Method and system for culturing chemosynthetic autotrophic bacteria

Abstract

This invention provides a method and system for culturing chemosynthetic autotrophic bacteria that enable safe operation while maintaining high culture efficiency. [Solution] A method for culturing chemosynthetic autotrophic bacteria comprises a gas supply step of supplying hydrogen, oxygen, and carbon dioxide to a culture medium containing chemosynthetic autotrophic bacteria; a mixed gas recovery step of recovering gases not dissolved in the culture medium as a mixed gas; and a maintenance step of maintaining the mixed gas so that the oxygen content is less than 5% by volume or the hydrogen content is less than 4% by volume. A culture system for chemosynthetic autotrophic bacteria comprises a gas supply means of supplying hydrogen, oxygen, and carbon dioxide to a culture medium containing chemosynthetic autotrophic bacteria; a mixed gas recovery means of recovering gases not dissolved in the culture medium as a mixed gas; and a maintenance means of maintaining the mixed gas so that the oxygen content is less than 5% by volume or the hydrogen content is less than 4% by volume.

Description

Technical Field

[0001] The present invention relates to a method and a culture system for culturing chemosynthetic autotrophic bacteria. More specifically, the present invention relates to a method and a culture system for culturing chemosynthetic autotrophic bacteria capable of converting carbon dioxide into organic substances using hydrogen and oxygen.

Background Art

[0002] From the viewpoint of CO2 fixation, in the culture of chemosynthetic autotrophic bacteria such as hydrogen-oxidizing bacteria that have attracted attention in recent years, it is necessary to simultaneously supply hydrogen and oxygen to the culture solution. However, since the mixed gas generated in the system has explosiveness, careful handling is required. So far, although the explosiveness has not been a major problem because it has been a laboratory-level culture, the problem becomes apparent in industrial culture. The following patent documents are known regarding this technology.

Prior Art Documents

Patent Documents

[0003]

Patent Document 1

Patent Document 2

Summary of the Invention

Problems to be Solved by the Invention

[0004] Patent Document 1 discloses a method of maintaining the mixed gas outside the explosion range by reducing the supply amount of oxygen to the culture solution. In this regard, chemosynthetic autotrophic bacteria are inherently bacteria capable of extremely efficient growth and are thus attracting attention as a means for CO2 fixation. However, when attempting to achieve highly efficient growth, it is predicted that oxygen will become insufficient as growth progresses. In the method of Patent Document 1, it is necessary to reduce the amount of oxygen supplied to the culture solution. As a result, it directly leads to a reduction in the culture rate and a decrease in the CO2 fixation rate. Patent Document 2 discloses a method of using fluorocarbons with excellent oxygen solubility as a culture medium. However, there are concerns that fluorocarbons may inhibit or reduce the growth of chemosynthetic autotrophic bacteria, leading to a decrease in the CO2 fixation rate.

[0005] This invention has been made in view of the above circumstances, and aims to provide a method and system for culturing chemosynthetic autotrophic bacteria that enables safe operation while maintaining high culture efficiency of chemosynthetic autotrophic bacteria. [Means for solving the problem]

[0006] This invention includes the following inventions. [1] A method for culturing chemosynthetic autotrophic bacteria using hydrogen and oxygen, A gas supply process that supplies hydrogen, oxygen, and carbon dioxide to a culture medium containing chemosynthetic autotrophic bacteria, A mixed gas recovery step for recovering gases not dissolved in the culture medium as a mixed gas, A method for culturing chemosynthetic autotrophic bacteria, comprising a maintenance step of maintaining the mixed gas so that it is as described in A or B below. A: The proportion of oxygen in the mixed gas is less than 5% by volume. B: The proportion of hydrogen in the mixed gas is less than 4% by volume. [2] The method for culturing chemosynthetic autotrophic bacteria according to [1] above, wherein the maintenance step is a step of removing some or all of the oxygen contained in the mixed gas to reach A. [3] The method for culturing chemosynthetic autotrophic bacteria according to [1] or [2] above, wherein the maintenance step is a step of mixing air with the mixed gas to reach B. [4] The method for culturing chemosynthetic autotrophic bacteria according to any one of [1] to [3] above, wherein the maintenance step is a step of adding a gas other than hydrogen and oxygen to the mixed gas to reach A and / or B. [5] The method for culturing chemosynthetic autotrophic bacteria according to any one of [1] to [4] above, wherein the oxygen to be removed is oxygen separated from the mixed gas via a gas separation membrane. [6] A method for culturing chemosynthetic autotrophic bacteria according to any one of [1] to [5] above, wherein hydrogen and / or carbon dioxide separated from the mixed gas via a gas separation membrane is used in the gas supply step. [7] A method for culturing chemosynthetic autotrophic bacteria according to any one of [1] to [6] above, wherein the gas supply step is a step of supplying each gas in a ratio of 50 to 90% by volume for hydrogen, 5 to 25% by volume for oxygen, and 5 to 25% by volume for carbon dioxide relative to the total of hydrogen, oxygen, and carbon dioxide. [8] A culture system for chemosynthetic autotrophic bacteria using hydrogen and oxygen, A gas supply means for supplying hydrogen, oxygen, and carbon dioxide to a culture medium containing chemosynthetic autotrophic bacteria, A mixed gas recovery means for recovering gases not dissolved in the culture medium as a mixed gas, A culture system for chemosynthetic autotrophic bacteria, comprising a maintenance means for maintaining the mixed gas such that it is as described in A or B below. A: The proportion of oxygen in the mixed gas is less than 5% by volume. B: The proportion of hydrogen in the mixed gas is less than 4% by volume. [9] The culture system for chemosynthetic autotrophic bacteria described in [8] above, wherein the maintenance means is a means of removing some or all of the oxygen contained in the mixed gas to reach A.

[10] The culture system for chemosynthetic autotrophic bacteria according to [8] or [9] above, wherein the maintenance means is a means of mixing air with the gas mixture to reach B.

[11] A culture system for chemosynthetic autotrophic bacteria according to any one of [8] to

[10] above, wherein the maintenance means is a means of adding a gas other than hydrogen and oxygen to the mixed gas to achieve A and / or B.

[12] A culture system for chemosynthetic autotrophic bacteria according to any one of [8] to

[11] above, wherein the oxygen to be removed is oxygen separated from the mixed gas via a gas separation membrane.

[13] A culture system for chemosynthetic autotrophic bacteria according to any one of [8] to

[12] above, wherein hydrogen and / or carbon dioxide separated from the mixed gas via a gas separation membrane is used in the gas supply means.

[14] A culture system for chemosynthetic autotrophic bacteria according to any one of [8] to

[13] above, wherein the gas supply means is a means for supplying each gas in a ratio of 50-90% by volume for hydrogen, 5-25% by volume for oxygen, and 5-25% by volume for carbon dioxide relative to the total of hydrogen, oxygen, and carbon dioxide. [Effects of the Invention]

[0007] The present invention provides a method for culturing chemosynthetic autotrophic bacteria that enables safe operation while maintaining high cultivation efficiency of chemosynthetic autotrophic bacteria. The culture system for chemosynthetic autotrophic bacteria of the present invention enables safe operation while maintaining high culture efficiency for chemosynthetic autotrophic bacteria. [Brief explanation of the drawing]

[0008] [Figure 1] This is an explanatory diagram illustrating an example of a culture system (1A). [Figure 2] This is an explanatory diagram illustrating another example of a culture system (1B). [Figure 3] This is an explanatory diagram illustrating another example of a culture system (1C). [Modes for carrying out the invention]

[0009] The present invention will be described below with reference to the drawings. The matters described herein are illustrative and illustrative to illustrate embodiments of the present invention, and are intended to provide what is considered to be the most effective and straightforward explanation of the principles and conceptual features of the present invention. In this regard, it is not intended to describe structural details of the present invention beyond what is necessary for a fundamental understanding of the invention, but rather to make it clear to those skilled in the art, through the description and drawings, how some forms of the present invention are actually realized.

[0010] [1] Method for culturing chemoautotrophic bacteria The present invention relates to a method for culturing chemoautotrophic bacteria using hydrogen and oxygen, and is characterized by comprising a gas supply step, a mixed gas recovery step, and a maintenance step.

[0011] "Chemoautotrophic bacteria" are bacteria that grow (proliferate) by chemoautotrophic metabolism, and are bacteria that utilize energy from the oxidation reaction of inorganic substances to reduce carbon dioxide (CO2) and synthesize organic substances. Examples of such bacteria include hydrogen-oxidizing bacteria (hydrogen bacteria), methanogens, methane-oxidizing bacteria, nitrifying bacteria, nitrite bacteria, acetic acid bacteria, sulfur-oxidizing bacteria, iron-oxidizing bacteria, CO-assimilating bacteria, anammox bacteria, and the like. Only one type of these may be used, or two or more types may be used in combination.

[0012] Among these, as the chemoautotrophic bacteria used in this method, it is preferable that the bacteria can utilize at least the oxidation reaction of hydrogen (free hydrogen, an inorganic substance) as an energy source and can utilize carbon dioxide as a carbon source. For this reason, hydrogen-oxidizing bacteria are preferable as the chemoautotrophic bacteria used in this method. That is, hydrogen-oxidizing bacteria are bacteria that utilize energy from the oxidation reaction of free hydrogen to reduce carbon dioxide and synthesize organic substances. Since hydrogen-oxidizing bacteria have a high growth rate among chemoautotrophic bacteria, the effect of increasing the bacterial concentration in a short time can be obtained by selecting hydrogen-oxidizing bacteria. In addition, since hydrogen-oxidizing bacteria do not require light supply unlike photosynthetic bacteria, the effect of being able to perform culturing in a more space-saving manner can be obtained by selecting hydrogen-oxidizing bacteria. In addition, the organic substances synthesized by chemoautotrophic bacteria are not limited and include carbohydrates, amino acids, and proteins. Among these, examples of carbohydrates include various organic acids such as hydroxybutyric acid (hydroxybutanoic acid) and hydroxypentanoic acid (hydroxypentanoic acid), and polymers thereof (such as polyhydroxyalkanoic acid). Only one type of these may be used, or two or more types may be used in combination.

[0013] Furthermore, from the viewpoint of CO2 fixation efficiency, bacteria with high efficiency in converting carbon dioxide into organic matter are preferable. For example, bacteria that use carbon dioxide as their sole carbon source (i.e., bacteria that synthesize all organic matter in their bodies from carbon dioxide) can be used. Furthermore, the chemosynthetic autotrophic bacteria used in this method can be any bacteria capable of using carbon dioxide as a carbon source, as described above. For example, it is not limited to bacteria capable of using sugars and / or lipids as carbon sources in addition to carbon dioxide.

