Therefore, according to the activity control apparatus 1 shown in FIG. 9, biogas does not leak from the container 5. Further, since the gas generated from the counter electrode 10 does not leak into the container 5, the gas generated from the counter electrode 10 is mixed into the biogas to reduce the methane concentration of the biogas, or from the counter electrode 10. The dissolved gas does not dissolve in the culture solution 4 and does not adversely affect the growth and function of the hydrogen-utilizing methane bacteria. Furthermore, since the container 5 has a sealed structure, there is an advantage that the inside of the container 5 can be easily controlled in an anaerobic environment.

  In addition, although the activity control apparatus 1 shown in FIG. 9 includes the gas recovery means 15 and the culture medium collection means 16 as in the first embodiment, as described above, the gas recovery method and the culture medium collection are performed. The method is not limited to those using these means. Further, as in the first embodiment, a means for adding and supplying a substance to the culture solution 4 may be provided.

  Details of the activation control device 1 shown in FIG. 9 will be described below. However, configurations other than those described below are substantially the same as those in the first embodiment, and a description thereof will be omitted.

  The container 5 has a sealed structure including at least a part of the ion exchange membrane 6. Examples of the material of the container 5 include, but are not limited to, glass, plastic, metal subjected to insulation treatment, concrete, and the like. In FIG. 9, the opening 5a provided below the liquid level of the culture solution 4 in the sealed container 5 is closed by the ion exchange membrane 6. However, the form and structure of the container 5 are particularly limited. Not. For example, the entire container 5 may be a bag-shaped container formed of the ion-exchange membrane 6, or only one surface of the bag-shaped container may be formed of the ion-exchange membrane 6, or a part of one surface may be ion-exchanged. You may make it comprise only with the film | membrane 6. FIG. When the ion exchange membrane 6 is partially used, the other portions may be made of the above-mentioned material such as glass, or other membrane materials other than the ion exchange membrane 6, such as the culture solution 4 and the components in the culture solution 4. You may comprise by the film | membrane material which does not permeate | transmit both (a hydrogen-utilizing methane bacterium is included). In short, the culture solution 4 accommodated in the container 5 may be a container having a structure that can come into contact with the ion exchange membrane 6 constituting at least a part of the container 5.

  The counter electrode 10 is brought into contact with at least a part of the surface opposite to the contact surface of the ion exchange membrane 6 with the culture solution 4. In the present embodiment, the counter electrode 10 is a plate-like carbon electrode, but the shape and material of the counter electrode 10 are not limited to this, and the shape is that the contact with the ion exchange membrane 6 is essential. And a material capable of proceeding with a reaction that complements the exchange of electrons with respect to the redox reaction at the working electrode 9. In the present embodiment, the counter electrode 10 has a larger area than the ion exchange membrane 6 so that the entire ion exchange membrane 6 is completely covered with the counter electrode 10, and the ion exchange membrane 6 and the counter electrode 10 are covered. If the counter electrode 10 is brought into contact with at least a part of the surface opposite to the contact surface of the ion exchange membrane 6 with the culture solution 4, the culture solution 4 is passed through the ion exchange membrane 6. Therefore, it is not always necessary to bring the ion exchange membrane 6 and the counter electrode 10 into contact with each other so that the entire ion exchange membrane 6 is completely covered with the counter electrode 10. However, by completely covering the entire ion exchange membrane 6 with the counter electrode 10, the counter electrode 10 can function as a protective material for the ion exchange membrane 6, and the ion transmission surface from the culture solution 4 increases. As a result, there is an advantage that the potential controllability of the culture solution 4 can be improved, which is preferable. As a method of completely covering the entire ion exchange membrane 6 with the counter electrode 10, for example, ion exchange is performed by closing the opening 5 a by applying an adhesive around the opening 5 a of the container 5 and bonding the counter electrode 10. The entire membrane 6 and the counter electrode 10 may be brought into contact with each other, or an ion exchange membrane formed by applying an adhesive around the opening 5a of the container 5 and applying it to at least a part of the surface of the counter electrode 10 By bonding 6, the counter electrode 10 may be brought into contact with the entire ion exchange membrane 6 that closes the opening 5 a while closing the opening 5 a with the ion exchange membrane 6. Examples of the agent for coating and forming the ion exchange membrane 6 include a Nafion dispersion, but are not limited thereto. Alternatively, a Nafion dispersion may be applied to the surface of the counter electrode 10 and the ion exchange membrane 6 may be attached before the Nafion dispersion is dried. In this case, the adhesion and contact properties of the ion exchange membrane 6 to the surface of the counter electrode 10 can be made sufficient.

