Gas-phase microbial reactions

Abstract

The present disclosure provides a conversion of a gaseous substrate with the use of a microorganism. In one aspect, the present disclosure provides a microbial gas-phase reaction system for converting a gaseous substrate with the use of a microorganism. This microbial gas-phase reaction system comprises at least one member selected from among a carrier having the microorganism immobilized thereon, a gas supply part for supplying the gaseous substrate to the gas phase of the microbial gas-phase reaction system, and a water supply system for supplying water to the carrier. In one aspect, the present disclosure provides a method for converting a gaseous substrate with the use of a microorganism. This method comprises a step for exposing a surface of a carrier, on which the microorganism is immobilized, to a gas phase containing the gaseous substrate.

Description

Technical Field

[0001] The present disclosure relates to the conversion of gaseous substrates by microorganisms.

Background Art

[0002] Microorganisms can carry out chemical reactions under mild conditions without requiring large-scale equipment as compared with chemical reactions by physicochemical methods through various enzymatic reactions. Therefore, the production of compounds using microorganisms has been studied.

[0003] When converting a gaseous substrate using a microorganism, not only is it necessary to input a large amount of energy for aeration and stirring in the supply of the substrate, but also the reaction efficiency is low because the dissolution efficiency of hydrophobic gaseous substances is low.

Summary of the Invention

Means for Solving the Problems

[0004] As a result of intensive research, the present inventors have achieved efficient and / or stable conversion of gaseous substrates using microorganisms. Therefore, the present disclosure provides a method for converting a gaseous substrate using a microorganism, and a gas-phase microbial reaction system for converting a gaseous substrate using a microorganism.

[0005] Therefore, the present disclosure provides the following. (Item 1) A gas-phase reaction chamber comprising a gas supply unit for continuously or intermittently supplying a gaseous substrate from the outside, A carrier installed in the gas-phase reaction chamber or a carrier constituting at least a part of the wall of the gas-phase reaction chamber, which is water-permeable or water-absorbent but the surface on the gas phase side is not partially flooded, and the microorganism cells are fixed so that at least a part of the microorganism cells are in direct contact with the gas phase in the gas-phase reaction chamber, A gas discharge unit connected to the gas-phase reaction chamber, A gas-phase microbial reaction system comprising the above. (Item 2) The gas-phase microbial reaction system according to item 1, further comprising a moisture supply system for supplying moisture from outside the gas-phase reaction chamber to the extent that the microbial cells fixed to the carrier do not dry out. (Item 3) The aforementioned water supply system is configured such that the water content of the carrier is 1-50% (v / v), and is one of the gas-phase microbial reaction systems according to the above items. (Item 4) A gas-phase microbial reaction system according to any of the above items, further comprising a mechanism for recovering products resulting from the conversion of the gaseous substrate by the microorganisms. (Item 5) A gas-phase microbial reaction system according to any of the above items, wherein the gas supply unit is configured to passively take in the gaseous substrate by diffusion from outside the gas-phase reaction chamber. (Item 6) A gas-phase microbial reaction system according to any of the above items, wherein the carrier is a thin film. (Item 7) A gas-phase microbial reaction system according to any of the above items, further comprising a liquid discharge section connected to the gas-phase reaction chamber. (Item 8) A gas-phase microbial reaction system according to any of the above items, wherein the moisture supply system is configured to supply the moisture to the carrier as a liquid stream. (Item 9) A gas-phase microbial reaction system according to any of the above items, wherein the water supply system includes an aqueous phase chamber through which a liquid flows, the carrier is positioned between the aqueous phase chamber and the gas-phase reaction chamber, and the carrier is positioned such that the surface of the carrier on which the microorganisms are immobilized faces the gas-phase reaction chamber. (Item 9A) A gas-phase microbial reaction system according to any of the above items, wherein the carrier is configured to separate the gas-phase reaction chamber from the aqueous-phase chamber. (Item 9B) A gas-phase microbial reaction system according to any of the above items, wherein the flow path from the supply to the discharge of the gaseous substrate is configured in a spiral shape. (Item 9C) A gas-phase microbial reaction system according to any of the above items, wherein the flow path from the supply to the discharge of the gaseous substrate is configured to follow the surface of the carrier. (Item 10) A gas-phase microbial reaction system according to any of the above items, wherein the reaction vessel includes a plurality of gas phases and a plurality of liquid phases, and is configured such that the gas phases and liquid phases are alternately stacked. (Item 11) A gas-phase microbial reaction system according to any of the above items, wherein the carrier includes a plurality of tubular members, the plurality of tubular members are bundled together in close contact to form a columnar unit, or the plurality of tubular members are arranged at intervals from each other within a sheath tube to form a columnar unit including the plurality of tubular members and the sheath tube, and the plurality of tubular members and the sheath tube constitute the wall of the gas-phase reaction chamber or the aqueous-phase chamber. (Item 12) A gas-phase microbial reaction system according to any of the above items, further comprising a rotating mechanism connected to the carrier, wherein the rotating mechanism comprises a rotating shaft to which the carrier is fixed. (Item 13) A gas-phase microbial reaction system according to any of the above items, wherein the moisture supply system is configured to supply the moisture to the carrier by capillary action. (Item 14) A gas-phase microbial reaction system according to any of the above items, wherein the moisture supply system is configured to supply the moisture to the carrier in an aerosol or gaseous state. (Item 15) The moisture supply system is configured such that the moisture in a gaseous state condenses on the carrier, and is a gas-phase microbial reaction system according to any of the above items. (Item 16) A gas-phase microbial reaction system according to any of the above items, further comprising a power source for driving the aforementioned rotating mechanism. (Item 17) A method for converting a gaseous substrate using microorganisms, The steps include supplying a gaseous substrate to the gas-phase reaction chamber from the outside, either continuously or intermittently, In the gas-phase reaction chamber, exposing at least a part of the surface of the microorganism-fixed carrier in the carrier to the gas phase in the gas-phase reaction chamber A method comprising (Item 18) The method according to any one of the above items, further comprising the step of supplying moisture to the microorganism-supported carrier from outside the gas-phase reaction chamber. (Item 19) The method according to any one of the above items, controlling the moisture content of the carrier to be 1 to 50%. (Item 20) [[ID=,14]]A carrier for fixing microorganisms, or a carrier on which microorganisms are fixed, A gas-phase reaction chamber, or a gas-phase reaction chamber member configured to provide the gas-phase reaction chamber by assembling, A gas supply unit for continuously or intermittently supplying a gaseous substrate from the outside, A gas discharge unit A kit comprising The gas-phase reaction chamber may be provided in a state including at least one of the carrier, the gas supply unit, and the gas discharge unit, or may be provided separately from the carrier, the gas supply unit, and the gas discharge unit. The gas-phase reaction chamber is configured to accommodate the carrier. The gas-phase reaction chamber is configured such that the microorganisms fixed on the carrier come into contact with the gaseous substrate. Kit. (Item 21) The kit according to any one of the above items, further comprising a moisture supply system for supplying moisture to the carrier from outside the gas-phase reaction chamber. (Item 22) The kit according to any one of the above items, further comprising a support configured to support the carrier.

[0006] In the present disclosure, it is intended that the above one or more features may be provided in further combinations in addition to the explicitly stated combinations. Further embodiments and advantages of the present disclosure will be recognized by those skilled in the art upon reading the following detailed description as needed.

Advantages of the Invention

[0007] The present disclosure can provide efficient and / or stable conversion of gaseous substrates using microorganisms, and thus can be advantageously used for removal of unwanted gaseous substances and / or acquisition of desired products. In addition, the conversion of gaseous substrates according to the present disclosure can provide an energy-saving gas-phase microbial reaction system that is efficient and stable.