[0014] The species of このようなchemically synthesized independent phytotrophic bacteria (special に, hydroacidifying bacteria) are limited to されるものではないが, example えば, Alcaligenes genus (A. hydrogenophilus, A.ruhlandii, A.latus, A.paradoxus, etc.), Aquaspirillum Genus (A.autotrophicum, etc.), Arthrobacter (A.globiformis, etc.), Azospirillum (A.lipoferum, etc.), Calderobacterium (C.hydrogenophilum, etc.), Cupriavid *C. us* (C. necator, C. oxalaticus, C. pauculus, C. respiraculi, C. taiwanensis, C. metallidurans, etc.), *Derxia* (D. gummosa, etc.), *Flavobacterium* (F. autothermophilum, etc.), *Hydrogenibacillus* (H. schlegelii, etc.), *Hydrogenobacter* (H. thermophilus, etc.), *Hydrogenophaga* (H. aquatica, H. atypica, H. bi... sanensis,H.caeni,H.crassostreae,H.defluvii,H.flava,H.laconesensis,H.luteola,H.intermedia,H.palleronii,H.pseudoflava,H.soli,H. taeniospiralis, etc.), Hydrogenophilus (H.thermoluteolus), Hydrogenovibrio (H.marinus, etc.), Kyrpidia (K.tusciae, etc.), Microcyclus (M.aquatic us, M.ebruneus, etc.), Mycobacterium (M.gordonae, M.llatzerense, etc.), Nocardia (N.autotrophica, N.opaca, etc.), Paracoccus (P.denitrificans, P.pa ntotrophus, etc.), Pseudomonas (P.facilis, P.flava, P.pseudoflava, P.hydrogenovora, P.hydrogenothermophila, P.palleronii, P.thermophila, P.Examples include the genera *Saccharophila*, *Renobacter* (e.g., *R. vacuolatum*), *Rhizobium* (e.g., *R. japonicuni*), *Rhodobacter* (e.g., *R. sphaeroides*, *R. capsulatus*), *Rhodococcus* (e.g., *R. opacus*, *R. erythropolis*), *Rhodopseudomonas* (e.g., *R. sphaeroides*, *R. palustris*), *Rhodospirillum* (e.g., *R. rubrum*), *Ralstonia* (e.g., *R. eutropha*), *Wautersia* (e.g., *W. eutropha*), and *Xanthobacter* (e.g., *X. autotrophicus*, *X. flavus*). These can be used individually or in combination of two or more species.

[0015] The origin of these bacteria is not limited, and those isolated from various environments (aquatic environments such as soil, ponds, rivers, lakes, swamps, and the sea) can be used. Furthermore, the chemosynthetic autotrophic bacteria (especially hydrogen-oxidizing bacteria) mentioned above may not only be strains that inherently possess organic matter synthesis genes (organic matter synthesis enzyme genes), but also recombinant strains (transformed organisms) into which acquired genes encoding organic matter (organic matter synthesis enzymes derived from other bacteria, etc.) have been introduced. When using recombinant strains, only one type may be used, or two or more types may be used in combination. In addition, either non-recombinant strains or recombinant strains may be used alone, or in combination. Among the chemosynthetic autotrophic bacteria (especially hydrogen-oxidizing bacteria) used in this method, the following species can be suitably used: Cupriavidus necator, Cupriavidus oxalaticus, Cupriavidus pauculus, Cupriavidus respiraculi, Cupriavidus taiwanensis, and Cupriavidus metallidurans. Of these, Cupriavidus necator (e.g., strain H16) is particularly suitable.

[0016] The "gas supply process" is the process of supplying hydrogen, oxygen, and carbon dioxide to a culture medium containing chemosynthetic autotrophic bacteria. The supplied hydrogen, oxygen, and carbon dioxide are all gases. Furthermore, these hydrogen, oxygen, and carbon dioxide are all substrates for the chemosynthetic autotrophic bacteria being cultured. The supply of these gases may be discontinuous (i.e., in batches as needed) or continuous. If the gas is supplied continuously, it becomes continuous aeration culture.

[0017] Culture medium is a culture medium used for culturing chemosynthetic autotrophic bacteria, and typically contains water as its main liquid component. By using a liquid medium (i.e., culture medium) instead of a solid medium, it is possible to simplify the control of culture conditions (controlling the amount of substrate supplied, measuring substrate mass, controlling pH, etc.) and improve culture efficiency. Furthermore, it is possible to simplify the separation of the culture (chemosynthetic autotrophic bacteria) from the medium after cultivation.

[0018] As described above, the culture medium only needs to be able to supply hydrogen, oxygen, and carbon dioxide, and is not limited to other matters, but may contain other substrates (inorganic nutrients) besides hydrogen, oxygen, and carbon dioxide. Examples of other substrates include inorganic salts. The types of inorganic salts are not limited, but examples include nitrogen sources (ammonia, ammonium salts, nitrates, etc.), phosphorus sources (phosphates, etc.), and other necessary bioelements (alkaline earth metal salts such as MgSO4 and CaCl2, transition metal salts such as FeCl3 and NiCl2, etc.). These may be used individually or in combination of two or more. Furthermore, the culture medium may contain buffering components (Na2HPO4, KH2PO4, NaHCO3, etc.) to buffer the pH fluctuation (pH decrease) associated with the dissolved carbon dioxide being supplied. These components other than hydrogen, oxygen, and carbon dioxide may be supplied to the culture medium as needed, may be contained in the culture medium from the beginning, or may be used in combination.

[0019] Hydrogen (free hydrogen) is a component that is oxidized, and it is the component that provides the energy necessary for carbon dioxide reduction during organic synthesis through oxidation. The source of hydrogen is not limited; manufactured hydrogen may be used, or hydrogen produced as a by-product in various facilities and equipment may be used. Among these, examples of hydrogen produced include hydrogen produced by steam reforming of fossil fuels (natural gas, petroleum, etc.), hydrogen produced by partial oxidation reforming of fossil fuels (natural gas, petroleum, etc.), hydrogen produced by reforming alcohols (methanol, ethanol, etc.), and hydrogen produced by electrolysis of water. These may be used individually or in combination of two or more. Furthermore, examples of by-product hydrogen include hydrogen produced during the manufacturing process of caustic soda and hydrogen produced during the manufacturing process of chlorine. These may be used individually or in combination of two or more. In addition, if by-product hydrogen is contained in the mixed gas, the mixed gas may be used as is, or it may be used as a mixed gas with a higher hydrogen concentration after passing through a filter, concentrator, separator, etc., or it may be used as purified hydrogen after passing through a filter, concentrator, separator, etc.

[0020] Means and methods for separating or concentrating hydrogen include gas separation membranes, cryogenic separation, and pressure swing adsorption (PSA). These may be used individually or in combination of two or more. Among these, gas separation membranes include polyimide membranes, polyamide membranes, and palladium alloy membranes. These may be used individually or in combination of two or more.

[0021] Furthermore, while hydrogen may be supplied together with oxygen and / or carbon dioxide, it is preferable to supply hydrogen separately from oxygen. Supplying hydrogen and oxygen together requires controlling the concentrations of hydrogen and oxygen in the mixture to stay outside the explosive range, necessitating a concentration adjustment mechanism. In contrast, separate supply eliminates the need for this control and reliably prevents the reaction between hydrogen and oxygen.

[0022] Oxygen (free oxygen) is an electron acceptor and typically constitutes the reaction [H2 + 1 / 2O2 → H2O]. While this method supplies at least oxygen as an electron acceptor, it does not prevent the supply of other electron acceptors besides oxygen. Other components that can function as electron acceptors include carbon dioxide and nitrate ions (NO). 3- Examples include the following. These may be used individually or in combination of two or more.

[0023] The source of oxygen is not limited; manufactured oxygen may be used, oxygen produced as a by-product in various facilities and equipment may be used, or air (atmosphere) may be used. Among these, examples of manufactured oxygen include oxygen produced by the electrolysis of water and oxygen separated from air. These may be used individually or in combination of two or more. Furthermore, examples of by-product oxygen include combustion exhaust gas (combustion exhaust gas containing excess oxygen). These may be used individually or in combination of two or more. Since combustion exhaust gas contains carbon dioxide along with oxygen, it can be suitably used in the gas supply process. In addition, combustion exhaust gas containing excess oxygen may be used as is, or it may be used as a mixed gas with a higher oxygen concentration after being filtered, concentrated, or separated.

[0024] Means and methods for separating or concentrating oxygen include gas separation membranes, zeolite adsorption methods, and cryogenic separation methods. These may be used individually or in combination of two or more. Among these, gas separation membranes include polyimide membranes, polyamide membranes, and ceramic membranes. These may be used individually or in combination of two or more. Furthermore, similar to the supply of hydrogen, oxygen may be supplied together with oxygen and / or carbon dioxide, but it is preferable to supply hydrogen separately from oxygen. Furthermore, since carbon monoxide (CO) can inhibit the growth of hydrogen bacteria, if CO is present in the combustion exhaust gas, it is preferable to remove the carbon monoxide or convert it to carbon dioxide by oxidation.

[0025] Carbon dioxide is a carbon source for chemosynthetic autotrophic bacteria. The source of carbon dioxide is not limited; it can be manufactured carbon dioxide, carbon dioxide produced as a by-product in various facilities and equipment, or air (atmosphere). Typically, carbon dioxide can be supplied from either by-product carbon dioxide or the atmosphere. Examples of by-products of carbon dioxide include combustion exhaust gas, exhaust gas from steelmaking, and exhaust gas from petrochemical plants. These may be used individually or in combination of two or more. Since these exhaust gases contain oxygen along with carbon dioxide, they can be suitably used in gas supply processes. Furthermore, the exhaust gas containing carbon dioxide may be used as is, or it may be used as a mixed gas with a higher carbon dioxide concentration after being filtered, concentrated, separated, etc.

[0026] Methods and techniques for separating or concentrating carbon dioxide include gas separation membranes, cryogenic separation, pressure swing adsorption (PSA), and chemical absorption using alkaline solutions. These may be used individually or in combination of two or more. Examples of gas separation membranes include polyimide membranes, polyamide membranes, and ceramic membranes. These may be used individually or in combination of two or more.

[0027] Furthermore, carbon dioxide can be supplied together with hydrogen and / or oxygen, but when using a mixed gas, it is preferable to use (1) a mixed gas containing carbon dioxide and hydrogen but not oxygen, or (2) a mixed gas containing carbon dioxide and oxygen but not hydrogen, rather than using a mixed gas containing all three components of carbon dioxide, hydrogen, and oxygen simultaneously.