  Here, the counter electrode 10 is preferably a porous body. In this case, the gas generated at the contact surface between the ion exchange membrane 6 and the counter electrode 10 can easily pass through the surface opposite to the contact surface. In addition, the counter electrode 10 is made a porous body, and the ion exchange membrane 6 is attached using a Nafion dispersion liquid, whereby the ion exchange membrane 6 and the counter electrode 10 are separated by the penetration of the Nafion dispersion liquid into the pores of the porous body. The contact area can be increased to facilitate the progress of the electrochemical reaction, which is preferable.

  The above-described embodiment is an example of a preferred embodiment of the present invention, but is not limited thereto, and various modifications can be made without departing from the gist of the present invention.

  For example, in the first embodiment and the second embodiment described above, the method of the present invention in the case of increasing the methane generation activity has been described based on FIGS. 5 to 9, but the above embodiment has the methane generation activity. Needless to say, the present invention is applicable to the method of the present invention in the case of lowering.

  In addition, the activity control apparatus for carrying out the present invention, for example, as shown in FIG. 10, allows the culture solution 4 and the electrolyte 4a to permeate ions and hydrogen-utilizing methane bacteria, not the ion exchange membrane 6. The culture solution 4 is separated by a non-impermeable member 40, or the activity control tank 7 and the counter electrode tank 8 are formed in separate containers, and a salt bridge 41 (agar or the like containing a saturated electrolyte solution such as KCl). And electrolyte 4a may be brought into contact (liquid junction). Also in this case, the movement of the hydrogen-assimilating methane bacteria in the culture solution 4 to the counter electrode tank 8 can be prevented, and the salt bridge 41 allows the flow of ion current. Further, since the redox substance 3 contained in the culture solution 4 does not permeate the counter electrode tank 8, the controllability of the potential of the culture solution 4 is also ensured.

Examples of the present invention will be described below, but the present invention is not limited to these examples.
<Experimental apparatus and experimental method>
Experiments were performed using the experimental apparatus shown in FIG. A 250 mL glass vial (manufactured by Duran) was used as the culture container 5, and three openings were provided below the liquid level of the culture solution 4. The reference electrode 11 (manufactured by Toa DDK, HS-205C, silver / silver chloride electrode) was inserted into the first opening 5b, and the reference electrode 11 and the culture solution 4 were brought into contact with each other. The second opening 5c is provided to collect the culture solution 4, and the culture solution 4 can be collected by opening and closing the lid. The third opening 5 a was closed with the counter electrode 10 through the ion exchange membrane 6. The counter electrode 10 was a porous plate-like carbon electrode. As the ion exchange membrane 6, a Nafion membrane N117 (manufactured by DuPont) was used, and a 20% Nafion dispersion DE2021 (manufactured by DuPont) was applied to the lower half of the counter electrode 10, and the ion exchange membrane 6 was attached thereto. A plate-like carbon electrode was immersed in the culture solution 4 as the working electrode 9. Then, the working electrode 9, the counter electrode 10, and the reference electrode 11 are connected to a three-electrode type potential control device (potentiostat, Toho Giken, PS-08) 12, and the potential of the working electrode 9 can be strictly controlled. It was. In addition, the culture solution 4 was put into the culture vessel 5 until about the eighth minute, and a head space was secured above the liquid level. The culture container 5 is provided with a lid 18, and an upper surface 18 a of the lid 18 is provided with a silicone rubber that is an elastic material. The syringe needle of the syringe can be inserted to collect a gaseous substance from the head space in the culture container 5, and The hole produced by inserting the injection needle was closed when the injection needle was removed.

The composition of the culture solution 4 was as follows.
[Composition of culture solution (/ L)]
・ KH ₂ PO ₄ 0.3 g
・ (NH ₄ ) ₂ SO ₄ 1.5 g
・ NaCl 0.6g
· MgSO ₄ · _7H 2 O 0.12g
・ CaCl ₂ · 2H ₂ O 0.08 g
・ FeSO ₄・ 7H ₂ O 4mg
・ K ₂ HPO ₄ 0.15g
・ Na ₂ CO ₃ 4.0 g
・ Trace vitamin 10ml
・ Trace element solution 10ml