Brief Description of the Drawings

[0008] [Figure 1] This is a specific configuration example of the bioreactor (gas-phase microbial reaction system) of the present disclosure. [Figure 2] (A) It is a top view from the upper part of the bioreactor of FIG. 1, showing a configuration in which gas is supplied from one side line and discharged through the other side line. (B) It is a cross-sectional view from the side of the bioreactor of FIG. 1. [Figure 3] It shows continuous methane decomposition by the bioreactor of FIG. 1 using Methylococcus capsulatus (Bath). The vertical axis indicates the methane decomposition rate (mL methane / hr / mL gas phase). (A) The result when air containing 20% methane is supplied at a flow rate of 0.5 mL / min. The horizontal axis indicates the cell density (dry weight g / L gas phase). (B) The result when air containing 20% methane is supplied and microorganisms with a cell density of 33 g-dry weight / L gas phase are used. The horizontal axis indicates the gas flow rate (mL / min). (C) The result when air containing 20% methane is supplied at a flow rate of 0.5 mL / min. The immobilized microorganism concentration used is 33, 31, 28, or 25 g-dry weight / L gas phase as indicated by each symbol in the graph. The horizontal axis indicates the supplied methane concentration (v / v%). [Figure 4]Figure 1 shows the results of methane decomposition and methane-to-methanol conversion when air containing 20% ​​methane was supplied at a flow rate of 0.1 mL / min to a bioreactor using Methylococcus capsulatus (Bath). The results of methane decomposition and methanol production are shown when the feed solution composition was changed stepwise at 0-3 hours (12.9 mM phosphate buffer), 3-6 hours (0.1 × inorganic nitrate (NMS) medium + 10 mM formic acid), 6-9 hours (0.1 × inorganic nitrate (NMS) medium + 10 mM formic acid + 1 μM cyclopropanol), and 9-12 hours (0.1 × inorganic nitrate (NMS) medium + 10 mM formic acid + 10 μM cyclopropanol). The vertical axis shows the residual methane concentration in the exhaust gas (%v / v) and the accumulated methanol concentration in the liquid phase chamber (μM). The horizontal axis shows time (hr). [Figure 5] Figure 1 shows the effect of the presence of a liquid phase on the methane decomposition rate by Methylococcus capsulatus (Bath) in the bioreactor. (A) Results when air containing 22.8% (v / v) methane is supplied at a flow rate of 0.1 mL / min. The horizontal axis shows the reaction time. The vertical axis shows the methane concentration at the gas supply section (△), the methane concentration at the gas discharge section when the aqueous phase chamber is left empty (〇), and the methane concentration at the gas discharge section when water is added to the aqueous phase chamber (□), respectively. (B) Results when air containing 22.8% (v / v) methane is supplied at flow rates of 0.05, 0.1, and 0.2 mL / min and the aqueous phase chamber is left empty (gas phase), and when air containing 22.8% (v / v) methane is supplied at a flow rate of 0.1 mL / min and water is added to the aqueous phase chamber. The vertical axis shows the methane decomposition rate (mL methane / hr / mL gas phase). [Figure 6] This is a specific example of the configuration of the bioreactor (gas-phase microbial reaction system) described herein. [Figure 7](A) A side view of the outer and inner units of the bioreactor shown in Figure 6, which is one embodiment of the gas-phase reaction chamber of the present disclosure. (1): Upper cover of the bioreactor (equipped with ports for injecting gaseous substrate and water vapor). (2): Lower cover of the bioreactor. (3): Connector for assembling (2) and (4). (4): Liquid collection chamber. (5): Motor for rotating the inner unit (motion mechanism). (6): Bundle of rod-shaped members for supporting (7) (support for the carrier). (7): Fan composed of a carrier on which microorganisms are immobilized (carrier). (8): Aqueous solution. (B) A side view of the outer unit of the bioreactor shown in Figure 6. (9): Port for injecting gaseous substrate and water vapor (gas supply unit, water supply system). (10): Port for discharging and / or sampling gas (gas discharge unit). (11): Port for discharging and / or sampling product-containing liquid (product recovery mechanism). (C) A side view of the inner unit of the bioreactor shown in Figure 6. (12): Motor for rotating the inner unit (corresponding to (5) of (A)). (13): Bundle of rod-shaped members for supporting (14) (corresponding to (6) of (A)). (14): Fan composed of a carrier on which microorganisms are immobilized (corresponding to (7) of (A)). (15): Member for supplying water to the carrier using capillary action (part of the water supply system). (16): Aqueous solution supply port (part of the water supply system). (17): Aqueous solution (corresponding to (8) of (A)). [Figure 8] Figure 6 shows a specific example of the fan section of the inner unit of the bioreactor. (A) A photograph from above. (B) A photograph from the side. (C) A side view. (D) A perspective view. (E) A bird's-eye view. (F) A side view of the fan section's axis. (G) A perspective view of the fan section's axis. [Figure 9] Figure 6 shows a specific example of the fan section of the inner unit of the bioreactor. (A) A photograph from above. (B) A photograph from the side. (C) A side view. (D) A perspective view. (E) A bird's-eye view. (F) A side view of the fan section's axis. (G) A perspective view of the fan section's axis. [Figure 10]Figure 6 shows the continuous methane decomposition using a bioreactor with Methylococcus capsulatus (Bath). The upper panel shows the results when using a bioreactor equipped with a 10-blade fan (dry cell weight 1.4g). The lower panel shows the results when using a bioreactor equipped with a 20-blade fan (dry cell weight 1.3g). The horizontal axis of each graph shows the supply rate of the methane-air mixed gas (mL / min). The vertical axis of the graphs in the left column shows the methane concentration at the outlet (v / v%). The vertical axis of the graphs in the right column shows the methane decomposition rate (mL methane / hr / mL gas phase). [Figure 11] Figure 6 shows the results of methane decomposition in a bioreactor using Methylococcus capsulatus (Bath). The results were obtained when air containing 20% ​​(v / v) methane was supplied at a flow rate of 8 mL / min. The horizontal axis represents the reaction time. The vertical axis represents the methane concentration at the gas supply section (□), the methane concentration at the gas outlet section when using a 10-blade fan (◇), and the methane concentration at the gas outlet section when using a 20-blade fan (△), respectively. [Figure 12] (A) A specific configuration example of a part of the bioreactor (gas-phase microbial reaction system) of the present disclosure, which has a gas phase and a liquid phase stacked on top of each other. (B) A unit of the bioreactor of (A) is shown. A carrier is placed between the gas phase and the liquid phase. [Figure 13](A) This is a specific configuration example of a bioreactor (gas-phase microbial reaction system) of the present disclosure, which comprises a carrier including a plurality of tubular members. The plurality of tubular members are arranged at intervals from each other within a sheath tube, and the plurality of tubular members and the sheath tube form a columnar unit. In one embodiment, the interior of the plurality of tubular members functions as an aqueous phase chamber, and the exterior of the plurality of tubular members and the interior of the sheath tube functions as a gas-phase reaction chamber. In another embodiment, the interior of the plurality of tubular members functions as a gas-phase reaction chamber, and the exterior of the plurality of tubular members and the interior of the sheath tube functions as an aqueous phase chamber. (B) This shows a carrier formed by bundling a plurality of tubular members with a hexagonal cross-section (honeycomb) in close contact with each other to form a columnar unit. This shows that the interior of some of the tubular members functions as an aqueous phase chamber, and the interior of some of the tubular members functions as a gas-phase reaction chamber. (C) This shows one embodiment of the operation of the bioreactor of (A), in which a liquid phase is formed inside the tubular members and a gas phase is formed outside the tubular members. (D)(A) shows one embodiment of the operation of the bioreactor, in which a gas phase is formed inside the tubular member and a liquid phase is formed outside the tubular member. [Figure 14] This is a schematic diagram of an exemplary embodiment of the gas-phase microbial reaction system of the present disclosure. (1) gas-phase reaction chamber, (2) carrier (located within and / or constituting at least a portion of the wall of the gas-phase reaction chamber), (3) gas supply unit, (4) gaseous substrate, (5) moisture supply system (if necessary), (6) moisture (if necessary), (7) space within the gas-phase reaction chamber containing the gas phase, (8) gas, (9) gas discharge unit. [Figure 15] (A) An overhead view of the bioreactor of Example 6, showing a configuration in which gas is supplied from one line and discharged from the other line. (B) A side cross-sectional view of the bioreactor of Example 6. [Figure 16] This is a schematic diagram of the bioreactor of Example 6 when a methane-utilizing bacteria immobilization filter is placed between the gas phase and the liquid phase, with the side on which the bacteria are immobilized facing the gas phase. [Figure 17]This is a schematic diagram of the bioreactor of Example 6 when a methane-utilizing bacteria immobilization filter is placed between the gas phase and the liquid phase, with the side on which the bacteria are immobilized facing the liquid phase. [Figure 18] This graph shows the results of methane-to-methanol conversion when the filter is positioned so that the side with the fixed bacteria faces either the gas phase (A) or the liquid phase (B). The vertical axis shows the residual methane concentration in the exhaust gas (%v / v) and the accumulated methanol concentration in the liquid phase chamber (mM). The horizontal axis shows time (hr). [Figure 19] This is a schematic diagram of the bioreactor of Example 6, in which a methane-utilizing bacteria immobilization filter is placed between the gas phase and the liquid phase, with the side on which the bacteria are immobilized facing the gas phase, and the operation is performed continuously for a long period of time. [Figure 20] The results of long-term methane-to-methanol conversion in the bioreactor of Example 6 are shown. The vertical axis shows the residual methane concentration (%v / v) in the exhaust gas and the methanol concentration (mM) in the effluent extracted from the liquid phase. The horizontal axis shows time (hr). [Figure 21] This is a schematic diagram of the system used in Example 7. [Figure 22] The results of toluene decomposition in the bioreactor of Example 7 are shown. The vertical axis represents the toluene removal rate (%) at the bioreactor outlet. The horizontal axis represents time (min). [Modes for carrying out the invention]

[0009] The present disclosure is described below in best form. Throughout this specification, singular expressions should be understood to include the concept of their plural form unless otherwise specified. Accordingly, singular articles (e.g., "a," "an," "the" in English) should be understood to include the concept of their plural form unless otherwise specified. Furthermore, terms used herein should be understood to have the meaning commonly used in the art unless otherwise specified. Accordingly, unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this disclosure pertains. In case of any conflict, this specification (including definitions) shall prevail.

[0010] The following provides definitions of terms used specifically in this specification and / or basic technical concepts as appropriate.

[0011] (Definition, etc.) In this specification, "microorganisms" means organisms that include not only bacteria and archaea, but also microalgae, yeasts, and mosses.

[0012] In this specification, “substrate” refers to any substance utilized by a microorganism or an enzyme secreted by a microorganism, and typically refers to a substance that is converted into another substance.