[0028] As described above, gases with increased concentrations of the three substrate gases (individual gases, mixed gases) can be produced, for example, using a separator that utilizes a gas separation membrane. That is, each of the above gases permeates the gas separation membrane at a different rate. For example, when a polyimide membrane is used as a gas separation membrane, hydrogen has the highest permeation rate, followed by carbon dioxide, and oxygen has the lowest permeation rate among these three gases. In this way, by utilizing the differences in permeation rates through the gas separation membrane, it is possible to easily prepare gases that are substantially hydrogen only, mixed gases with a high concentration of hydrogen, gases that are substantially carbon dioxide only, mixed gases with high concentrations of hydrogen and carbon dioxide, gases that are substantially oxygen only, mixed gases with a high concentration of oxygen, and so on.

[0029] Furthermore, the mixed gas recovered in the mixed gas recovery process described later is typically a gas containing high concentrations of hydrogen, oxygen, and carbon dioxide. Therefore, the recovered mixed gas can be reused in the gas supply process. In particular, the hydrogen and / or carbon dioxide separated from the mixed gas (recovered mixed gas) via the gas separation membrane can be suitably used as the supply gas in the gas supply process. More specifically, for example, by using a polyimide membrane as a gas separation membrane, the permeation rates of hydrogen and carbon dioxide can be made higher than those of oxygen. In other words, the gas separation membrane can create differences in permeation rates for each gas. Consequently, the gas that permeates through the polyimide membrane (the gas on the permeation side of the polyimide membrane) can be a mixture of gases with high concentrations of hydrogen and carbon dioxide. On the other hand, the oxygen concentration in this mixture can be low (especially the oxygen content in the mixture can be less than 5% by volume).

[0030] When the gas mixture recovered in the gas mixture recovery process is reused, the gas mixture can be modified through a maintenance process described later to (A) reduce the oxygen content to less than 5% by volume and / or (B) reduce the hydrogen content to less than 4% by volume. This will be explained later. Furthermore, in polyimide membranes, the permeability rate of hydrogen is higher than that of carbon dioxide. Therefore, this difference in permeability can be used to separate hydrogen and carbon dioxide. However, since the membrane can be safely used as long as hydrogen and oxygen are not present together, this separation is not always necessary.

[0031] Furthermore, in the gas supply process, hydrogen, oxygen, and carbon dioxide are supplied to a single tank containing the culture medium. These gases may be supplied simultaneously, or only specific gases may be supplied. That is, for example, all three gases—hydrogen, oxygen, and carbon dioxide—may be supplied simultaneously, or hydrogen and oxygen may be supplied, but carbon dioxide may not be supplied initially, and carbon dioxide may be supplied after the hydrogen and oxygen have been supplied. Thus, the three gases do not necessarily have to be supplied simultaneously, and may be supplied according to their dissolved amounts in the culture medium as needed. Furthermore, naturally, in the gas supply process, only one tank containing the culture medium may be used, but multiple tanks can also be used simultaneously. When multiple tanks are used, the recovery of the mixed gas (mixed gas recovery process), which will be described later, can be performed individually in each tank, or the non-dissolved gases generated in multiple tanks can be recovered collectively.

[0032] Furthermore, while the gas can be supplied in any way, from the viewpoint of efficient dissolution of the gas into the culture medium, it is preferable to supply the gas into the culture medium. That is, the gas can be supplied into the culture medium in the tank (culture tank) containing the culture medium. Moreover, from the viewpoint of more efficient dissolution of the gas into the culture medium, and from the viewpoint of more efficiently obtaining the stirring effect brought about by excess gas (gas that was supplied to the culture medium but not dissolved) in the culture medium, it is preferable to supply the gas from a deeper position in the culture tank. That is, for example, as shown in Figure 1, the gas can be discharged from the opening of a gas supply pipe inserted to near the bottom of the culture tank.

[0033] In the gas supply process, the supply concentrations of hydrogen, oxygen, and carbon dioxide are not limited and can be supplied individually or in combination at appropriate concentrations. However, from the viewpoint of the growth efficiency of chemosynthetic autotrophic bacteria, it is preferable to supply them in a range where, for example, the proportion of each component relative to the total of these three gases is 50-90% by volume for hydrogen, 5-25% by volume for oxygen, and 5-25% by volume for carbon dioxide. This ratio can be further adjusted to 60-88% by volume for hydrogen, 6-20% by volume for oxygen, and 6-20% by volume for carbon dioxide; 70-86% by volume for hydrogen, 7-18% by volume for oxygen, and 7-18% by volume for carbon dioxide; or 75-85% by volume for hydrogen, 8-15% by volume for oxygen, and 8-15% by volume for carbon dioxide. Within these preferred ranges, chemosynthetic autotrophic bacteria can be produced more efficiently.

[0034] The "mixed gas recovery process" is a process for recovering gases that are not dissolved in the culture medium as a mixed gas. Normally, it is difficult to supply only the amount of each gas corresponding to its dissolved amount to the culture medium, so an amount exceeding the dissolved amount of each gas is supplied to the culture medium. This replenishes the substrate consumed in the culture medium and effectively keeps the dissolved amount of each gas saturated at all times. Therefore, when gas is supplied in this way, gases that are not absorbed by the culture medium are released from the surface of the culture medium. At this time, it is difficult to recover the gases that have passed through the culture medium individually, so they are recovered as a mixed gas of hydrogen, oxygen, and carbon dioxide.

[0035] The mixed gas can be recovered in any way. For example, it can be recovered by providing a recovery space above the container holding the culture medium and recovering the gas present in that space. Alternatively, it can be recovered by providing a duct above the container holding the culture medium and recovering the gas released from the surface of the culture medium through the duct. In this case, the mixed gas can be recovered without substantially providing a recovery space, or by reducing the volume of the recovery space if one is provided. That is, substrate (gas) that was not dissolved in the culture medium can be recovered without being released outside the system and without waste.

[0036] The "maintenance process" is a process of maintaining the gas mixture so that either "(A) the proportion of oxygen in the gas mixture is less than 5 volume%" or "(B) the proportion of hydrogen in the gas mixture is less than 4 volume%". The gas mixture recovered in the gas mixture recovery process contains hydrogen, oxygen, and carbon dioxide (and other gases if other gases are supplied). When hydrogen and oxygen coexist, the gas mixture may become explosive. Specifically, the gas mixture can become explosive in a wide concentration range, where hydrogen accounts for 4 to 95 volume percent and oxygen accounts for 5 volume percent or more, relative to a total of 100 volume percent of hydrogen and oxygen. Therefore, the explosive concentration range can be avoided by (A) controlling the concentration of each gas in the gas mixture so that the proportion of oxygen in the gas mixture is less than 5 volume percent, and / or (B) controlling the concentration of each gas in the gas mixture so that the proportion of hydrogen in the gas mixture is less than 4 volume percent.

[0037] The above (A) and (B) can be achieved in any way, but for example, they can be achieved by any of the following operations (M1) to (M3), or a combination of these operations. (M1): Remove some or all of the oxygen contained in the recovered gas mixture to achieve (A) above. (M2): The recovered gas mixture is mixed with air to achieve (B) above. (M3): Add other gases (gases other than hydrogen and oxygen) to the recovered gas mixture to achieve (A) and / or (B) above.

[0038] When performing the operation described in (M1) above, the maintenance step can be a step to achieve (A) above by removing some or all of the oxygen contained in the recovered gas mixture. This oxygen removal operation can also be described as an operation to reduce the oxygen concentration. That is, the maintenance step can be a step to achieve (A) above by reducing the oxygen concentration contained in the recovered gas mixture. Oxygen can be removed by any method, such as using a gas separation membrane, chemical adsorption (amine-based, etc.), or physical adsorption (activated carbon, zeolite, etc.). These methods may be used individually or in combination of two or more. Of these, gas separation membrane removal is preferable from the viewpoint of energy efficiency and equipment simplification.

[0039] When using a gas separation membrane, oxygen can be removed (the oxygen concentration reduced) by passing the recovered gas mixture through the membrane. For example, when a polyimide membrane is used as a gas separation membrane, the permeation rate of oxygen through the polyimide membrane is lower than that of hydrogen and carbon dioxide. On the other hand, as mentioned above, the recovered gas mixture is a gas with high concentrations of hydrogen, oxygen, and carbon dioxide. Therefore, the gas that permeates through the polyimide membrane (the gas on the permeation side of the polyimide membrane) can be obtained as a gas mixture with a low oxygen concentration (in particular, the proportion of oxygen in the gas mixture is less than 5% by volume). On the other hand, gases that permeate slowly through the polyimide membrane can be recovered as gases that have not permeated the polyimide membrane (gases on the non-permeable side of the polyimide membrane), and thus recovered as a mixed gas with a high oxygen concentration. Therefore, this gas can be reused in the gas supply process for oxygen supply.

[0040] When performing the operation described in (M2) above, the maintenance step can be a step in which air (atmosphere) is mixed with the recovered gas mixture to achieve (B) above. That is, air contains no hydrogen at all, or if it does, the amount of hydrogen is very small (for example, less than 1 ppm by volume). Therefore, when air is mixed with the recovered gas mixture, the hydrogen concentration in the entire gas mixture will decrease. In other words, by monitoring the hydrogen concentration in the recovered gas mixture, the amount of air required to reduce the proportion of hydrogen in the gas mixture to less than 4% by volume can be calculated, and by mixing in that amount of air, (B) can be achieved through the operation described in (M2).

[0041] When performing the operation described in (M3) above, the maintenance step can be a step in which other gases (gases other than hydrogen and oxygen) are added to the recovered gas mixture to achieve (A) and / or (B) above. As described above, when performing the operation described in (M2), i.e., mixing air with the recovered gas mixture, (B) is easily achieved, but (A) tends to be difficult to achieve. This is because air contains oxygen. In this respect, when gases other than hydrogen and oxygen are mixed, it is easier to achieve (A) and (B) simultaneously.

[0042] While the type of gas other than hydrogen and oxygen is not limited, from the viewpoint of preferring less reactive gases, examples include noble gases (helium, neon, argon, krypton, xenon), nitrogen (free nitrogen), and carbon dioxide. These may be used individually or in combination of two or more. Among these, carbon dioxide is preferred from the viewpoint of maximizing the dissolved amount of hydrogen and oxygen. That is, among the above gases, nitrogen and carbon dioxide can be used inexpensively, but of these, nitrogen, when dissolved in the culture medium, reduces the dissolved amount of hydrogen and oxygen, which are the substrates. Therefore, from the viewpoint of maximizing the dissolved amount of substrates, carbon dioxide is preferred. Although carbon dioxide is more soluble in water than other gases and tends to lower the pH of the aqueous solution when dissolved, the pH of the culture medium generally changes to the acidic side as the culture progresses. Therefore, it is preferable to adjust the pH of the culture medium constantly or periodically, and the acidification by carbon dioxide can be corrected by such pH adjustment.