The composition of the trace element solution of the culture solution 4 was as follows.
[Composition of trace element solution (/ L)]
・ Na ₂ -EDTA 0.64g
· MgSO ₄ · _7H 2 O 6.2g
・ MnSO ₄ / 4H ₂ O 0.55 g
・ NaCl 1.0g
· FeSO ₄ · _7H 2 O 0.1g
・ CoCl ₂ · 6H ₂ O 0.17 g
・ ZnSO ₄ · 7H ₂ O 0.18 g
・ CuSO ₄ 0.05g
・ KAl (SO ₄ ) ₂ · 12H ₂ O 0.018 g
・ H ₃ BO ₃ 0.01 g
・ Na ₂ MoO ₄ .2H ₂ O 0.011 g
NiCl ₂ · _{6H 2} O 0.025g

The composition of trace vitamins in the culture solution 4 was as follows.
[Minor vitamin composition (/ L)]
・ Biotin 2.0mg
・ Folic acid 2.0mg
・ Pyridoxine-HCl 10.0mg
・ Thiamine-HCl 5.0mg
・ Riboflavin 5.0mg
・ Nicotinic acid 5.0mg
・ Calcium pantothenate 5.0mg
・ Vitamin B ₁₂ 0.1mg
・ P-aminobenzoic acid 5.0mg
・ Lipoic acid 5.0mg

The culture temperature (culture solution temperature) during the experiment was 55 ° C., and the culture solution 4 was continuously stirred by the stirring bar 19. Moreover, AQDS (anthraquinone-2,6-disulfonic acid) was added to the culture solution 4 so that it might become 0.5 mM. In addition, one injection needle is inserted into the upper silicone rubber stopper, and another injection needle is inserted into the rubber stopper portion of the culture medium sampling port 5C, from which H ₂ / CO ₂ = 80/20 (volume ratio). By aeration of the mixed gas, the culture solution and the gas phase above the culture solution were replaced with the mixed gas. The pH of the culture solution 4 was 7.2.

  Experiments were conducted using Methanothermobacter thermautotrophicus DSM1053 as a hydrogen-assimilating methane bacterium.

<Analysis method>
1. Chemical analysis The amount of biogas produced from the culture vessel 5 is measured by the water displacement method, and biogas is produced by gas chromatography equipped with a thermal conductivity detector (GC390B, manufactured by GL Science) and an activated carbon packed column (manufactured by GL Science). The amount of methane gas produced was calculated by analyzing the gas composition.

2. Biological analysis The cell density in the culture solution 4 was measured with a microscope Nikon ECLIPSE E400 (Nikon) and a bacterial counter hemocytometer (Sunlead glass).

Example 1
Experiments were performed by adding hydrogen-assimilating methane bacteria to the culture solution 4 so that the initial cell density was 6.0 × 10 ⁷ cells / mL. In addition, this initial cell density corresponds to the cell density in the stationary phase in a normal culture method, and it has been confirmed that no increase in cell density is observed under any of the following experimental conditions.

  By controlling the set potential of the working electrode 9 to +0.2 V, -0.2 V, -0.5 V, and -0.8 V with respect to the reference electrode 11 (silver / silver chloride electrode), the redox of the culture solution 4 Culture tests were performed under conditions where the potentials were electrically controlled to +0.2 V, -0.2 V, -0.5 V, and -0.8 V, respectively.

As a comparative test, L-cysteine · HCl · H ₂ O and Na ₂ S · 9H ₂ O were added to the culture solution 4 so as to have final concentrations of 0.5 g / L, respectively, without energization. A culture test was conducted under the condition that the oxidation-reduction potential of Solution 4 was chemically adjusted to -0.45V. This condition is a condition corresponding to a redox potential at which a hydrogen-assimilating methane bacterium capable of being formed in the environment by an added chemical component can grow, that is, a reference potential.

  FIG. 1 shows the results of measuring changes over time in the amount of methane produced in culture tests under various conditions. At −0.5V, there was almost no difference from the comparative test, but at −0.8V, there was a tendency for the amount of methane production to increase significantly compared to the comparative test, and methane production calculated from the slope of the approximate line. The speed was found to be 1.7 times better than the comparative test. On the other hand, at +0.2 V and -0.2 V, a tendency that the amount of methane produced was significantly reduced as compared with the comparative test was observed.

  Here, in Example 1, since it was confirmed that no increase in the cell density was observed under any of the experimental conditions, the increase or decrease in the amount of methane produced was due to the methane production of one hydrogen-utilizing methane bacterium. It was presumed to have been caused by the increase or decrease in activity.

  That is, from the results of the culture test in Example 1, it became clear that the methane production activity of the hydrogen-utilizing methane bacterium itself can be controlled by electrical control of the oxidation-reduction potential of the culture solution 4.