[0013] In this specification, "conversion" of a substrate by microorganisms refers to the transformation of a substrate substance into another substance. In one embodiment, the conversion results in the transformed substance as a product, and the recovery of this product is intended. In one embodiment, the conversion is decomposition, and the recovery of the transformed substance is not intended.

[0014] In this specification, "products" of microorganisms refer to any substance produced by microorganisms, and include the microbial cells themselves, intracellular components, extracellular components, cell membranes, cell walls, secretions, etc., as well as preparations obtained by operations such as lysis and purification of these.

[0015] In this specification, "moisture content" refers to the volume percentage of water in an object. Moisture content can be measured by any suitable method, but for example, it can be calculated by determining the volume of absorbed water using the specific gravity from the weight difference before and after drying the object, and then dividing by the volume of the object. In this specification, the moisture content of a carrier refers to the moisture content of the carrier itself, excluding microorganisms (for example, before microorganisms are immobilized).

[0016] In this specification, "carrier" means a part for immobilizing microorganisms or the like, which is installed in the gas-phase reaction chamber of this disclosure or constitutes part of the wall of the gas-phase reaction chamber of this disclosure. The carrier may be made of any material that can immobilize microorganisms, and preferably contains a small amount of water or is configured to allow water to pass through.

[0017] In this specification, "support for the carrier" refers to the part for fixing the carrier, and is also called the carrier holder. The support for the carrier is used to fix the carrier inside or on the wall of a gas-phase reaction chamber so that at least a portion of the immobilized microorganisms are exposed to the gas phase.

[0018] In this specification, the term "liquid phase" refers to a space in which liquids make up the majority of the three forms of matter: gas, liquid, and solid. The liquid phase in this specification also includes spaces in which gases (e.g., bubbles in a gaseous substrate) or solids are dispersed in a liquid.

[0019] In this specification, the term "gas phase" refers to a space in which gases constitute the majority of the three forms of matter: gases, liquids, and solids. In this specification, the gas phase also includes spaces in which liquids (e.g., water) are dispersed within a gas.

[0020] In this specification, “continuous supply” of a substance means that a substance (e.g., a gas (e.g., a substrate in a gaseous state), a liquid (e.g., water, a substrate in a liquid state)) is continuously delivered to the target. In this specification, “intermittent supply” of a substance (also referred to as “semi-intermittent supply” herein) means that a substance (e.g., a gas (e.g., a substrate in a gaseous state), a liquid (e.g., water, a substrate in a liquid state)) is delivered to the target intermittently. In one embodiment, continuously or intermittently delivering a gaseous substance into a gas-phase reaction chamber from the outside through piping or an intake (e.g., a gas supply unit) is also a continuous or intermittent supply of a substance to the gas phase in the gas-phase reaction chamber. In one embodiment, continuously vaporizing a liquid substrate and continuously delivering the gaseous substrate into the gas-phase reaction chamber through piping or an intake (e.g., a gas supply unit) is also a continuous supply.

[0021] In this specification, "immersion" of the carrier means a state in which at least a portion of the surface of the carrier facing the gas-phase reaction chamber is submerged in water.

[0022] In this specification, "not to the extent that it dries up" means a state in which there is no water dripping from the outside of the cell, and only the water that the cell naturally contains is maintained. The surface of the cell may be wet, but not so wet that water droplets are falling from it.

[0023] In this specification, "partially open system" for microbial transformation means that the microorganism is not exposed to the complete external environment, but is placed within some kind of enclosure such as a chamber, and that substances (e.g., gases (e.g., substrates in a gaseous state), liquids (e.g., water, substrates in a liquid state, nutrients, inhibitors, inducers, etc.)) are continuously or intermittently delivered to the microorganism from the outside. Alternatively, "partially open system" means that a portion of the chamber is open, and through this opening, continuous or intermittent exchange of substances with the external environment occurs.

[0024] In this specification, "gas-phase microbial reaction system" means a configuration that performs the conversion of a gaseous substrate using the microorganisms of this disclosure, and originally refers to a system or organization for achieving a purpose, in which multiple elements are systematically configured and interact with each other, and multiple various devices are configured to communicate with each other as needed.

[0025] In this specification, "gas supply unit" means the part that supplies a gaseous substrate to the gas phase in the gas-phase microbial reaction system of this disclosure.

[0026] In this specification, “moisture supply system” means the part of the gas-phase microbial reaction system of this disclosure that supplies moisture to a carrier on which microorganisms are immobilized.

[0027] In this specification, "gas discharge section" means the section in the gas-phase microbial reaction system of this disclosure that removes gas from the gas-phase reaction chamber.

[0028] In this specification, “liquid discharge section” means the section in the gas-phase microbial reaction system of this disclosure that removes liquid from the gas-phase reaction chamber.

[0029] In this specification, “kit” means a unit in which the parts to be provided (e.g., parts of the gas-phase microbial reaction system of this disclosure, microorganisms, instructions, etc.) are provided, usually divided into two or more compartments. It is advantageous that the kit preferably includes instructions or manuals describing how to use or operate the provided parts (e.g., parts of the gas-phase microbial reaction system of this disclosure, microorganisms, etc.).

[0030] In this specification, "program" is used in the ordinary sense used in this art, referring to a sequence of instructions for a computer to perform a particular task, treated as a "thing" under Japanese patent law, and sometimes referred to as a "program product" to clarify that it is perceptible or tangible. All computers operate according to programs. In modern computers, programs can be represented as data, stored on recording media or storage devices, or provided from the cloud.

[0031] In this specification, “recording medium” refers to a recording medium that stores a program causing the Disclosure to be executed. The recording medium may be any type of recording medium that can record a program, is computer-readable, and allows the program read by that recording medium to be executed and implemented on other devices such as computers. For example, it may be, but is not limited to, an external storage device such as a ROM, HDD, magnetic disk, or flash memory such as a USB memory that can be stored internally.

[0032] In this specification, the term "approximately" refers to plus or minus 10% of the indicated value unless otherwise specified. When "approximately" is used for temperature, it refers to plus or minus 5°C of the indicated temperature; when "approximately" is used for pH, it refers to plus or minus 0.5 of the indicated pH.

[0033] (Preferred embodiment) Preferred embodiments of the Disclosure are described below. The embodiments provided below are provided for a better understanding of the Disclosure, and it will be understood that the scope of the Disclosure should not be limited to the descriptions below. Accordingly, it will be obvious that those skilled in the art can make appropriate modifications within the scope of the Disclosure, taking into consideration the descriptions herein. It will also be understood that the embodiments of the Disclosure below can be used individually or in combination.

[0034] In one aspect, this disclosure provides the conversion of gaseous substrates by microorganisms. Any means to achieve this are intended to be within the scope of this disclosure. For example, even without explicit description, a description of the conditions for the conversion of gaseous substrates by microorganisms should be understood to simultaneously disclose various forms of means for carrying out this disclosure, such as parts of a gas-phase microbial reaction system for achieving those conditions and steps in a method for achieving those conditions. For ease of understanding, a gas-phase microbial reaction system may be described as a whole, but characteristic parts thereof may also be within the scope of this disclosure. Also for ease of understanding, parts of a gas-phase microbial reaction system may be described along with their connections and / or positional relationships, but the parts do not need to actually be assembled in those connections and / or positional relationships; they may be within the scope of this disclosure if they are configured to be assembled in those connections and / or positional relationships. The conversion of gaseous substrates by microorganisms in this disclosure may be industrial gaseous substrate treatment or household gaseous substrate treatment (e.g., toilet odor control products).

[0035] In one aspect, the present disclosure provides a gas-phase microbial reaction system for converting a gaseous substrate by microorganisms. This gas-phase microbial reaction system comprises at least a gas-phase reaction chamber providing a gas-phase space for converting the gaseous substrate into another substance, a carrier on which microorganisms are immobilized so as to be in direct contact with the gas in the chamber, and a gas supply unit for supplying the gaseous substrate to the gas phase of the gas-phase microbial reaction system. The operation of the gas-phase microbial reaction system may or may not require a power source. A gas-phase microbial reaction system requiring a power source may be switched between operating and non-operating states by toggling a switch. In one embodiment, the gas-phase microbial reaction system of the present disclosure is a partially open system, and the gas-phase reaction chamber on which microorganisms convert the gaseous substrate includes supply channels (e.g., a gas supply unit, a water supply system) for continuously or intermittently supplying substances (e.g., gaseous substrate, water) from outside the gas-phase reaction chamber into the gas-phase reaction chamber. In one embodiment, the supply channel may be in gaseous and / or liquid communication with a gas-phase reaction chamber in which microorganisms convert a gaseous substrate, or it may be configured to be switchable between gaseous and / or liquid communication and a blocked state. In one embodiment, the gas-phase microbial reaction system includes a gas discharge section and / or liquid discharge section separate from this supply channel. In one embodiment, a moisture supply system (e.g., including a moisture supply channel) is provided for supplying moisture to the carrier from outside the gas-phase reaction chamber.

[0036] In one aspect, the present disclosure provides a method for converting a gaseous substrate by microorganisms. This method includes the step of exposing at least a portion of the surface on which microorganisms are immobilized on a carrier to a gaseous phase containing a gaseous substrate. In one embodiment, the method of the present disclosure is carried out in a partially open system, and the reaction in which microorganisms convert a gaseous substrate is carried out in a gaseous reaction chamber in which substances (e.g., gaseous substrate, water) are supplied from outside the gaseous reaction chamber into the gaseous reaction chamber.