[0043] This method allows for safe operation while maintaining high culture efficiency of chemosynthetic autotrophic bacteria. Specifically, increasing the culture volume for efficient bacterial cultivation increases the amount of hydrogen and oxygen used. Consequently, the amount of gas mixture recovered also increases, raising the risk of explosion. However, this method allows for continuous control of the recovered gas mixture to stay outside the explosive range during the maintenance process. Furthermore, this control can be achieved through low-cost and simple methods such as oxygen removal, air introduction, and nitrogen filling.

[0044] In addition to the gas supply step, mixed gas recovery step, and maintenance step described above, this method may include other steps in addition to the above steps, to the extent that the effects of the present invention are achieved. Only one of these other steps may be used, or two or more may be used in combination. Furthermore, these other steps may be added before the gas supply step, between the above-described steps, or after the maintenance step. Other processes include pH adjustment, temperature adjustment, culture medium recovery, and culture medium replenishment. These processes may be performed individually or in combination of two or more. Of the above steps, the pH adjustment step is the process of adjusting the pH of the culture medium to a pH suitable for culturing chemosynthetic autotrophic bacteria. Generally, the pH of the culture medium tends to become acidic during cultivation, so neutralization can be performed using ammonia (such as aqueous ammonia). Of the above, the temperature adjustment step is the process of adjusting the temperature of the culture medium to a temperature suitable for culturing chemosynthetic autotrophic bacteria. Of the above steps, the culture medium recovery step is a step in which chemosynthetic autotrophic bacteria that have grown through cultivation are removed from the culture medium. That is, since the culture efficiency tends to decrease as bacteria grow in the culture medium, it is preferable to perform an operation to reduce the bacterial density continuously or periodically from the viewpoint of maintaining high culture efficiency. Of the above, the culture medium replenishment step is the step of replenishing the culture medium. In addition to the substrate supplied as a gas, the culture medium may contain various substrates as described above. These components are consumed during cultivation and can be supplied as needed, but it is also possible to replenish with a new culture medium that already contains these components.

[0045] This method allows for the safe and efficient cultivation of chemosynthetic autotrophic bacteria while maintaining high cultivation efficiency. Specifically, organic substances (carbohydrates, amino acids, proteins, etc.) produced by these bacteria can be obtained safely and efficiently through the cultivation of chemosynthetic autotrophic bacteria. This method can be widely applied to various fields such as resin (plastic) manufacturing, fiber manufacturing, fuel manufacturing, reagent manufacturing, industrial raw material manufacturing, food ingredient manufacturing, and feed manufacturing (fish and shellfish feed, poultry feed, pig feed, dairy cow feed, beef cattle feed).

[0046] Examples of organic matter produced by bacteria include polyhydroxyalkanoates (PHA), polyhydroxybutyric acid (PHB), 4-hydroxybenzoic acid (PHBA), hydroxyisobutyric acid, hydroxybutyric acid, methylcitric acid, lactic acid, carboxylic acids, acetic acid, etc.), polysaccharides, amino acids, peptides, proteins (e.g., lysine, arginine, tryptophan, methionine, cyanophycin, organophosphorus hydrolases, etc.), fatty acids (e.g., linear saturated fatty acids, etc.), amines, esters (e.g., lactones, phosphate esters, etc.), nucleic acids, enzymes, vitamins, physiologically active substances, and bioplastics (polyesters, etc.). These can be used individually or in combination of two or more.

[0047] [2] Culture systems for chemosynthetic autotrophic bacteria This culture system 1 is a culture system for chemosynthetic autotrophic bacteria using hydrogen and oxygen, and is characterized by comprising a gas supply means 3, a mixed gas recovery means 5, and a maintenance means 7. In other words, System 1 can be described as a specific system for performing the aforementioned method of culturing chemosynthetic autotrophic bacteria. The terminology used in System 1 and the terminology used in the aforementioned culture method are basically the same, and the explanations in the aforementioned culture method can be similarly applied to the explanation of this System 1.

[0048] The culture medium 11 may contain chemosynthetic autotrophic bacteria and, as mentioned above, inorganic salts, etc. The culture medium 11 is usually housed in a culture tank 10. The culture tank 10 may be a batch type or a continuous type. It may also be a single-tank type or a multi-tank type. In the case of a multi-tank type, the processes may be arranged to proceed in parallel or in series. Furthermore, in this culture system and the culture method described above, the bacteria in the culture medium may be cultured by being directly contained in the culture medium (e.g., the bacteria are suspended in the medium), or they may be cultured by being supported on a carrier that is in contact with the culture medium.

[0049] The culture tank 10 may be an open-type tank that is open to the external environment (atmosphere, etc.), or a closed-type tank that is not open to the external environment (atmosphere, etc.). Of these, a closed-type culture tank is preferred. By using a closed-type culture tank, it is possible to easily control the internal environment of the culture tank. That is, it is possible to reduce the influence of the external environment outside the culture tank. In addition, it is possible to easily recover the substrate (hydrogen, oxygen, carbon dioxide, etc.), and by circulating it as needed, the substrate can be utilized efficiently.

[0050] The culture vessel 10 may be made of any material, but for example, metal (stainless steel, etc.) or glass can be used. These materials may be used individually or in combination of two or more. Using these materials makes the culture vessel less susceptible to external environmental influences (heat, pressure, humidity, etc.) compared to using other materials. Furthermore, excellent corrosion resistance can be obtained.

[0051] Furthermore, the culture tank 10 does not necessarily have an insulating structure, but it may have one. By having an insulating structure, heat exchange between the inside and outside of the culture tank can be suppressed compared to the case without an insulating structure. As the insulating structure, vacuum insulation, insulating materials, etc., can be used. These may be used alone or in combination of two or more types. Vacuum insulation can be achieved, for example, by making the culture tank itself (the outer shell of the culture tank) a double-walled structure or more, reducing the amount of gas between each layer or reducing the pressure to a state where virtually no gas is present, and maintaining that state. Furthermore, the shape of the culture tank is not limited, but the upper (top) and lower (bottom) can be dome-shaped. This can improve pressure resistance. It can also improve the circulation performance of the gas (substrate). Furthermore, the intermediate part (body) located between the upper and lower parts can be cylindrical, for example.

[0052] The size of the culture vessel 10 is not limited, but as mentioned above, problems associated with the mixing of hydrogen and oxygen are less likely to become apparent in small-scale cultures, while these problems are more likely to become apparent in large-scale cultures. From this perspective, although the amount of culture medium 11 is not limited, for example, the effects of using this culture system and the culture method described above can be more easily obtained in cultures of 100 L or more. This amount of culture medium can be further set to 1,000 L or more, 5,000 L or more, or 10,000 L or more. On the other hand, the upper limit of the amount of culture medium is not limited, and can be, for example, 1,000,000 L or less. These upper and lower limits can be any combination of each other, and the numerical range specified by combining these upper and lower limits is also disclosed in the present invention. That is, for example, it can be 100 L or more and 1,000,000 L or less.

[0053] The culture tank 10 may be equipped with control means for controlling its internal environment (temperature, pressure, humidity, etc.). That is, for example, it may be equipped with temperature control means, pressure control means, humidity control means, etc. Of these, the temperature control means may include a heating mechanism (such as a heater jacket) and / or a cooling mechanism (such as a cooling mechanism using refrigerant circulation or a cooling mechanism using airflow). Furthermore, a pressurizing mechanism (such as a pressurizing pump) can be provided as a pressure control means. By providing a pressurizing mechanism, the amount of gaseous substrates such as hydrogen, oxygen, and carbon dioxide dissolved in the culture medium can be increased. Therefore, the culture efficiency of chemosynthetic autotrophic bacteria can be improved by pressurizing. A depressurization mechanism such as an exhaust valve or sub-tank can also be provided. A sub-tank refers to a culture area from which a portion of the gas contained in the culture area can be moved as the pressure rises. This movement allows for adjustment of the pressure within the culture area, but since the gas contained in the sub-tank is not exhausted to the outside, the gas in the sub-tank can also be reused. Furthermore, a dryer can be provided as a means of humidity control. In particular, when gas separation is used, a dryer can be provided in front of the gas separation mechanism.

[0054] The "gas supply means (3)" described above is a means for supplying hydrogen, oxygen, and carbon dioxide to the culture medium 11 containing chemosynthetic autotrophic bacteria. In other words, it is a means for supplying a gaseous substrate. Furthermore, as will be described later, the gas supply means 3 may also supply gases other than hydrogen, oxygen, and carbon dioxide. It can also be used in conjunction with the maintenance means 7. The configuration of the gas supply means 3 is not limited, but it may include a hydrogen supply mechanism 31, an oxygen supply mechanism 33, a carbon dioxide supply mechanism 35, and other gas supply means (means for supplying gases other than hydrogen, oxygen, and carbon dioxide). As mentioned above, the gas supply means 3 may be provided separately for each gas, but it may also be a supply means common to two or more gases. That is, examples include a gas supply means for supplying hydrogen and carbon dioxide, and a gas supply means for supplying oxygen and carbon dioxide. Only one of these may be used, or two or more may be used in combination. Furthermore, the configuration of each of the gas supply means described above is not limited, but each can be composed of, for example, a storage tank, a supply mechanism, a flow control mechanism, a flow pipe, etc. One of these may be used alone, or two or more may be used in combination.

[0055] Of the above, the storage tank is a tank that stores the gas that will be supplied to the culture tank. Although it is possible to connect the gas source and the supply mechanism directly without using a storage tank, using a storage tank makes the gas supply more stable. In other words, for example, it prevents the amount of gas to be supplied from falling below the required amount. If storage tanks are provided, each gas may have its own separate storage tank, or, for example, a storage tank 73 in which hydrogen and carbon dioxide coexist, a storage tank in which oxygen and carbon dioxide coexist, etc., can be provided. From the viewpoint of making it easier to manage individual flow rates, it is preferable to have separate storage tanks. On the other hand, if storage tanks are not provided, gas can be supplied by connecting the gas generation source and the culture tank through a flow pipe.