  Further, from the results of the culture test in Example 1, by controlling the oxidation-reduction potential of the culture solution 4 to a larger value on the minus side than -0.8V, an effect approximated to the case of controlling to -0.8V is obtained. It was estimated that Furthermore, since there was a tendency for the amount of methane produced to decrease significantly at +0.2 V and −0.2 V compared to the comparative test, the oxidation-reduction potential of the culture solution 4 was set to a larger value on the plus side than the reference potential. If controlled, the methane production activity can be reduced, and if controlled to a larger value on the plus side than -0.2 V, the methane production activity can be reliably reduced, and from -0.2 V to +0.2 V It was estimated that the methanogenic activity could be more reliably reduced by controlling to.

(Example 2)
Experiments were conducted by adding hydrogen-utilizing methane bacteria to the culture solution 4 so that the initial cell density was 1.5 × 10 ⁶ cells / mL, and the electric potential of redox potential for the growth of hydrogen-utilizing methane bacteria. The effects of dynamic control were examined.

  By controlling the set potential of the working electrode 9 to +0.4 V, −0.1 V, −0.4 V, and −0.8 V with respect to the reference electrode 11 (silver / silver chloride electrode), the redox of the culture solution 4 Culture tests were performed under conditions where the potentials were electrically controlled to + 0.4V, -0.1V, -0.4V, and -0.8V, respectively.

  A comparative test was performed under the same conditions as in Example 1.

  The cell density of the culture solution 4 after culturing for one week under various conditions is shown in FIG. In addition, the numerical value of the vertical axis | shaft in FIG. 2 is a relative value when the microbial cell density in a comparative test is set to 1. It was revealed that the cell density significantly increased at −0.8 V as compared with the comparative test.

  Next, FIG. 3 shows the amount of methane produced when culturing for one week under various conditions. In addition, the numerical value of the vertical axis | shaft in FIG. 3 is a relative value when the methane production amount in a comparative test is set to 1. Moreover, the methane production amount in FIG. 3 is the value converted into the methane production amount per microbial cell. It was revealed that the amount of methane produced significantly increased at −0.8 V compared with the comparative test.

  Here, in Example 1, it became clear that the methanogenic activity of the hydrogen-utilizing methane bacterium itself can be improved by setting it to -0.8 V. Therefore, the result obtained in Example 2 is It was estimated that the growth was promoted as a result of improving the methanogenic activity (respiratory activity) of the cells of hydrogen-utilizing methane bacteria per se at −0.8 V.

4 Culture solution (environment)
9 Electrode (working electrode)

Claims (6)

The reference potential is the oxidation-reduction potential at which hydrogen-utilizing methane bacteria that can be formed in the environment by chemical components that are inherent or added can grow.
An electrode is brought into contact with the environment containing the hydrogen-assimilating methane bacterium, a potential is applied to the electrode, and the oxidation-reduction potential of the environment is -0.8 V including -0.8 V based on the silver / silver chloride electrode potential. And controlling the methane production activity per individual hydrogen-utilizing methane bacterium to be higher than the methane production activity per individual hydrogen-utilizing methane bacterium at the reference potential. A method for controlling methane production activity of hydrogen-utilizing methane bacteria. The method for controlling methane production activity of a hydrogen-assimilating methane bacterium according to claim 1, wherein the environment in contact with the electrode has the reference potential. The reference potential is the oxidation-reduction potential at which hydrogen-utilizing methane bacteria that can be formed in the environment by chemical components that are inherent or added can grow.
An electrode is brought into contact with an environment containing the hydrogen-utilizing methane bacterium and having the reference potential, and a potential is applied to the electrode so that the oxidation-reduction potential of the environment is increased to a plus side with respect to the reference potential. and a hydrogen-utilizing methane bacteria, characterized in that to lower than the hydrogen-assimilating methanogens 1 wherein the methanogenic activity per individual in the reference potential hydrogen-utilizing methanogens 1 per individual methanogenic activity Method for controlling methane production activity. The method for controlling methane production activity of hydrogen-utilizing methane bacteria according to claim 1 or 3, wherein the hydrogen-assimilating methane bacteria are contained in the environment at a cell density corresponding to a growth stage in a stationary phase. The method for controlling methane production activity of hydrogen-assimilating methane bacteria according to claim 1, wherein the oxidation-reduction potential of the environment controlled by the electrode is -0.8 V on the basis of the silver / silver chloride electrode potential. 4. The hydrogen assimilation according to claim 3, wherein the oxidation-reduction potential of the environment controlled by the electrode is a potential greater than −0.2 V including −0.2 V on the basis of the silver / silver chloride electrode potential. Method for controlling the methanogenic activity of marine methane bacteria.