[0037] In one embodiment, a gaseous substrate is supplied to the gas phase, and the gaseous substrate is delivered to microorganisms via the gas phase. The gaseous substrate may be supplied to the gas phase by vaporization of a liquid or sublimation of a solid, or it may be supplied to the gas phase in a gaseous state through piping or the like. In one embodiment, the gaseous substrate is supplied to the gas phase continuously or intermittently. In one embodiment, the present disclosure (e.g., the system or method of the present disclosure) may have a predetermined period during which the gaseous substrate is continuously supplied to the gas phase for a certain duration, for example, 1 minute or more, 5 minutes or more, 10 minutes or more, 20 minutes or more, 30 minutes or more, 45 minutes or more, 1 hour or more, 5 hours or more, 10 hours or more, 20 hours or more, 1 day or more, 3 days or more, 5 days or more, or 7 days or more. In one embodiment, the present disclosure (e.g., the system or method of the present disclosure) may include periods during which the gaseous substrate is not supplied to the gas phase (interrupted), for example, periods of 1 second or more, 10 seconds or more, 30 seconds or more, 1 minute or more, 5 minutes or more, 10 minutes or more, 20 minutes or more, 30 minutes or more, 45 minutes or more, 1 hour or more, 2 hours or less, 5 hours or less, 12 hours or less, or 24 hours or less. In one embodiment, an airflow (e.g., a unidirectional airflow) may be generated in the gas phase to actively deliver the gaseous substrate to the microorganism. In one embodiment, the gaseous substrate may be delivered to the microorganism by passive diffusion from the ambient air.

[0038] In one embodiment, the supply of gaseous substrate to the gas phase may be controlled. Any suitable amount of gaseous substrate can be supplied to the gas phase, for example, 0.001 mL / hr or more, 0.01 mL / hr or more, 0.1 mL / hr or more, 0.5 mL / hr or more, 1 mL / hr or more, 5 mL / hr or more, 10 mL / hr or more, 50 mL / hr or more, 100 mL / hr or more, 100 mL / hr or more, 500 mL / hr or more, 5000 mL / hr or more, 10000 mL / hr or more, or 10000 mL / hr or more per 1 L volume of the gas phase. In another embodiment (e.g., a passive diffusion embodiment), the net supply of gas to the gas phase may be zero, and only the diffusion of the internal substance may occur.

[0039] In one embodiment, microorganisms are immobilized on a carrier. By immobilizing microorganisms on a carrier, chemical reactions can be carried out in a gas phase where there is no liquid to suspend the microorganisms. In one embodiment, microorganisms are immobilized on the surface of a carrier. In one embodiment, microorganisms are immobilized on the surface of a carrier that is exposed to the gas phase. In one embodiment, the carrier itself can partition the gas phase reaction chamber and the aqueous phase chamber, and a structure can be adopted in which the surface of the carrier on which the microorganisms are immobilized faces the gas phase reaction chamber. In this case, compared to a method in which the carrier is floated between the liquid phase and the gas phase, for example, there is no part where the liquid phase and the gas phase are in direct contact, so direct evaporation of the liquid is suppressed, and mass transfer between gas and liquid without going through the microorganism layer is also suppressed, and it is thought that the efficiency of substrates coming into contact with microorganisms is improved. In one embodiment, the carrier is placed between the gas phase and the liquid phase. For example, if a carrier placed between a gas phase and a liquid phase can supply water to the gas phase-side surface of the carrier, but is permeable to water to the extent that the gas phase side of the carrier is not immersed, then a layered structure in which the gas phase and liquid phase are alternately arranged with the carrier in between, or an aggregate structure in which the gas phase and liquid phase are arranged inside and outside each of multiple hollow columnar carriers, can be formed, thereby improving the area per unit volume that can be used for the conversion reaction of the gaseous substrate.

[0040] In one embodiment, when a carrier is placed between the gas phase and the liquid phase, liquid water is not supplied to the carrier from the gas phase side. For example, this can be achieved by employing a structure in which the carrier is surrounded by a liquid-impermeable material, and / or a structure in which there are no liquid-permeable materials or channels other than the carrier between the gas-phase reaction chamber and the aqueous-phase chamber. In one embodiment, the thickness of the carrier placed between the gas-phase reaction chamber and the aqueous-phase chamber can be, for example, 0.01-100 mm, 0.01-50 mm, 0.01-10 mm, 0.01-5 mm, 0.01-1 mm, 0.01-0.5 mm, 0.01-0.1 mm, 0.05-100 mm, 0.05-50 mm, 0.05-10 mm, 0.05-5 mm, 0.05-1 mm, 0.05-0.5 mm, 0.1-100 mm, 0.1-50 mm, 0.1-10 mm, 0.1-5 mm, 0.1-1 mm, 0.5-100 mm, 0.5-50 mm, 0.5-10 mm, 0.5-5 mm, 1-100 mm, 1-50 mm, 1-10 mm, 5-100 mm, 5-50 mm, etc. In one embodiment, the carrier includes a plurality of tubular members. The plurality of tubular members may be bundled together in close contact to form a columnar unit, or they may be arranged at intervals within the outer tube to form a columnar unit including the plurality of tubular members and the outer tube. When a gas phase is formed inside the tubular members, the tubular members can be said to constitute the wall of the gas-phase reaction chamber, and when a liquid phase is formed inside the tubular members, the tubular members can be said to constitute the wall of the aqueous-phase chamber. Furthermore, when a gas phase is formed between the tubular members and the outer tube, the tubular members and the outer tube can be said to constitute the wall of the gas-phase reaction chamber, and when a liquid phase is formed between the tubular members and the outer tube, the tubular members and the outer tube can be said to constitute the wall of the aqueous-phase chamber.

[0041] In one embodiment, the carrier is placed in the gas phase. In one embodiment, the conversion rate of the gaseous substrate (the reduction rate of the discharged gaseous substrate relative to the supplied gaseous substrate) can be improved by lengthening the flow path from the supply to the discharge of the gaseous substrate, even if the volume of the gas phase is the same. In one embodiment, the conversion rate of the gaseous substrate can be improved by configuring the flow path so that the flow of the supplied gas becomes turbulent (for example, by selecting the shape, surface properties, etc.), even if the volume of the gas phase is the same. In one embodiment, the conversion rate of the gaseous substrate can be improved even if the volume of the gas phase is the same by minimizing the volume of the gas phase that is not in contact with the immobilized microorganisms. In one embodiment, the conversion rate of the gaseous substrate can be improved even if the volume of the gas phase is the same by increasing the specific surface area of ​​the microbial immobilization surface of the carrier that is in contact with the gas phase. In one embodiment, the conversion rate of the gaseous substrate can be improved even if the volume of the gas phase is the same by stirring the gas in the gas phase, but since the energy required to stir the gas is much less than the energy required to stir the liquid, a gas phase stirring mechanism may be used.

[0042] In one embodiment, water is supplied to the carrier. Unless otherwise specified herein, water means any form of water molecules, including not only liquid water but also gaseous water (water vapor) and solid water (ice). In one embodiment, water is supplied to the surface of a carrier on which microorganisms are immobilized. This embodiment also includes cases in which water is introduced from one side of a thick carrier, passes through the interior of the carrier, and is supplied to microorganisms immobilized on the other side. The passage of water through the interior of the carrier may be by passive transport using gravity and capillary action, by active transport through the application of water pressure and manipulation such as centrifugal force, or by utilizing the lift generated by the vaporization of water. In one embodiment, water is supplied to the carrier as a liquid stream. In one embodiment, water is supplied to the carrier via an aerosol or gaseous state (e.g., dispersed in the gas phase). In one embodiment, water is supplied to the carrier by condensation of gaseous water on the carrier. For example, the condensation of water is controlled by the material and / or structure of the carrier, the temperature of the carrier and / or the gas phase, the amount of water in the gas phase, and the amount of water supplied from the water supply system.

[0043] In one embodiment, the water content of the carrier during the conversion of a gaseous substrate by the microorganisms of this disclosure may be 70% or less, 60% or less, 50% or less, 40% or less, 30% or less, 20% or less, 15% or less, 10% or less, 7% or less, 5% or less, or 3% or less. Many microorganisms require water for their survival or to perform their functions. In one embodiment, the water content of the carrier during the conversion of a gaseous substrate by the microorganisms of this disclosure may be 0.01% or more, 0.05% or more, 0.1% or more, 0.5% or more, or 1% or more. In one embodiment, the water content of the carrier during the conversion of the gaseous substrate by the microorganisms of the present disclosure is 0.01-70%, 0.01-60%, 0.01-50%, 0.01-40%, 0.01-30%, 0.01-20%, 0.01-10%, 0.01-7%, 0.01-5%, 0.1-60%, 0.1-20%, 0.1-10%, The moisture content may be 0.1-7%, 0.1-5%, 0.5-60%, 0.5-20%, 0.5-10%, 0.5-7%, 0.5-5%, 1-60%, 1-20%, 1-10%, 1-7%, 1-5%, 2-60%, 2-20%, 2-10%, 2-7%, 2-5%, 3-60%, 3-20%, 3-10%, 3-7%, or 3-5%. On the other hand, a lower moisture content may increase the frequency with which microorganisms come into contact with the gaseous substrate, thereby increasing the conversion rate of the gaseous substrate. Therefore, controlling the moisture content may improve the stability and / or efficiency in the conversion of the gaseous substrate in this disclosure. In particular, when the gaseous substrate is continuously supplied, water loss from the carrier can easily occur, so controlling the moisture content may be advantageous. In one embodiment, microorganisms are immobilized on a carrier such that at least a portion of the microbial cells are in direct contact with the gas phase in a gas-phase reaction chamber without being submerged in water.