[0056] The supply mechanism is a mechanism for supplying the gas to be supplied to the culture tank, and can utilize, for example, a blower or a pump (gas pump). That is, for example, the gas can be supplied by a pump or blower. Alternatively, the gas can be supplied by liquefying and storing it, then gasifying it under reduced pressure. When a supply mechanism is provided, each gas may have its own separate supply mechanism, or, for example, a supply mechanism common to hydrogen and carbon dioxide, or a supply mechanism common to oxygen and carbon dioxide, etc., can be provided. From the viewpoint of making it easier to control individual flow rates, it is preferable to provide separate supply mechanisms.

[0057] A flow control mechanism is a mechanism that manages the amount of gas to be supplied, and can utilize, for example, valves, flow meters (gas flow meters), etc. When a flow control mechanism is provided, each gas may have its own separate supply mechanism, or, for example, a flow control mechanism common to hydrogen and carbon dioxide, or a flow control mechanism common to oxygen and carbon dioxide, etc., may be provided. From the viewpoint of facilitating individual flow control, it is preferable to provide separate flow control mechanisms.

[0058] A flow pipe is a structure for circulating the gas to be supplied. For example, a flow pipe can be interposed between a storage tank and a supply mechanism, or between a culture tank and a supply mechanism. As long as the gas can be circulated, the shape, material, size, etc. of the flow pipe are not limited and can be appropriately selected according to the culture content, but a pipe with a void that is connected to allow gas circulation inside can be used.

[0059] The supply of gas by the gas supply means may be performed directly or indirectly to the culture medium. An example of direct gas supply is directly bubbling the supply gas into the culture medium in the culture tank. An example of indirect gas supply is supplying gas (by bubbling, etc.) to the water or other components constituting the culture medium to form a liquid in which the substrate is dissolved, and then mixing this liquid with the culture medium to supply gas to the culture medium. Another example is supplying gas to the gas phase space (a space in the culture tank that is not filled with culture medium, etc.) when the culture tank has a gas phase space, and dissolving the substrate into the culture medium from the phase boundary between the gas phase space and the culture medium. One of these methods may be used alone, or two or more may be used in combination.

[0060] In the gas supply means, the supply amounts of hydrogen, oxygen, and carbon dioxide are not limited and can be supplied individually or in combination at appropriate concentrations. However, from the viewpoint of the growth efficiency of chemosynthetic autotrophic bacteria, it is preferable to supply these gases in a range where the proportion of each component to the total of the three gases is 50-90% by volume for hydrogen, 5-25% by volume for oxygen, and 5-25% by volume for carbon dioxide. This ratio can be further adjusted to 60-88% by volume for hydrogen, 6-20% by volume for oxygen, and 6-20% by volume for carbon dioxide; 70-86% by volume for hydrogen, 7-18% by volume for oxygen, and 7-18% by volume for carbon dioxide; or 75-85% by volume for hydrogen, 8-15% by volume for oxygen, and 8-15% by volume for carbon dioxide. Within these preferred ranges, chemosynthetic autotrophic bacteria can be produced more efficiently.

[0061] The "mixed gas recovery means (5)" described above is a means for recovering gases that are not dissolved in the culture medium 11 as a mixed gas. In other words, it is a means for recovering the excess gas (substrate) supplied to the culture medium 11 by the gas supply means 3. The configuration of the mixed gas recovery means 5 is not limited, but for example, it can consist of a recovery tank 51, a supply mechanism, a flow rate control mechanism, a flow pipe 53, etc. One of these may be used alone, or two or more may be used in combination.

[0062] Of the above, the recovery tank 51 is a tank in which gases not dissolved in the culture medium are recovered as a mixed gas. Specifically, if there is space above the culture tank (for example, if there is an upper void in the culture tank where no culture medium is stored), this space (the upper part of the culture tank) can be used as a recovery tank. Also, if there is no space above the culture tank (for example, if there is no upper void in the culture tank where no culture medium is stored), a recovery tank can be provided separately from the culture tank. In this case, for example, the mixed gas can be recovered into the recovery tank by connecting the upper end of the culture tank and the recovery tank with a flow pipe.

[0063] The supply mechanism 53 can be used as a mechanism for supplying the mixed gas to the recovery tank, or as a mechanism for supplying the gas recovered in the recovery tank to other parts (for example, a separator). Specifically, a blower, a pump (gas pump), etc., can be used. On the other hand, if a supply mechanism is not provided, supply can be performed by utilizing the positive pressure of the gas released from the culture medium. The flow control mechanism is a mechanism that manages the amount of mixed gas to be recovered, and can utilize, for example, valves, flow meters (gas flow meters), etc. A flow pipe is a structure for circulating a gas mixture, and as long as it allows gas to circulate, its shape, material, size, etc., are not limited, and a pipe or the like with a connected void that allows gas to circulate inside can be used.

[0064] The above-mentioned "maintenance means (7)" is a means for maintaining the gas mixture such that (A) the proportion of oxygen in the gas mixture is less than 5 volume%, or (B) the proportion of hydrogen in the gas mixture is less than 4 volume%. In other words, it is a means for maintaining the gas mixture recovered by the gas mixture recovery means so that it is in the state of (A) or (B) above.

[0065] The gas mixture recovered by the gas mixture recovery means 5 contains hydrogen, oxygen, and carbon dioxide (and other gases if other gases are supplied). When hydrogen and oxygen coexist in the gas mixture, the mixture may become explosive. Specifically, the gas mixture may become explosive in a wide concentration range, where hydrogen accounts for 4 to 95 volume percent and oxygen accounts for 5 volume percent or more relative to a total of 100 volume percent of hydrogen and oxygen. Therefore, by controlling the concentration of each gas in the gas mixture so that (A) and / or (B) are met, the concentration range in which it becomes explosive can be avoided.

[0066] The above (A) and (B) may be achieved in any way, but for example, they can be achieved by making the maintenance means 7 one of the means (S1) to (S3) below, or a combination of several of these means. (S1): A means of removing some or all of the oxygen contained in the recovered gas mixture to reduce the oxygen content in the gas mixture to less than 5% by volume. (S2): A method for mixing air with the recovered gas mixture to reduce the proportion of hydrogen in the gas mixture to less than 4% by volume. (S3): A means of adding another gas (a gas other than hydrogen and oxygen) to the recovered gas mixture to reduce the proportion of oxygen in the gas mixture to less than 5% by volume, and / or reduce the proportion of hydrogen in the gas mixture to less than 4% by volume.

[0067] When the maintenance means 7 functions as the means described in (S1) above, the maintenance means 7 can remove some or all of the oxygen contained in the recovered gas mixture to reduce the oxygen content in the gas mixture to less than 5% by volume. This oxygen removal operation can also be described as an operation to reduce the oxygen concentration. That is, the maintenance means can be used to reduce the oxygen concentration contained in the recovered gas mixture to achieve (A) above.

[0068] The configuration of the maintenance means when it functions as means (S1) is not limited, but for example, it can be configured by a gas separation mechanism 71. Specifically, this can be done by gas separation membranes (polyimide membranes, polyamide membranes, ceramic membranes, etc.), cryogenic separation, pressure swing adsorption (PSA), zeolite adsorption, chemical absorption using an alkaline solution, etc. One of these may be used alone, or two or more may be used in combination. Among these, a gas separation membrane (separation is possible by the pressure difference before and after the separation membrane) can be preferably used from the viewpoint of energy efficiency, suppression of CO2 generation, and simplicity of equipment.

[0069] When using a gas separation membrane, oxygen can be removed (the oxygen concentration reduced) by passing the recovered gas mixture through the membrane. For example, when a polyimide membrane is used as a gas separation membrane, the permeation rate of oxygen through the polyimide membrane is lower than that of hydrogen and carbon dioxide. On the other hand, as mentioned above, the recovered gas mixture is a gas with high concentrations of hydrogen, oxygen, and carbon dioxide. Therefore, the gas that permeates through the polyimide membrane (the gas on the permeation side of the polyimide membrane) can be obtained as a gas mixture with a low oxygen concentration (in particular, the proportion of oxygen in the gas mixture is less than 5% by volume). On the other hand, gases that permeate slowly through the polyimide membrane can be recovered as gases that have not permeated the polyimide membrane (gases on the non-permeable side of the polyimide membrane), thereby recovering a mixed gas with a high oxygen concentration. Therefore, this gas can be reused for oxygen supply in the gas supply means. Furthermore, oxygen removal (reduction of oxygen concentration) may be performed after the recovery of the gas mixture, but it may also be performed simultaneously with the recovery of the gas mixture, or before the recovery of the gas mixture.

[0070] When the maintenance means 7 is configured to function as means (S1), it can include other components besides the gas separation mechanism. Other components include a gas concentration meter 13 (such as an oxygen concentration meter). By using a gas concentration meter, it is possible to confirm that the proportion of oxygen in the gas mixture has fallen below 5 volume percent, or to predict that the proportion of oxygen in the gas mixture will fall below 5 volume percent. Furthermore, the gas mixture in which the oxygen concentration has been reduced to less than 5% by volume by means (S1) is rich in hydrogen and carbon dioxide, and can therefore be reused as a hydrogen source, a carbon dioxide source, etc. In this case, a supply mechanism, flow rate control mechanism, flow pipe, etc., can be provided for sending the gas mixture in which the oxygen content has been reduced to less than 5% by volume to the gas supply means. The details of these are the same as those of the configurations described for the gas supply means.

[0071] When the maintenance means 7 functions as the means described in (S2) above, the maintenance means 7 can be a means (such as an air supply mechanism) that mixes air (atmosphere) with the recovered gas mixture to reduce the proportion of hydrogen in the gas mixture to less than 4 volume percent. That is, air contains no hydrogen at all, or if it does, the amount of hydrogen is very small (for example, less than 1 volume ppm). Therefore, when air is mixed with the recovered gas mixture, the hydrogen concentration in the entire gas mixture with air added will decrease. Furthermore, while air supply may be performed after the recovery of the gas mixture, it may also be performed simultaneously with the recovery of the gas mixture, or before the recovery of the gas mixture. By supplying air simultaneously and / or before recovery, the risk of explosion of the gas mixture can be more reliably reduced.

[0072] In cases where air is supplied simultaneously, one configuration involves supplying air to the recovery path and, when gas is recovered in the recovery tank 51, controlling the system so that the proportion of hydrogen in the gas mixture is always less than 4 volume percent. Alternatively, in cases where air is supplied before recovery, one configuration involves supplying air in advance into the recovery tank 51 for recovering the gas mixture and, when gas is recovered in the recovery tank 51, controlling the system so that the proportion of hydrogen in the gas mixture is always less than 4 volume percent.