[0044] In one embodiment, the water content of the carrier during the conversion of a gaseous substrate by the microorganisms of the present disclosure can be controlled by adjusting the amount of water present in the gas phase, selecting the material and / or structure of the carrier (e.g., hydrophilic / hydrophobic balance, utilization of capillary action, carrier thickness), designing the channels for supplying water to the carrier (including channels for supplying water by capillary action), adjusting the carrier's movement speed, adjusting the contact level between the carrier and the liquid phase and / or adjusting the water pressure, adjusting the temperature of the gas phase and / or the carrier, etc.

[0045] In one embodiment, additive components such as nutrients utilized by microorganisms (e.g., low molecular weight water-soluble substances such as organic acids, trace elements, salts, and amino acids), agents that regulate metabolism (e.g., inhibitors and inducers of specific metabolic pathways), components that enhance the activity of the microorganisms used, surfactants, drying agents, components for maintaining microorganisms for extended periods, components that suppress the growth of other microorganisms, excipients, and antioxidants may be delivered to the microorganisms by the movement of water in a liquid state. For example, a portion of the water channel to the location of the microorganisms may be in direct or indirect contact (e.g., via piping or a water-permeable material) with an additive component storage unit (which may be in a solid state or prepared at a high concentration), or the liquid phase containing the additive components may be brought into contact with a carrier on which the microorganisms are immobilized. Here, the water channel may or may not have a water flow in a constant direction, and as long as there is fluid communication from the additive component storage unit to the location of the microorganisms, the additive components can be delivered to the microorganisms by diffusion.

[0046] In one embodiment, the carrier is water-permeable or water-absorbent. Examples of water-permeable or water-absorbent carrier forms include, but are not limited to, filters, fibers, sponges, and other porous materials. In one embodiment, the carrier is configured to allow water to pass through by capillary action. For example, the water permeability or water absorption of the carrier can be prepared by the material and / or structure of the carrier. In one embodiment, the carrier is made of a material and surface structure having a water contact angle of 0°~90°, 10°~90°, 30°~90°, 50°~90°, 70°~90°, 0°~70°, 10°~70°, 30°~70°, 50°~70°, 0°~50°, 10°~50°, 30°~50°, 0°~30°, 10°~30°, or 0°~10°. In one embodiment, the carrier is made of a material having a surface structure such that the contact angle with water is 0°~90°, 10°~90°, 30°~90°, 50°~70°, 0°~50°, 10°~50°, 30°~50°, 0°~30°, 10°~30°, or 0°~10°. 3 This density has a weight of 0.01-2g, 0.02-2g, 0.05-2g, 0.1-2g, 0.2-2g, 0.5-2g, 0.01-1g, 0.02-1g, 0.05-1g, 0.1-1g, 0.2-1g, 0.5-1g, 0.01-0.5g, 0.02-0.5g, 0.05-0.5g, 0.1-0.5g, 0.01-0.2g, 0.02-0.2g, 0.05-0.2g, 0.1-0.2g, 0.01-0.1g, 0.02-0.1g, or 0.05-0.1g per unit.

[0047] The material of the carrier is not particularly limited as long as it can immobilize microorganisms. Examples include carbon fiber (PAN-based, pitch-based, phenolic resin-based, etc.), glass fiber, cellulose fiber, polyethylene resin, polypropylene resin, polyurethane resin, polystyrene resin, polyvinyl chloride resin, polyvinyl acetate resin, polyvinyl alcohol resin, polyethylene glycol resin, acrylic resin, ceramics, silicon, metal, charcoal, activated carbon, nonwoven fabric, and composites thereof. Examples of carrier shapes include fan-shaped, hollow columnar (hollow cylindrical, hollow hexagonal columnar, etc.), tubular, cubic, rectangular, columnar, spherical, disc-shaped, sheet-shaped, membrane-shaped, and bead-shaped.

[0048] Microorganisms can be immobilized on a carrier by any suitable method, for example, by passing a microbial suspension through a filter carrier, spraying a microbial suspension onto a carrier, immersing a carrier in a microbial suspension, or physically and chemically binding microorganisms to a carrier using a linker (e.g., a bifunctional reagent, antibody, etc.). For information on microbial immobilization techniques, see, for example, "Wastewater Treatment by Microbial Immobilization Method (edited by Ryuichi Sudo, Industrial Water Research Association)" and "Water Treatment by Microbial Immobilization Method - Carrier Immobilization Method, Comprehensive Immobilization Method, Biological Activated Carbon Method (New Water Treatment Series (1)) (authors Kazuhiro Mochizuki, Katsutoshi Hori, Hideki Tachimoto, NTS Corporation)." In one embodiment, surface adhesion proteins possessed by microbial cells may be utilized. In this case, it is also useful to introduce and express the gene for an external adhesion protein into microorganisms that do not possess adhesive properties. The amount of microorganisms immobilized on the carrier can be set appropriately. For example, 0.1% or more, 0.5% or more, 1% or more, 5% or more, 10% or more, 50% or more, or 100% or more of the carrier weight (dry weight) of microorganisms may be immobilized.

[0049] In one embodiment, the carrier may move. The movement of the carrier may increase the frequency of contact between the microorganisms and the gaseous substrate, thereby improving the conversion efficiency of the gaseous substrate. In one embodiment, the movement of the carrier is active, and for example, a movement mechanism is used to move the carrier. The carrier may or may not be connected to the movement mechanism. For example, a movement mechanism not connected to the carrier may move the carrier via electromagnetic force, wind power, etc. Examples of carrier movement include rotational motion and vibrational motion. Power sources for driving the movement mechanism include, but are not limited to, electricity (e.g., supplied from photovoltaic systems such as solar panels, batteries, or general power sources), wind power, etc. In one embodiment, the shape of the rotation mechanism may be rod-shaped (e.g., a rotation axis) or plate-shaped (e.g., a disc). The manner in which the carrier is fixed to the rotation mechanism is arbitrary, but in one embodiment, the carrier may be fixed at an angle (e.g., nearly perpendicular) to the rotation mechanism. In one embodiment, any number of blades, for example, 1 to 100 blades, such as 1, 2, 3, 4, 5, 6, 8, 10, 12, 16, or 20 blades, may be fixed to the rotating shaft. In one embodiment, the rotating mechanism may include at least part of a moisture supply system, and moisture can be supplied to the carrier via the rotating mechanism. For example, moisture can be supplied to the carrier by connecting the rotating mechanism to a liquid reservoir via a water-permeable material component as needed. Generally, when the chamber is spherical, the gas flow inside becomes uniform, and there is no stagnation of flow at corners. Also, when the chamber is spherical, it becomes easier to simulate the gas flow, scale-up becomes easier, and the structural design for axial rotation (compared to, for example, vibrational motion) becomes simpler.

[0050] In one embodiment, the movement of the carrier may result in the movement of water within and / or on the carrier. In one embodiment, rapid rotational or oscillating motion of the carrier may remove weakly bound liquid water from the carrier. In one embodiment, the movement of the carrier may promote the vaporization of water within and / or on the carrier into the gas phase. As liquid water leaves the carrier, fluid-communicated liquid water may be attracted to the carrier and move, so the movement of the carrier may be accompanied by the supply of additional components to the microorganisms. In one embodiment, the movement of the carrier may promote the movement of liquid water within and / or on the carrier. For example, rotational motion may generate centrifugal force, which can move water from the center of the carrier outward, thereby changing the water content of parts of the carrier. In one embodiment, the rotational or vibrational motion of the carrier can be performed at speeds of approximately 1 to 10,000 rpm, approximately 1 to 1,000 rpm, approximately 1 to 100 rpm, approximately 10 to 10,000 rpm, approximately 10 to 1,000 rpm, approximately 10 to 10,000 rpm, or approximately 100 to 1,000 rpm.

[0051] In one embodiment, the products generated by the conversion of a gaseous substrate by the microorganisms of the present disclosure can be recovered. The method for recovering the products is not particularly limited, but for example, the products can be recovered by releasing or volatilizing them into the gas phase, dissolving them in liquid water, or adsorbing them onto a solid phase surface.

[0052] (portion) In one embodiment, the conversion of a gaseous substrate by the microorganisms of the Disclosure is carried out using a gas-phase microbial reaction system, which may have one or more parts. Therefore, these one or more parts are also within the scope of the Disclosure. Any part of the present disclosure for carrying out the conversion of a gaseous substrate by microorganisms is intended, but examples of such parts include a carrier on which microorganisms are immobilized or for immobilizing microorganisms, a gas-phase reaction chamber or components for constructing such a chamber, a support for immobilizing the carrier (also called a carrier holder) (for example, inside or on the wall of the gas-phase reaction chamber such that at least a portion of the immobilized microorganisms are exposed to the gas phase), a kinetic mechanism for moving the carrier, a power source for moving the kinetic mechanism, a gas supply unit for supplying the gaseous substrate to the gas phase (including a liquid reservoir that produces the gaseous substrate by vaporization, and a solid that produces the gaseous substrate by vaporization or sublimation), a water supply system for supplying water to the carrier (including a liquid reservoir), a chamber for containing the liquid phase (aqueous phase chamber), a gas discharge unit for removing gas from the gas-phase reaction chamber, a liquid discharge unit for removing liquid from the gas-phase reaction chamber, and a mechanism for recovering products resulting from the conversion of the gaseous substrate by microorganisms. In this specification, a system comprising parts having multiple functions may be described, such a system may comprise multiple parts, each having a different function, or one part may have multiple functions. For example, a system including a gas supply unit and a water supply system may include a part that supplies a gaseous substrate and water (e.g., (9) in Figure 7(B)). Alternatively, one or more functions may be performed by combining multiple parts (components). Multiple parts may be supplied separately, integrally molded (e.g., 3D printing or injection modeling), or assembled. In one embodiment, one or more of the liquid discharge unit, gas supply unit, and gas discharge unit are connected to the gas-phase reaction chamber. A kit containing one or more parts constituting the gas-phase microbial reaction system of the Disclosure is also contemplated in this Disclosure. In addition to the parts, the kit may include instructions that describe how to use the parts and / or how to operate the gas-phase microbial reaction system.

[0053] In one embodiment, the gas-phase microbial reaction system and / or a part thereof of the Disclosure may be controlled by a program. The program may be configured to implement on a computer the method of converting a gaseous substrate by microorganisms of the Disclosure. In one embodiment, the program may implement on a computer a method of performing or controlling at least one of the steps of supplying a gaseous substrate to the gas phase, supplying water to a carrier, and moving the carrier. The Disclosure also provides a recording medium storing the program of the Disclosure. The gas-phase microbial reaction system of the Disclosure may include such a recording medium.

[0054] (Operating environment) In this disclosure, any gaseous substrate that can be converted by microorganisms may be used, but in one embodiment, gaseous substrates used in the microbial conversion of this disclosure include methane, ethane, ethylene, acetylene, propane, propylene, formaldehyde, acetaldehyde, ammonia, carbon dioxide, nitrogen oxides, hydrogen sulfide, sulfur oxides, terpenes, methanol, ethanol, toluene, naphthalene, camphor, and paradichlorobenzene. In one embodiment, the conversion of this disclosure is aimed at removing harmful substances such as malodorous substances, allergens, and caustic substances. In one embodiment, the conversion of this disclosure is aimed at producing a desired product. Those skilled in the art can appropriately select microorganisms depending on the type of gaseous substrate.

[0055] In one embodiment, the conversion of the gaseous substrate according to this disclosure may produce a desired product, which may include, but is not limited to, alcohols (such as methanol), carboxylic acids, aldehydes, ketones, terpenes, and geranic acid. In one embodiment, an aromatic component may be produced.

[0056] In this disclosure, any microorganism capable of converting a gaseous substrate may be used. Those skilled in the art can appropriately select a microorganism based on the type of gaseous substrate and, if necessary, the type of desired product. In addition to bacteria, other microorganisms such as yeasts, microalgae, and mosses can also be selected. An appropriate water content can be selected depending on the microorganism used. Furthermore, in addition to wild-type strains, genetically modified microorganisms, molecularly bred strains created by synthetic biology techniques, and genome-edited microorganisms can also be used.

[0057] In one embodiment, the conversion of a gaseous substrate by the microorganisms of the Disclosure is carried out by maintaining the microorganisms in any temperature environment such as 0-100°C, 5-70°C, 10-50°C, 15-40°C, 20-35°C, less than 70°C, less than 60°C, less than 50°C, less than 40°C, less than 30°C, less than 25°C, less than 20°C, less than 15°C, less than 10°C, less than 5°C, less than 0°C, about 70°C, about 60°C, about 50°C, about 40°C, about 30°C, about 25°C, about 15°C, about 10°C, about 5°C, or about 0°C.

[0058] In one embodiment, the conversion of a gaseous substrate by the microorganisms of this disclosure may be carried out in conjunction with any operation that provides assistance for the survival of the microorganisms used and / or improves the conversion efficiency, such as light irradiation or voltage application.

[0059] (General technology) The molecular biological, biochemical, and microbiological methods used herein are well-known and commonly used in the field.

[0060] (Note) In this specification, "or" is used when "at least one" of the items listed in the text can be adopted. The same applies to "or else". In this specification, when it is specified that "within the range" of "two values", that range includes the two values ​​themselves.

[0061] The present disclosure will be described below based on examples, but the above description and the following examples are provided for illustrative purposes only and not to limit the present invention. Accordingly, the scope of the present invention is not limited to the embodiments or examples specifically described herein, but is limited only by the claims. [Examples]

[0062] Examples are described below. Where necessary, the handling of organisms used in the following examples complied with the standards stipulated by Nagoya University, the supervisory authority, and the Cartagena Protocol. Specifically, the reagents used were those described in the examples, but equivalent products from other manufacturers (Sigma-Aldrich, Fujifilm, Wako Pure Chemical Industries, Nakai, R&D Systems, USCN Life Science INC, Kanto Chemical, Funakoshi, Tokyo Chemical Industries, Merck, etc.) can be substituted.

[0063] A carrier on which methane-utilizing bacteria (Methylococcus capsulatus (Bath)) were immobilized was prepared using the following procedure.

[0064] A glass fiber filter (GF / F 47, GE Healthcare) was placed in a filter holder, and a freeze-thawed microbial suspension was passed through the filter using a syringe. Washing solution was flowed in both forward and reverse directions to remove weakly bound cells. All of the liquid passed through the filter was collected, and the absorbance of the collected solution was measured to calculate the amount of cells contained in the collected solution. The weight of cells fixed to the filter (dry weight) was calculated by subtracting the amount of cells in the collected solution from the amount of cells in the cell suspension.

[0065] The methane concentration in the recovered gas and the methanol concentration in the liquid phase were quantified using gas chromatography (GC 2014, Shimadzu Corporation) equipped with a flame ionization detector and a 25% sorbitol gas port B (60 / 80) glass column (GL Science). Gas chromatography was performed under the following conditions. Carrier gas: Nitrogen Flow rate: 40mL / min Column temperature: 100℃ Detector temperature: 150℃

[0066] (Example 1) The bioreactor (gas-phase microbial reaction system) shown in Figures 1 and 2 was assembled. A mixed gas of methane and air was flowed through a single gas supply line. The bioreactor was configured so that the flow path from the gas supply section to the gas discharge section was long. The operation was carried out under atmospheric pressure. The bioreactor was configured so that the liquid phase circulated at a flow rate of 30 mL / min. The initial volume of the liquid phase was approximately 10 mL. A methane-utilizing bacteria immobilization filter was placed between the gas phase and the liquid phase, with the side containing the immobilized bacteria facing the gas phase.

[0067] A bioreactor equipped with a support structure immobilized with Methylococcus capsulatus (Bath) was supplied with a methane-air mixture (20% methane content), and methane decomposition experiments were conducted at 42°C. The methane concentration in the exhaust gas was measured under various cell densities (dry cell weight per unit volume of gas), gas flow rates, and methane concentrations in the supplied gas, and the methane decomposition rate was calculated from its change over time.

[0068] The results are shown in Figure 3. Efficient and continuous methane decomposition using a bioreactor was achieved.

[0069] Next, methane decomposition and methane-to-methanol conversion experiments were conducted when air containing 20% ​​methane was supplied at a flow rate of 0.1 mL / min. For the first 6 hours, only methane decomposition was performed by supplying only buffer solution or a culture medium containing formic acid as a reducing agent. After 6 hours, 1 μM of cyclopropanol, a methanol dehydrogenase inhibitor, was added to produce methanol from methane. However, at 9 hours, the culture medium was replaced with fresh medium, and the concentration of cyclopropanol was further increased to 10 μM.

[0070] The results are shown in Figure 4. Stable methane decomposition and efficient continuous methanol production were achieved using a bioreactor.

[0071] Furthermore, using this bioreactor equipped with a support structure immobilized with Methylococcus capsulatus (Bath), the decomposition of methane in the aqueous phase chamber was observed both in the presence and absence of the liquid phase. Air containing 22.8% (v / v) methane was supplied at 42°C, and the methane concentration at the gas supply and discharge sections was measured under various conditions, including the presence or absence of the liquid phase and different gas flow rates, and the methane decomposition rate was calculated.

[0072] The results are shown in Figure 5. The methane concentration at the gas discharge point remained constant at 14.6% in the absence of the liquid phase and at 19.4% in the presence of the liquid phase. The methane consumption rate in the absence of the liquid phase was approximately 2.5 times higher than in the presence of the liquid phase, indicating that the water content of the carrier may be important for the efficient conversion of the gaseous substrate. Furthermore, under conditions in the absence of the liquid phase, the methane decomposition rate improved with increasing gas flow velocity. This indicates that gas flow velocity can affect the water content on the carrier surface and thus the methane decomposition rate.

[0073] (Example 2) The bioreactor (gas-phase microbial reaction system) shown in Figures 6-9 was assembled. Details are as described in Figure 7.