[0073] The configuration of the maintenance means 7 when it functions as means (S2) is not limited, but it can be configured, for example, with an air supply mechanism, a gas concentration meter (such as a hydrogen concentration meter), etc. Among these, the air supply mechanism can be, for example, a blower, a pump (gas pump), etc. Furthermore, by using a gas concentration meter, it is possible to confirm that the proportion of hydrogen in the gas mixture has fallen to less than 4 volume percent, or to predict that the proportion of hydrogen in the gas mixture will fall to less than 4 volume percent. That is, by monitoring the concentration of hydrogen contained in the gas mixture to be recovered, the amount of air required to reduce the proportion of hydrogen in the gas mixture to less than 4 volume percent can be calculated.

[0074] When the maintenance means 7 functions as the means described in (S3) above, the maintenance means 7 can be a means (other gas supply mechanism 75, etc.) that adds another gas (a gas other than hydrogen and oxygen) to the recovered mixed gas to reduce the oxygen concentration to less than 5 volume% and / or the hydrogen concentration to less than 4 volume% by adding another gas (a gas supply mechanism 75, etc.). As described above, the means described in (S2), i.e., the means of mixing air with the recovered mixed gas, makes it easy to reduce the hydrogen concentration to less than 4 volume%, but tends to make it difficult to reduce the oxygen concentration to less than 5 volume%. This is because air contains oxygen. In this respect, when a gas other than hydrogen and oxygen is mixed, it is easier to achieve both of the above simultaneously. In other words, the other gases can either contain no hydrogen or oxygen at all, or if they do contain them, their content can be extremely small (for example, less than 1 ppm by volume each). Therefore, when other gases are mixed with the recovered gas mixture, both the hydrogen and oxygen concentrations in the overall gas mixture will decrease. Furthermore, the supply of other gases may be carried out after the recovery of the gas mixture, simultaneously with the recovery of the gas mixture, or before the recovery of the gas mixture. By supplying other gases simultaneously and / or before the recovery, the risk of explosion of the gas mixture can be more reliably reduced.

[0075] In the case of simultaneously supplying other gases, one configuration is to supply other gases to the recovery path and, when the gas is recovered in the recovery tank 51, control the system so that the ratio of hydrogen and oxygen in the mixed gas is always below a predetermined value. Alternatively, in the case of supplying other gases before recovery, one configuration is to supply other gases in advance into the recovery tank 51 for recovering the mixed gas and, when the gas is recovered in the recovery tank 51, control the system so that the ratio of hydrogen and oxygen in the mixed gas is always below a predetermined value.

[0076] The configuration of the maintenance means when it functions as a means (S3) is not limited, but for example, it can be configured with an other gas supply mechanism 75 (a mechanism for supplying other gases), a gas concentration meter (hydrogen concentration meter, oxygen concentration meter, instrument capable of measuring the concentration of other gases, etc.). Among these, the other gas supply mechanism can be, for example, a blower, a pump (gas pump), etc. Furthermore, by using a gas concentration meter, it is possible to confirm that the ratio of hydrogen and oxygen in the mixed gas has fallen below a predetermined value, or to predict that the ratio of hydrogen and oxygen in the mixed gas will fall below a predetermined value. That is, by monitoring the concentration of hydrogen and / or oxygen contained in the mixed gas to be recovered, it is possible to calculate the amount of other gas necessary to make the ratio of hydrogen and / or oxygen in the mixed gas fall below a predetermined value.

[0077] While the type of gas other than hydrogen and oxygen is not limited, from the viewpoint of preferring less reactive gases, examples include noble gases (helium, neon, argon, krypton, xenon), nitrogen (free nitrogen), and carbon dioxide. These may be used individually or in combination of two or more. Among these, nitrogen and / or carbon dioxide are preferred from a cost standpoint, and nitrogen is more preferred from the viewpoint of the culture efficiency of chemosynthetic autotrophic bacteria. That is, carbon dioxide is more soluble in water than other gases, and when dissolved, it tends to lower the pH of the aqueous solution. For this reason, using carbon dioxide as a means to achieve (A) and / or (B) tends to easily alter the environment of the culture medium. In this respect, nitrogen does not have such disadvantages, is less reactive, is cost-effective, and is therefore preferable from various viewpoints.

[0078] This system enables safe operation while maintaining high culture efficiency for chemosynthetic autotrophic bacteria. Specifically, increasing the culture volume for efficient bacterial cultivation increases the amount of hydrogen and oxygen used. Consequently, the amount of gas mixture recovered also increases, raising the risk of explosion. However, this system allows the recovery gas mixture to be continuously controlled outside the explosion range by the maintenance means 7. Furthermore, this control can be achieved through low-cost and simple methods such as oxygen removal, air introduction, and nitrogen filling.

[0079] In addition to the gas supply means 3, mixed gas recovery means 5, and maintenance means 7 described above, this system may also include other means to the extent that the effects of the present invention are achieved. Only one type of other means may be used, or two or more types may be used in combination. Furthermore, the other means may be added before the gas supply means, between the above-described means, or after the maintenance means. Other means include pH adjustment means, temperature adjustment means, culture medium recovery means, and culture medium replenishment means. These may be used individually or in combination of two or more. Of the above, the pH adjustment means is a means of adjusting the pH of the culture medium to a pH suitable for culturing chemosynthetic autotrophic bacteria. Generally, the pH of the culture medium tends to become acidic during cultivation, so for example, a neutralization operation can be performed using ammonia (ammonia water, etc.). Of the above, the temperature control means is a means of adjusting the temperature of the culture medium to a temperature suitable for culturing chemosynthetic autotrophic bacteria. Of the above, the culture medium recovery means is a means of removing chemosynthetic autotrophic bacteria that have grown through cultivation from the culture medium. That is, since the culture efficiency tends to decrease as bacteria grow in the culture medium, it is preferable to perform operations to reduce the bacterial density continuously or periodically from the viewpoint of maintaining high culture efficiency. Of the above, the culture medium replenishment means is a means for replenishing the culture medium. In addition to the substrate supplied as a gas, the culture medium may contain various substrates as described above. These components are consumed during cultivation and can be supplied as needed, but it is also possible to replenish with a new culture medium that has these components pre-formulated.

[0080] This system allows for the safe and efficient cultivation of chemosynthetic autotrophic bacteria while maintaining high cultivation efficiency. In other words, organic matter (carbohydrates, amino acids, proteins, etc.) produced by these bacteria can be obtained safely and efficiently through the cultivation of chemosynthetic autotrophic bacteria. This system can be widely used in various fields such as resin (plastic) manufacturing, fiber manufacturing, fuel manufacturing, reagent manufacturing, industrial raw material manufacturing, food ingredient manufacturing, and feed manufacturing (fish and shellfish feed, poultry feed, pig feed, dairy cow feed, beef cattle feed). [Examples]

[0081] The present invention will be described below by several embodiments. [Culture System 1A] Culture system 1A (see Figure 1) comprises a culture tank 10, a gas supply means 3, a mixed gas recovery means 5, and a maintenance means 7. This culture system 1A is a system that supplies a mixed gas of hydrogen and carbon dioxide, and / or carbon dioxide, to a recovery tank (the gas containment space within the culture tank) to maintain an oxygen ratio of less than 5% by volume or a hydrogen ratio of less than 4% by volume.

[0082] The culture tank 10 is constructed as a stainless steel pressure-resistant tank that can be sealed. The culture tank 10 is also capable of containing 10 L or more of culture medium 11, and has a gas containment space 51 above the area containing the culture medium 11. In this example, the gas containment space 51 functions as the recovery tank 51 of the mixed gas recovery means 5. In other words, a culture medium layer and a gas layer exist inside the culture tank 10.

[0083] Furthermore, the culture tank 10 is equipped with a gas concentration meter 15 capable of measuring the concentration of dissolved gases (hydrogen, oxygen, and carbon dioxide) in the culture medium, and a gas concentration meter 13 capable of measuring the concentration of a mixed gas (hydrogen, oxygen, and carbon dioxide) contained in the gas containment space. In addition, it is equipped with a pH meter for measuring the pH of the culture medium, a pressure gauge for measuring the pressure inside the culture tank 10, and a vent (pressure relief valve) to avoid excessive pressure. Furthermore, the culture tank 10 is equipped with a stirring mechanism 19 (stirring blade) at its bottom, enabling stirring of the culture medium 11. Stirring increases and stabilizes the dissolved amount of the substrate supplied in gaseous form. The stirring blade can be rotated at, for example, 50 rpm to 1500 rpm.

[0084] The culture medium (100 vol%) contains hydrogen-oxidizing bacteria (Cupriavidus necator strain H16), disodium hydrogen phosphate dodecahydrate (0.56 vol%), potassium dihydrogen phosphate (0.06 vol%), magnesium sulfate heptahydrate (0.01 vol%), calcium chloride (0.0005 vol%), ammonium chloride (0.12 vol%), iron(III) chloride hexahydrate (0.0004 vol%), and nickel(II) chloride hexahydrate (0.009 vol%).

[0085] The gas supply means 3 includes three supply mechanisms: a hydrogen supply mechanism 31, an oxygen supply mechanism 33, and a carbon dioxide supply mechanism 35. Of these, the hydrogen supply mechanism 31 comprises a hydrogen supply source (not shown) and a hydrogen gas flow pipe 311. The flow pipe 311, which extends from the hydrogen tank, extends to the culture medium containment area in the culture tank 10 and is open at the bottom of the culture tank 10. That is, hydrogen gas can be released from the depths of the contained culture medium 11. In this example, a hydrogen tank filled with compressed purified hydrogen is used as the hydrogen supply source. Furthermore, the hydrogen can be recycled.

[0086] In this example, the oxygen supply mechanism 33 utilizes air (atmosphere) as the oxygen source. Specifically, the oxygen supply mechanism 33 includes an oxygen separator 331 (first oxygen separator) for concentrating or separating oxygen contained in the air, and an oxygen gas flow pipe 333. As the oxygen separator 331 (oxygen generator), an oxygen generator that removes nitrogen by physical adsorption is used. Specifically, one end of the oxygen gas flow pipe 333 is connected to the upstream side of the oxygen separator 331, and the other end is left open near the bottom of the culture tank 10, so that oxygen gas can be released from deep within the culture medium 11 contained in the culture tank 10. The remaining oxygen after separation is discharged from the flow pipe 334.