[0074] In the following preliminary experiment, neither water nor culture medium was added to the chip. First, 2 mL of 12.9 mM phosphate buffer (1.83 g Na2HPO4 and 1.76 g KH2PO4 in 1 L of distilled water, pH 7) was added, and moisture was transferred to the filter by capillary action through tissue paper. No water was added during operation. The decomposition test was started with the filter initially moistened. The moisture content of the 10-blade fan was 53% (v / v), and the moisture content of the 20-blade fan was 58%. The bioreactor has a volume of approximately 150 mL. A filter with Methylococcus capsulatus (Bath) fixed to it was used for the fan blades. In this case, water was not added to the liquid reservoir (16) in Figure 7(C), and since moisture was not supplied to the fan blades via (15) in Figure 7(C) during operation, the moisture content of the fan decreased to less than 20% at the end of operation.

[0075] The decomposition of methane at 42°C was observed using a bioreactor with a 10-blade or 20-blade fan for continuous supply. The fan was rotated at 100 rpm. The methane concentration in the exhaust gas was measured under various methane supply conditions (mL / min), and the methane decomposition rate was calculated.

[0076] The results are shown in Figure 10. Efficient continuous methane decomposition was achieved in both bioreactors. In particular, the methane decomposition rate was approximately three times faster when using a 20-blade fan compared to when using a 10-blade fan.

[0077] Similarly, the methane concentration at the gas outlet was measured when air containing 20% ​​(v / v) methane was supplied at a flow rate of 8 mL / min.

[0078] The results are shown in Figure 11. Stable, continuous methane decomposition was achieved in both bioreactors.

[0079] (Example 3) Assemble a bioreactor (gas-phase microbial reaction system) comprising the parts shown in Figures 12 and 13. Efficient and / or stable conversion of a gaseous substrate by microorganisms is achieved.

[0080] (Example 4) The Acinetobacter strain Tol5, genetically edited to control gene expression in the toluene degradation pathway, was applied to the bioreactor described in Figures 1 and 2 to convert gaseous toluene to methylcatechol. Stable toluene consumption and methylcatechol production were achieved.

[0081] (Example 5) A polyurethane carrier immobilized with a molecularly bred strain of Acinetobacter bacterium Tol5, which has been conferred geranic acid production capacity using synthetic biological methods, is packed into a gas-phase reaction chamber. A mixture of gaseous geraniol and air is continuously supplied from outside the chamber to carry out the conversion reaction to geranic acid. The discharge of a gas containing a stable concentration of geraniol from the outlet of the gas-phase reaction chamber is confirmed, and the adsorption and recovery of the produced geranic acid onto the polyurethane carrier is achieved.

[0082] (Example 6) The bioreactor (gas-phase microbial reaction system) shown in Figure 15 was assembled. A mixture of methane and air was flowed through a single gas supply line. Unlike the bioreactor in Example 1, the flow path is not vortex-shaped. Similar to Example 1, a methane-assimilating bacterial fixation filter was placed between the gas phase and the liquid phase (bacterial fixation amount: 9.95 g (dry weight) / m³). 2The device was positioned so that the side with the fixed bacteria was facing the gas phase (Figure 16), or so that the side with the fixed bacteria was facing the liquid phase (Figure 17), and a methane-to-methanol conversion experiment was performed by supplying a methane-air mixed gas (methane content 20%). The gas flow rate was 3 mL / min, the medium circulation flow rate was 10 mL / min, and the medium was NMS medium containing 10 μM cyclopropanol and 10 mM sodium formate (KNO3 1.0 g / L; MgSO4·7H2O 1.0 g / L; CaCl2·2H2O 0.2 g / L; Na2HPO4 0.497 g / L; KH2PO4 0.39 g / L; Fe-EDTA 0.019 g / L; Na2MoO4·2H2O 0.5 mg / L; FeSO4·7H2O 0.5 mg / L; ZnSO4·7H2O 0.4 mg / L; Na2EDTA 0.25 mg / L; H3BO3 0.015 mg / L; CoCl2·6H2O 0.05 mg / L; MnCl2·4H2O 0.02 mg / L; NiCl2·6H2O (0.01 mg / L) was used. Note that in the conventional membrane reactor configuration, as shown in Figure 17, where the methane-utilizing bacterial immobilization filter is positioned so that the side with the immobilized bacteria faces the liquid phase, the microorganisms that were initially immobilized in the aqueous phase detached and became suspended.

[0083] Figure 18 shows the results of arranging the filter so that the side with the fixed bacteria faces either the gas phase (A) or the liquid phase (B). It was confirmed that using bacteria in the gas phase achieves more efficient methanol conversion compared to the conventional configuration of using bacteria in the liquid phase.

[0084] Similarly, a methane-utilizing bacterial immobilization filter was placed between the gas phase and the liquid phase, with the side containing the immobilized bacteria facing the gas phase (Figure 19), and a methane-to-methanol conversion experiment was conducted. In the above experiment (Figure 18), the liquid culture medium was circulated by a Perister pump, and no new medium was supplied, so the liquid portion was equivalent to a batch operation. On the other hand, in this experiment (Figure 19), the culture medium was circulated at the same flow rate (10 mL / min) as in the above experiment (Figure 18), while fresh medium was continuously supplied at 4 mL / hour, and the methanol-containing discharge liquid was continuously collected at the same rate as the medium supply rate (4 mL / hour). The gas flow rate was 0.2 mL / min.

[0085] The results are shown in Figure 20. Stable and efficient continuous methane-to-methanol conversion was achieved using a bioreactor.

[0086] (Example 7) The decomposition of toluene was tested in a fan-type bioreactor. The experiment was carried out according to the following procedure. A schematic of the system used in this example is shown in Figure 21.

[0087] Preparation of toluene samples 1 mL of toluene was pipetteed into a 125 mL flask. The flask was then sealed by attaching rubber plugs to both ports. The flask was then left to stand at room temperature for 1 hour before being used as a sample introduced into the bioreactor inlet. Toluene vapor had a saturated vapor pressure at 25°C.

[0088] Tol5 culture and immobilization Wild-type cells of Acinetobacter Tol5 (S. Ishii, J. Koki, H. Unno, K. Hori; Two Morphological Types of Cell Appendages on a Strongly Adhesive Bacterium, Acinetobacter sp. Strain Tol5, Appl. Environ. Microbiol. 70, (2004) 5026-5029) were cultured in 150 mL of LB medium for 24 hours. All cells were harvested and washed three times with deionized water by centrifugation at 6000 rpm for 10 minutes. The cell pellet was resuspended in 10 mL of deionized water and added dropwise to a fan unit using a pipette. 140 mg of Tol5 cells (dry weight) were harvested, and the cell concentration on the filter surface was 3 g (dry weight) / m². 2 That was the case.

[0089] Construction of a fan-type bioreactor A 150 mL gas chamber was assembled by screwing together two components: a lower cover with a gas inlet connector and an upper cover with a gas outlet connector. A fan-type bioreactor was assembled by setting a fan unit inside the gas chamber. To rotate the fan unit, a rotor was attached to the upper cover of the fan-type bioreactor and powered by a battery. PTFE gaskets were used to connect each component to prevent gas leaks. A syringe plunger was attached to the rotor's rotation axis and assembled with the fan unit (the central axis being a syringe barrel). The fan unit consisted of 20 filter paper blades.

[0090] Preparation of a fan-type bioreactor After immobilizing the cells onto the fan unit, the fan unit was assembled onto the rotor shaft. The gas chamber was sealed, and 5 mL of deionized water was injected into the bottom of the bioreactor to maintain saturation humidity within the system. A bioreactor without immobilized cells onto the fan unit was also prepared as a control.

[0091] Toluene vapor injection and bioreactor operation A tube for injecting pure air from an air cylinder was connected to one port of a flask containing toluene vapor, and the other port was connected to the inlet port of a fan-type bioreactor. The air flow rate was controlled by a gas blender. Air was injected into the flask at a flow rate of 10 mL / min and mixed with the toluene vapor, which was then injected into the bioreactor as the inlet gas sample. The toluene concentration in the inlet gas stream was 32.2 μM. The retention time of the toluene vapor in the bioreactor was 12.5 minutes. By rotating the fan at 25 rpm using a rotor, the toluene vapor was efficiently brought into contact with and reacted with microorganisms immobilized on the fan. This bioreactor operation was performed at room temperature.

[0092] Gas sample acquisition and quantification Gas samples (5 μL) were collected from the bioreactor inlet and outlet and injected into gas chromatography-mass spectrometry (GC / MS) using airtight syringes. Toluene concentration was then quantified using a GC / MS system including an MS detector (MSD 5975; Agilent Technologies) connected to a GC system (GC7820A; Agilent Technologies, Santa Clara, California) fitted with an Rtx-200 capillary column (30 m × 0.32 mm × 0.5 μm; RESTEC, Belfonte, Pennsylvania). The helium split ratio and flow rate were set to 10:1 and 2 mL / min, respectively. The operation program began with an isocratic step of 90°C for 1 minute, followed by a temperature increase of 25°C / min to a final temperature of 120°C, with ion fragments at m / z=65 and m / z=91 monitored in selective ion monitoring mode. Peaks indicating toluene were detected with a retention time of 2.1 minutes. The peak area for calculating toluene concentration in the gas phase was measured manually.

[0093] The toluene removal rate was determined by comparing the toluene concentration at the outlet of the control experiment (bioreactor without immobilized cells) and the test experiment (bioreactor with immobilized cells).

[0094] The results are shown in Figure 22. When toluene vapor was supplied to a fan-type bioreactor for a holding time of 12.5 minutes, approximately 37% of the toluene was continuously removed by Acinetobacter Tol5 cells, confirming the excellent performance of the bioreactor.

[0095] (Note) As described above, while the present disclosure has been illustrated using preferred embodiments thereof, it is understood that the scope of the present invention should be interpreted solely by the claims. Patents, patent applications and other documents cited herein should be incorporated herein by reference as if their contents were specifically described herein.

[0096] This application claims priority to Japanese Patent Application No. 2020-71961, filed with the Japan Patent Office on April 13, 2020, the entire contents of which are incorporated herein by reference. [Industrial applicability]

[0097] This disclosure provides an efficient and / or stable conversion of gaseous substrates using microorganisms, thereby facilitating the use of microorganisms for the removal of unwanted gases and / or the acquisition of desired products. Furthermore, since the conversion of gaseous substrates according to this disclosure may be efficient and / or stable, it may also contribute to the miniaturization of gas-phase microbial reaction systems.

Claims

1. A gas-phase reaction chamber equipped with a gas supply unit that continuously or intermittently supplies a gaseous substrate from the outside, A carrier installed in the gas-phase reaction chamber or a carrier constituting at least a part of the wall of the gas-phase reaction chamber, wherein the carrier is permeable to water or absorbent but its gas-phase side surface is not immersed in water, and at least a portion of the microbial cells are fixed on the surface of the carrier that is exposed to the gas phase so that at least a portion of the microbial cells are in direct contact with the gas phase in the gas-phase reaction chamber, A gas discharge unit connected to the gas-phase reaction chamber, A gas-phase microbial reaction system comprising, (A) A water supply system further comprising any of the following features (1) to (3): (1) (i) The gas-phase microbial reaction system comprises a plurality of gas phases and a plurality of liquid phases, configured such that the gas phases and the liquid phases are alternately layered, and / or (ii) The water supply system includes an aqueous phase chamber, the carrier includes a plurality of tubular members, the plurality of tubular members are bundled together in close contact to form a columnar unit, or the plurality of tubular members are arranged at intervals from each other in a sheath tube to form a columnar unit including the plurality of tubular members and the sheath tube, the plurality of tubular members and the sheath tube constitute the wall of the gas phase reaction chamber or the aqueous phase chamber. (2) The moisture supply system is configured to supply the moisture to the carrier in an aerosol or gaseous state, or (3) The water supply system is configured such that the water in a gaseous state condenses on the carrier, or (B) Further comprising a rotating mechanism connected to the carrier, the rotating mechanism comprising a rotating shaft to which the carrier is fixed, Gas-phase microbial reaction system.

2. The gas-phase microbial reaction system according to claim 1, wherein the moisture supply system is configured to supply moisture from outside the gas-phase reaction chamber to an extent that prevents the microbial cells fixed to the carrier from drying out.

3. The gas-phase microbial reaction system according to claim 2, wherein the moisture supply system is configured such that the moisture content of the carrier is 1 to 50% (v / v).

4. The gas-phase microbial reaction system according to any one of claims 1 to 3, further comprising a mechanism for recovering products resulting from the conversion of the gaseous substrate by the microorganisms.

5. The gas-phase microbial reaction system according to any one of claims 1 to 4, wherein the gas supply unit is configured to passively take in the gaseous substrate by diffusion from outside the gas-phase reaction chamber.

6. The gas-phase microbial reaction system according to any one of claims 1 to 5, wherein the carrier is a thin film.

7. The gas-phase microbial reaction system according to any one of claims 1 to 6, further comprising a liquid discharge unit connected to the gas-phase reaction chamber.

8. The gas-phase microbial reaction system according to any one of claims 1 to 7, wherein the moisture supply system is configured to supply the moisture to the carrier as a liquid stream.

9. The gas-phase microbial reaction system according to claim 8, wherein the water supply system includes an aqueous phase chamber through which a liquid flows, the carrier is disposed between the aqueous phase chamber and the gas-phase reaction chamber, and the carrier is positioned such that the surface of the carrier on which the microorganisms are immobilized faces the gas-phase reaction chamber.

10. The gas-phase microbial reaction system according to any one of claims 1 to 9, wherein the gas-phase microbial reaction system comprises a plurality of gas phases and a plurality of liquid phases, and is configured such that the gas phases and the liquid phases are alternately stacked.

11. The gas-phase microbial reaction system according to any one of claims 1 to 10, wherein the carrier includes a plurality of tubular members, the plurality of tubular members are bundled together in close contact to form a columnar unit, or the plurality of tubular members are arranged at intervals from each other within a sheath tube to form a columnar unit including the plurality of tubular members and the sheath tube, and the plurality of tubular members and the sheath tube constitute the wall of the gas-phase reaction chamber or the aqueous-phase chamber.

12. The gas-phase microbial reaction system according to any one of claims 1 to 11, further comprising a rotating mechanism connected to the carrier, wherein the rotating mechanism comprises a rotating shaft to which the carrier is fixed.

13. The gas-phase microbial reaction system according to any one of claims 1 to 12, wherein the moisture supply system is configured to supply the moisture to the carrier by capillary action.

14. The gas-phase microbial reaction system according to any one of claims 1 to 12, wherein the moisture supply system is configured to supply the moisture to the carrier in an aerosol or gaseous state.

15. The gas-phase microbial reaction system according to any one of claims 1 to 12, wherein the moisture supply system is configured such that the gaseous moisture condenses on the carrier.

16. The gas-phase microbial reaction system according to any one of claims 1 to 15, further comprising a power source for driving the rotating mechanism.

17. A method for converting a gaseous substrate using microorganisms, The steps include supplying a gaseous substrate to the gas-phase reaction chamber from the outside, either continuously or intermittently, The steps include: exposing at least a portion of the surface on which the microorganisms on the carrier are immobilized to the gas phase in the gas-phase reaction chamber; Includes, Here, at least a portion of the microorganisms are immobilized on a surface of the carrier that is exposed to the gas phase. (A) The method further comprises the step of supplying water to the microbial carrier from outside the gas-phase reaction chamber using a water supply system having any of the following features (1) to (3): (1) (i) The gas-phase reaction chamber and the moisture supply system are configured such that the gas phase and liquid phase are alternately layered, and / or (ii) The water supply system includes an aqueous phase chamber, the carrier includes a plurality of tubular members, the plurality of tubular members are bundled together in close contact to form a columnar unit, or the plurality of tubular members are arranged at intervals from each other in a sheath tube to form a columnar unit including the plurality of tubular members and the sheath tube, the plurality of tubular members and the sheath tube constitute the wall of the gas phase reaction chamber or the aqueous phase chamber. (2) The moisture supply system is configured to supply the moisture to the carrier in an aerosol or gaseous state, or (3) The water supply system is configured such that the water in a gaseous state condenses on the carrier, or (B) The method further includes the step of rotating the carrier by a rotating mechanism connected to the carrier, wherein the rotating mechanism comprises a rotating shaft on which the carrier is fixed. method.

18. The method according to claim 17, further comprising the step of supplying moisture to a microbial support from outside the gas-phase reaction chamber.

19. The method according to claim 17 or 18, wherein the moisture content of the carrier is controlled to 1 to 50%.

20. A carrier for immobilizing microorganisms, or a carrier on which microorganisms are immobilized, A gas-phase reaction chamber, or a gas-phase reaction chamber member configured to provide the gas-phase reaction chamber when assembled, A gas supply unit that continuously or intermittently supplies a gaseous substrate from the outside, Gas discharge section and A kit that includes, The gas-phase reaction chamber may be provided comprising at least one of the carrier, the gas supply unit, and the gas discharge unit, or it may be provided separately from the carrier, the gas supply unit, and the gas discharge unit. The gas-phase reaction chamber is configured to house the carrier, The gas-phase reaction chamber is configured such that the microorganisms immobilized on the carrier come into contact with a gaseous substrate, and at least a portion of the microorganisms are immobilized on a surface of the carrier that is exposed to the gas phase. (A) The kit further includes a water supply system for supplying water to the carrier from outside the gas-phase reaction chamber having any of the following features (1) to (3): (1) (i) The gas-phase reaction chamber and the moisture supply system are configured such that the gas phase and liquid phase are alternately stacked, and / or (ii) The water supply system includes an aqueous phase chamber, the carrier includes a plurality of tubular members, the plurality of tubular members are bundled together in close contact to form a columnar unit, or the plurality of tubular members are arranged at intervals from each other in a sheath tube to form a columnar unit including the plurality of tubular members and the sheath tube, the plurality of tubular members and the sheath tube constitute the wall of the gas phase reaction chamber or the aqueous phase chamber. (2) The moisture supply system is configured to supply the moisture to the carrier in an aerosol or gaseous state, or (3) The water supply system is configured such that the water in a gaseous state condenses on the carrier, or (B) The kit further comprises a rotating mechanism configured to be connected to the carrier, the rotating mechanism comprising a rotating shaft to which the carrier is fixed. kit.

21. The kit according to claim 20, further comprising a moisture supply system for supplying moisture to the carrier from outside the gas-phase reaction chamber.

22. The kit according to claim 20 or 21, further comprising a support configured to support the carrier.

23. The gas-phase microbial reaction system according to any one of claims 1 to 16, wherein the microbial cells express adhesion proteins.

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