[0087] In this example, the carbon dioxide supply mechanism 35 utilizes combustion exhaust gas as a carbon dioxide source. Specifically, the carbon dioxide supply mechanism 35 consists of a carbon dioxide separator 351 for concentrating or separating carbon dioxide contained in the combustion exhaust gas, a carbon dioxide storage tank 353, and a carbon dioxide gas flow pipe 355. A polyimide gas separation membrane (polyimide membrane) is used as the carbon dioxide separator 351. The permeation rate of carbon dioxide through the polyimide membrane is greater than that of oxygen and nitrogen. Therefore, it is possible to obtain a separated gas with a high concentration of carbon dioxide (it is also possible to obtain a gas that contains virtually only carbon dioxide). This concentrated carbon dioxide gas is used as the carbon dioxide source. Specifically, one end of the carbon dioxide gas flow pipe 355 is connected to the upstream side of the carbon dioxide separator 351, and the other end is left open near the bottom of the culture tank 10, thereby allowing carbon dioxide gas to be released from deep within the culture medium 11 contained in the culture tank 10.

[0088] Furthermore, the carbon dioxide separator 351 is also connected to the carbon dioxide storage tank 353, and a portion of the carbon dioxide obtained in the carbon dioxide separator 351 is stored in the carbon dioxide storage tank 353. The carbon dioxide storage tank 353 is equipped with a carbon dioxide gas flow pipe 355 that can supply carbon dioxide to the gas storage space 51 (recovery tank 51) above the culture medium 11 without passing through the culture medium 11. In other words, the carbon dioxide supply mechanism 35 has two routes: a route for supplying carbon dioxide to the culture medium 11 (first route) and a route for supplying it to the gas storage space 51 above the culture medium 11 (second route).

[0089] The second of these routes is used to fill the gas containment space 51 with carbon dioxide during the initial stages of system operation. Furthermore, if the proportion of oxygen and / or hydrogen in the mixed gas contained in the gas containment space 51 increases, carbon dioxide is supplied to the gas containment space 51 using the second route. This allows the proportion of oxygen in the mixed gas contained in the gas containment space 51 to be less than 5 volume%, and the proportion of hydrogen to be less than 4 volume%. In other words, the second route functions as both a carbon dioxide supply mechanism and a maintenance means 7 that functions as the means described in (S3) above.

[0090] Furthermore, as described above, the gas containment space 51 of the culture tank 10 functions as the recovery tank 51 of the mixed gas recovery means 5, and the recovery tank 51 is connected to the oxygen separator 71 (which is a second oxygen separator different from the first oxygen separator of the oxygen supply mechanism, but has the same structure), so that oxygen can be removed from the recovered mixed gas. In other words, the second oxygen separator 71 functions as the maintenance means 7 that functions as the means (S1) described above.

[0091] Furthermore, the second oxygen separator 71 is connected to the mixed gas storage tank 73. The mixed gas storage tank 73 is a tank for storing the mixed gas (a mixed gas with high concentrations of hydrogen and carbon dioxide, or consisting substantially only of these gases) from which oxygen has been removed via the second oxygen separator 71. Furthermore, a mixed gas flow pipe 751 extends from the mixed gas storage tank 73, allowing the mixed gas (a mixed gas with high concentrations of hydrogen and carbon dioxide, or consisting substantially only of these gases) to be supplied to the gas containment space 51 above the culture medium 11 without passing through the culture medium 11. In addition, the flow pipe extending from the mixed gas storage tank 73 also extends into the culture tank and is open near the bottom of the culture tank. That is, the mixed gas flow pipe 753 extends so that the mixed gas (a mixed gas with high concentrations of hydrogen and carbon dioxide, or consisting substantially only of these gases) can be released from the depths of the culture medium contained in the culture tank. As a result, hydrogen and carbon dioxide can be circulated and reused within the system as either a diluent gas and / or a substrate supply gas without exhausting them outside the system.

[0092] In this example, the aforementioned gas supply means 3 can be, for example, two independent supply mechanisms: a hydrogen and carbon dioxide supply mechanism and an oxygen supply mechanism, or two independent supply mechanisms: a hydrogen supply mechanism and an oxygen and carbon dioxide supply mechanism.

[0093] [Culture System 1B] Culture system 1B (see Figure 2) comprises a culture tank 10, a gas supply means 3, a mixed gas recovery means 5, and a maintenance means 7. This culture system 1B is a system that supplies air to the recovery tank (the gas-containing space within the culture tank) to maintain an oxygen ratio of less than 5% by volume or a hydrogen ratio of less than 4% by volume.

[0094] The culture tank 10 is similar to the recovery tank 51 in the culture system 1A, and is capable of containing 10 L or more of culture medium 11 (with the same composition and proportions of the culture medium). Above the area containing the culture medium 11 is a gas containment space 51. In this example, the gas containment space 51 functions as the recovery tank 51 of the mixed gas recovery means 5. That is, a culture medium layer and a gas layer exist within the culture tank 10.

[0095] Furthermore, the culture tank 10, like the culture system 1A, is equipped with a gas concentration meter 15 for measuring the concentration of dissolved gases (hydrogen, oxygen, and carbon dioxide) in the culture medium, and a gas concentration meter 13 for measuring the concentration of a mixed gas (hydrogen, oxygen, and carbon dioxide) contained in the gas containment space. The culture tank 10 is also equipped with a stirring mechanism 19 (stirring blade) at its bottom.

[0096] The gas supply means 3 includes three supply mechanisms: a hydrogen supply mechanism 31, an oxygen supply mechanism 33, and a carbon dioxide supply mechanism 35. Of these, the hydrogen supply mechanism 31 comprises a hydrogen supply source and a hydrogen gas flow pipe 311. The flow pipe 311, which extends from the hydrogen tank, extends to the culture medium containment area within the culture tank 10 and is open at the bottom of the culture tank 10. That is, hydrogen gas can be released from deep within the contained culture medium 11. In this example, there are two hydrogen sources. One is a hydrogen tank (not shown) filled with compressed purified hydrogen, and the other is a hydrogen separator 53b that separates the hydrogen contained in the recovered mixed gas. In other words, the hydrogen supply mechanism 31 can utilize both purified hydrogen and the hydrogen contained in the recovered mixed gas.

[0097] The oxygen supply mechanism 33 is the same as the oxygen supply mechanism 33 in culture system 1A, and uses air (atmosphere) as an oxygen source. The oxygen supply mechanism 33 includes an oxygen separator 331 (first oxygen separator) for concentrating or separating oxygen contained in the air, and an oxygen gas flow pipe 333. Similar to culture system 1A, the oxygen separator 331 uses a polyimide gas separation membrane (polyimide membrane). The remaining oxygen after separation is discharged through the flow pipe 334.

[0098] The carbon dioxide supply mechanism 35 utilizes combustion exhaust gas and carbon dioxide contained in the recovered mixed gas as carbon dioxide sources. In other words, it is similar to the carbon dioxide supply mechanism 35 in culture system 1A in that it uses combustion exhaust gas as a carbon dioxide source, but the carbon dioxide supply mechanism 35 in culture system 1B differs from the carbon dioxide supply mechanism 35 in culture system 1A in that it also utilizes carbon dioxide contained in the recovered mixed gas as a carbon dioxide source.

[0099] The carbon dioxide supply mechanism 35 includes a carbon dioxide separator 351 (first carbon dioxide separator) for concentrating or separating carbon dioxide contained in combustion exhaust gas, and a carbon dioxide gas flow pipe 355. Similar to culture system 1A, the carbon dioxide separator 351 uses a polyimide gas separation membrane (polyimide membrane). One end of the carbon dioxide gas flow pipe 355 is connected to the upstream side of the carbon dioxide separator 351, and the other end is left open near the bottom of the culture tank 10, allowing carbon dioxide gas to be released from deep within the culture medium 11 contained in the culture tank 10.

[0100] On the other hand, the carbon dioxide supply mechanism 35 in culture system 1B differs from the carbon dioxide supply mechanism 35 in culture system 1A, as mentioned above, in that it also utilizes the carbon dioxide contained in the recovered mixed gas as a carbon dioxide source. Specifically, the carbon dioxide supply mechanism 35 in culture system 1B is connected to the carbon dioxide separator 53a (second carbon dioxide separator) of the mixed gas recovery means 5. The configuration of the carbon dioxide separator 53a is the same as that of the carbon dioxide separator 351, and uses a polyimide gas separation membrane (polyimide membrane). The carbon dioxide contained in the mixed gas recovered by the mixed gas recovery means 5 is returned to the carbon dioxide supply mechanism 35 and reused in the culture tank 10.

[0101] Furthermore, as mentioned above, the gas containment space 51 of the culture tank 10 functions as the recovery tank 51 of the mixed gas recovery means 5, and as mentioned above, the recovery tank 51 is connected to a carbon dioxide separator 53a (a second oxygen separator different from the first carbon dioxide separator 351 of the carbon dioxide supply mechanism 35, but with the same structure) and a hydrogen separator 53b. This allows carbon dioxide and hydrogen to be extracted from the recovered mixed gas and reused.

[0102] Furthermore, the configuration of the hydrogen separator 53b is the same as that of the oxygen separator 331, and a polyimide gas separation membrane (polyimide membrane) is used. Since the permeation rate of hydrogen through the polyimide membrane is greater than that of oxygen and carbon dioxide, it is possible to obtain a separated gas with a high hydrogen concentration (it is also possible to obtain a gas that contains virtually only hydrogen). By refluxing this hydrogen-concentrated gas to the hydrogen supply mechanism 31, hydrogen can be reused.

[0103] Furthermore, the culture system 1B includes an air circulation pipe 75 as a maintenance means 7 that can supply air to the gas containment space 51 above the culture medium 11. The amount of air supplied can be controlled by a blower and a flow meter (not shown). That is, the type and amount of gas contained in the gas containment space 51 can be monitored by the gas concentration meter 13, and air can be introduced into the gas containment space 51 through the air circulation pipe 75 as needed to maintain an oxygen percentage of less than 5 volume% or a hydrogen percentage of less than 4 volume%.

[0104] In this example, the aforementioned gas supply means 3 can be, for example, two independent supply mechanisms: a hydrogen and carbon dioxide supply mechanism and an oxygen supply mechanism, or two independent supply mechanisms: a hydrogen supply mechanism and an oxygen and carbon dioxide supply mechanism.

[0105] [Culture System 1C] Culture system 1C (see Figure 3) comprises a culture tank 10, a gas supply means 3, a mixed gas recovery means 5, and a maintenance means 7. This culture system 1C is a system that supplies carbon dioxide to a recovery tank (the gas-containing space within the culture tank) to maintain an oxygen ratio of less than 5% by volume or a hydrogen ratio of less than 4% by volume.

[0106] The culture tank 10 is similar to the recovery tank 51 in the culture system 1A, and is capable of containing 10 L or more of culture medium 11 (with the same composition and proportions of the culture medium). Above the area containing the culture medium 11 is a gas containment space 51. In this example, the gas containment space 51 functions as the recovery tank 51 of the mixed gas recovery means 5. That is, a culture medium layer and a gas layer exist within the culture tank 10.

[0107] Furthermore, the culture tank 10, like the culture system 1A, is equipped with a gas concentration meter 15 for measuring the concentration of dissolved gases (hydrogen, oxygen, and carbon dioxide) in the culture medium, and a gas concentration meter 13 for measuring the concentration of a mixed gas (hydrogen, oxygen, and carbon dioxide) contained in the gas containment space. The culture tank 10 is also equipped with a stirring mechanism 19 (stirring blade) at its bottom.

[0108] The gas supply means 3 includes three supply mechanisms: a hydrogen supply mechanism 31, an oxygen supply mechanism 33, and a carbon dioxide supply mechanism 35. Of these, the hydrogen supply mechanism 31 comprises a hydrogen supply source and a hydrogen gas flow pipe 311. The flow pipe 311, which extends from the hydrogen tank, extends to the culture medium containment area within the culture tank 10 and is open at the bottom of the culture tank 10. That is, hydrogen gas can be released from deep within the contained culture medium 11. In this example, there are two hydrogen sources. One is a hydrogen tank (not shown) filled with compressed purified hydrogen, and the other is a hydrogen separator 53b that separates the hydrogen contained in the recovered mixed gas. That is, the hydrogen supply mechanism 31 can utilize both purified hydrogen and the hydrogen contained in the recovered mixed gas. Furthermore, the hydrogen can be recycled.

[0109] The oxygen supply mechanism 33 is the same as the oxygen supply mechanism 33 in culture system 1A, and uses air (atmosphere) as an oxygen source. The oxygen supply mechanism 33 includes an oxygen separator 331 for concentrating or separating oxygen contained in the air, and an oxygen gas flow pipe 333. Similar to culture system 1A, the oxygen separator 331 uses a polyimide gas separation membrane (polyimide membrane). The remaining oxygen after separation is discharged through the flow pipe 334.

[0110] The carbon dioxide supply mechanism 35 utilizes combustion exhaust gas and carbon dioxide contained in the recovered mixed gas as carbon dioxide sources. In other words, it is similar to the carbon dioxide supply mechanism 35 in culture system 1A in that it uses combustion exhaust gas as a carbon dioxide source, but the carbon dioxide supply mechanism 35 in culture system 1C differs from the carbon dioxide supply mechanism 35 in culture system 1A in that it also utilizes carbon dioxide contained in the recovered mixed gas as a carbon dioxide source.

[0111] The carbon dioxide supply mechanism 35 includes a carbon dioxide separator 351 for concentrating or separating carbon dioxide contained in the combustion exhaust gas and carbon dioxide contained in the recovered mixed gas, and a carbon dioxide gas flow pipe 355. Similar to culture system 1A, the carbon dioxide separator 351 uses a polyimide gas separation membrane (polyimide membrane). One end of the carbon dioxide gas flow pipe 355 is connected to the upstream side of the carbon dioxide separator 351, and the other end is left open near the bottom of the culture tank 10, allowing carbon dioxide gas to be released from deep within the culture medium 11 contained in the culture tank 10.

[0112] On the other hand, the carbon dioxide supply mechanism 35 in culture system 1C differs from the carbon dioxide supply mechanism 35 in culture system 1A, as mentioned above, in that it also utilizes the carbon dioxide contained in the recovered mixed gas as a carbon dioxide source. Specifically, the carbon dioxide supply mechanism 35 in culture system 1C is connected to recirculate the gas discharged as a residue in the hydrogen separator 53b of the mixed gas recovery means 5 to the upstream side of the carbon dioxide separator 351.

[0113] The configuration of the hydrogen separator 53b is the same as that of the oxygen separator 331, and uses a polyimide gas separation membrane (polyimide membrane). Since the permeation rate of hydrogen through the polyimide membrane is greater than that of oxygen and carbon dioxide, it is possible to obtain a separated gas with a high hydrogen concentration (it is also possible to obtain a gas that contains substantially only hydrogen). By refluxing this hydrogen-concentrated gas to the hydrogen supply mechanism 31, hydrogen can be reused. In other words, the gas containment space 51 of the culture tank 10 functions as the recovery tank 51 of the mixed gas recovery means 5, and this recovery tank 51 is connected to the hydrogen separator 53b. This allows hydrogen to be extracted from the recovered mixed gas and reused.

[0114] Furthermore, the culture system 1C includes a carbon dioxide flow pipe 77 as a maintenance means 7, which supplies carbon dioxide to the gas containment space 51 above the culture medium 11. The amount of carbon dioxide supplied can be controlled by a blower and a flow meter (not shown). That is, the type and amount of gas contained in the gas containment space 51 are monitored by a gas concentration meter 13, and carbon dioxide is introduced into the gas containment space 51 through the carbon dioxide flow pipe 77 as needed, thereby maintaining an oxygen percentage of less than 5 volume% or a hydrogen percentage of less than 4 volume%.

[0115] In this example, the aforementioned gas supply means 3 can be, for example, two independent supply mechanisms: a hydrogen and carbon dioxide supply mechanism and an oxygen supply mechanism, or two independent supply mechanisms: a hydrogen supply mechanism and an oxygen and carbon dioxide supply mechanism. [Industrial applicability]

[0116] According to the present invention, chemosynthetic autotrophic bacteria can be cultured safely while maintaining high culture efficiency. In other words, organic matter (carbohydrates, amino acids, proteins, etc.) produced by chemosynthetic autotrophic bacteria can be obtained safely and efficiently through the culture of these bacteria. The culture apparatus 1 of the present invention can be widely used in various fields such as resin (plastic) manufacturing, fiber manufacturing, fuel manufacturing, reagent manufacturing, industrial raw material manufacturing, food ingredient manufacturing, and feed (fish and shellfish feed, poultry feed, pig feed, dairy cow feed, beef cattle feed). [Explanation of Symbols]

[0117] 1; Culture system, 1A; Culture system, 1B; Culture system, 1C; Culture system, 10;Culture tank, 11;Culture solution, 13; Gas concentration meter, 15; Gas concentration meter, 19; stirring mechanism, 3; means of supplying gas, 31; Hydrogen supply mechanism, 311; Distribution pipe, 33; Oxygen supply mechanism, 331; Separator, 333; Flow pipe, 334; Flow pipe, 35; Carbon dioxide supply mechanism, 351; Separator, 353; Storage tank, 355; Flow pipe, 5; means for recovering mixed gases, 51; Recovery tank (gas containment space), 53; Feeding mechanism; 53a; Separator; 53b; Separator; 7; Maintenance means, 71; Separator, 73; Storage tank, 75; Distribution pipe, 751; Distribution pipe, 753; Distribution pipe, 77; Distribution pipe.

Claims

1. A method for culturing chemosynthetic autotrophic bacteria using hydrogen and oxygen, A gas supply process that supplies hydrogen, oxygen, and carbon dioxide to a culture medium containing chemosynthetic autotrophic bacteria, A mixed gas recovery step for recovering gases not dissolved in the culture medium as a mixed gas, A method for culturing chemosynthetic autotrophic bacteria, comprising a maintenance step of maintaining the mixed gas so that it is in state A or B as described below. A: The proportion of oxygen in the mixed gas is less than 5% by volume. B: The proportion of hydrogen in the mixed gas is less than 4% by volume.

2. The method for culturing chemosynthetic autotrophic bacteria according to claim 1, wherein the maintenance step is a step of removing some or all of the oxygen contained in the mixed gas to reach A.

3. The method for culturing chemosynthetic autotrophic bacteria according to claim 1, wherein the maintenance step is a step of mixing air with the mixed gas to reach B.

4. The method for culturing chemosynthetic autotrophic bacteria according to claim 1, wherein the maintenance step is a step of adding a gas other than hydrogen and oxygen to the mixed gas to achieve A and / or B.

5. The method for culturing chemosynthetic autotrophic bacteria according to claim 2, wherein the oxygen to be removed is oxygen separated from the mixed gas via a gas separation membrane.

6. A method for culturing chemosynthetic autotrophic bacteria according to claim 1, wherein hydrogen and / or carbon dioxide separated from the mixed gas via a gas separation membrane is used in the gas supply step.

7. The method for culturing chemosynthetic autotrophic bacteria according to claim 1, wherein the gas supply step is a step of supplying each gas in a ratio of 50 to 90% by volume for hydrogen, 5 to 25% by volume for oxygen, and 5 to 25% by volume for carbon dioxide, relative to the total of hydrogen, oxygen, and carbon dioxide.

8. A culture system for chemosynthetic autotrophic bacteria using hydrogen and oxygen, A gas supply means for supplying hydrogen, oxygen, and carbon dioxide to a culture medium containing chemosynthetic autotrophic bacteria, A mixed gas recovery means for recovering gases not dissolved in the culture medium as a mixed gas, A culture system for chemosynthetic autotrophic bacteria, comprising a maintenance means for maintaining the mixed gas such that it is in state A or B as described below. A: The proportion of oxygen in the mixed gas is less than 5% by volume. B: The proportion of hydrogen in the mixed gas is less than 4% by volume.

9. The cultivation system for chemosynthetic autotrophic bacteria according to claim 8, wherein the maintenance means is a means for removing some or all of the oxygen contained in the mixed gas to achieve A.

10. The cultivation system for chemosynthetic autotrophic bacteria according to claim 8, wherein the maintenance means is a means of mixing air with the mixed gas to reach B.

11. The culture system for chemosynthetic autotrophic bacteria according to claim 8, wherein the maintenance means is a means of adding a gas other than hydrogen and oxygen to the mixed gas to achieve A and / or B.

12. The culture system for chemosynthetic autotrophic bacteria according to claim 9, wherein the oxygen to be removed is oxygen separated from the mixed gas via a gas separation membrane.

13. A culture system for chemosynthetic autotrophic bacteria according to claim 8, wherein hydrogen and / or carbon dioxide separated from the mixed gas via a gas separation membrane is used in the gas supply means.

14. The culture system for chemosynthetic autotrophic bacteria according to claim 8, wherein the gas supply means is a means of supplying each gas in a ratio of 50 to 90% by volume for hydrogen, 5 to 25% by volume for oxygen, and 5 to 25% by volume for carbon dioxide, relative to the total of hydrogen, oxygen, and carbon dioxide.

Images